tretinoin and Vitamin-A-Deficiency

Structural birth defects are a common cause of abnormalities in newborns. While there are cases of structural birth defects arising due to monogenic defects or environmental exposures, many birth defects are likely caused by a complex interaction between genes and the environment. A structural birth defect with complex etiology is congenital diaphragmatic hernias (CDH), a common and often lethal disruption in diaphragm development. Mutations in more than 150 genes have been implicated in CDH pathogenesis. Although there is generally less evidence for a role for environmental factors in the etiology of CDH, deficiencies in maternal vitamin A and its derivative embryonic retinoic acid are strongly associated with CDH. However, the incomplete penetrance of CDH-implicated genes and environmental factors such as vitamin A deficiency suggest that interactions between genes and environment may be necessary to cause CDH. In this review, we examine the genetic and environmental factors implicated in diaphragm and CDH development. In addition, we evaluate the potential for gene-environment interactions in CDH etiology, focusing on the potential interactions between the CDH-implicated gene, Gata4, and maternal vitamin A deficiency.

Topics: Diaphragm; Hernias, Diaphragmatic, Congenital; Humans; Infant, Newborn; Mutation; Tretinoin; Vitamin A Deficiency

Childhood obesity has become a public health concern. As the importance of vitamin A (VA) in the body has become increasingly acknowledged, there is limited clinical trial evidence to substantiate the association between VA and childhood obesity. Vitamin A deficiency (VAD) increases the risk of childhood obesity, a finding consistently reported in pregnant women. VA could regulate the adipogenic process, inflammation, oxidative stress and metabolism-related gene expression in mature adipocytes. VAD disrupts the balance of obesity-related metabolism, thus affecting lipid metabolism and insulin regulation. Conversely, VA supplementation has a major impact on efficacy in obesity, and obese individuals typically have a lower VA status than normal-weight individuals. Several studies have attempted to identify the genetic and molecular mechanisms underlying the association between VA and obesity. In this review, we summarize and discuss recent new developments focusing on retinol, retinoic acid, and RBP4 and elucidate and provide an overview of the complex interrelationships between these critical components of VA and childhood obesity. However, the causal relationship between VA status and childhood obesity remains unclear. It is also unknown whether VA supplementation improves the overall obesogenic metabolic profile.

Topics: Child; Female; Humans; Insulin; Pediatric Obesity; Pregnancy; Retinol-Binding Proteins, Plasma; Tretinoin; Vitamin A; Vitamin A Deficiency

Vitamin A is a fat-soluble vitamin that plays an important role in skin immunity. Deficiencies in Vitamin A have been linked to impaired immune response and increased susceptibility to skin infections and inflammatory skin disease. This narrative review summarizes recent primary evidence that elucidates the role of vitamin A and its derivatives on innate immune regulators through mechanisms that promote skin immunity and sustain the skin microbiome.

Topics: Animals; Dermatitis; Humans; Immunity, Innate; Microbiota; Skin; Staphylococcus aureus; Tretinoin; Vitamin A; Vitamin A Deficiency

Vitamin A deficiency can cause human pathologies that range from blindness to embryonic malformations. This diversity is due to the lack of two major vitamin A metabolites with very different functions: the chromophore 11-cis-retinal (vitamin A aldehyde) is a critical component of the visual pigment that mediates phototransduction, while the signaling molecule all-trans-retinoic acid regulates the development of various tissues and is required for the function of the immune system. Since animals cannot synthesize vitamin A de novo, they must obtain it either as preformed vitamin A from animal products or as carotenoid precursors from plant sources. Due to its essential role in the visual system, acute vitamin A deprivation impairs photoreceptor function and causes night blindness (poor vision under dim light conditions), while chronic deprivation results in retinal dystrophies and photoreceptor cell death. Chronic vitamin A deficiency is the leading cause of preventable childhood blindness according to the World Health Organization. Due to the requirement of vitamin A for retinoic acid signaling in development and in the immune system, vitamin A deficiency also causes increased mortality in children and pregnant women in developing countries. Drosophila melanogaster is an excellent model to study the effects of vitamin A deprivation on the eye because vitamin A is not essential for Drosophila development and chronic deficiency does not cause lethality. Moreover, genetic screens in Drosophila have identified evolutionarily conserved factors that mediate the production of vitamin A and its cellular uptake. Here, we review our current knowledge about the role of vitamin A in the visual system of mammals and Drosophila melanogaster. We compare the molecular mechanisms that mediate the uptake of dietary vitamin A precursors and the metabolism of vitamin A, as well as the consequences of vitamin A deficiency for the structure and function of the eye.

Topics: Animals; Drosophila melanogaster; Mammals; Photoreceptor Cells; Retina; Retinal Pigment Epithelium; Retinaldehyde; Tretinoin; Vision, Ocular; Visual Perception; Vitamin A; Vitamin A Deficiency

Several models have been proposed to explain the neurodevelopmental syndrome induced by exposure of human embryos to alcohol, which is known as fetal alcohol spectrum disorder (FASD). One of the proposed models suggests a competition for the enzymes required for the biosynthesis of retinoic acid. The outcome of such competition is development under conditions of reduced retinoic acid signaling. Retinoic acid is one of the biologically active metabolites of vitamin A (retinol), and regulates numerous embryonic and differentiation processes. The developmental malformations characteristic of FASD resemble those observed in vitamin A deficiency syndrome as well as from inhibition of retinoic acid biosynthesis or signaling in experimental models. There is extensive biochemical and enzymatic overlap between ethanol clearance and retinoic acid biosynthesis. Several lines of evidence suggest that in the embryo, the competition takes place between acetaldehyde and retinaldehyde for the aldehyde dehydrogenase activity available. In adults, this competition also extends to the alcohol dehydrogenase activity. Ethanol-induced developmental defects can be ameliorated by increasing the levels of retinol, retinaldehyde, or retinaldehyde dehydrogenase. Acetaldehyde inhibits the production of retinoic acid by retinaldehyde dehydrogenase, further supporting the competition model. All of the evidence supports the reduction of retinoic acid signaling as the etiological trigger in the induction of FASD.

Topics: Animals; Embryo, Mammalian; Ethanol; Fetal Alcohol Spectrum Disorders; Humans; Models, Biological; Syndrome; Tretinoin; Vitamin A Deficiency

Vitamin A is an essential micronutrient throughout life. Its physiologically active metabolite retinoic acid (RA), acting through nuclear retinoic acid receptors (RARs), is a potent regulator of patterning during embryonic development, as well as being necessary for adult tissue homeostasis. Vitamin A deficiency during pregnancy increases risk of maternal night blindness and anemia and may be a cause of congenital malformations. Childhood Vitamin A deficiency can cause xerophthalmia, lower resistance to infection and increased risk of mortality. RA signaling appears to be essential for expression of genes involved in developmental hematopoiesis, regulating the endothelial/blood cells balance in the yolk sac, promoting the hemogenic program in the aorta-gonad-mesonephros area and stimulating eryrthropoiesis in fetal liver by activating the expression of erythropoietin. In adults, RA signaling regulates differentiation of granulocytes and enhances erythropoiesis. Vitamin A may facilitate iron absorption and metabolism to prevent anemia and plays a key role in mucosal immune responses, modulating the function of regulatory T cells. Furthermore, defective RA/RARα signaling is involved in the pathogenesis of acute promyelocytic leukemia due to a failure in differentiation of promyelocytes. This review focuses on the different roles played by vitamin A/RA signaling in physiological and pathological mouse hematopoiesis duddurring both, embryonic and adult life, and the consequences of vitamin A deficiency for the blood system.

Topics: Anemia, Iron-Deficiency; Animals; Cell Differentiation; Disease Models, Animal; Embryonic Development; Epigenesis, Genetic; Erythropoiesis; Erythropoietin; Female; Granulocytes; Hematopoiesis; Humans; Leukemia, Promyelocytic, Acute; Pregnancy; Receptors, Retinoic Acid; Signal Transduction; Tretinoin; Vitamin A; Vitamin A Deficiency

Vitamin A, a generic designation for an array of organic molecules that includes retinal, retinol and retinoic acid, is an essential nutrient needed in a wide array of aspects including the proper functioning of the visual system, maintenance of cell function and differentiation, epithelial surface integrity, erythrocyte production, reproduction, and normal immune function. Vitamin A deficiency is one of the most common micronutrient deficiencies worldwide and is associated with defects in adaptive immunity. Reports from epidemiological studies, clinical trials and experimental studies have clearly demonstrated that vitamin A plays a central role in immunity and that its deficiency is the cause of broad immune alterations including decreased humoral and cellular responses, inadequate immune regulation, weak response to vaccines and poor lymphoid organ development. In this review, we will examine the role of vitamin A in immunity and focus on several aspects of T cell biology such as T helper cell differentiation, function and homing, as well as lymphoid organ development. Further, we will provide an overview of the effects of vitamin A deficiency in the adaptive immune responses and how retinoic acid, through its effect on T cells can fine-tune the balance between tolerance and immunity.

Topics: Adaptive Immunity; Animals; Cell Differentiation; Clinical Trials as Topic; Dietary Supplements; Disease Models, Animal; Epithelial Cells; Humans; Immune Tolerance; Immunity, Cellular; Lymphocytes; Organogenesis; T-Lymphocytes; Th1 Cells; Th17 Cells; Th2 Cells; Thymus Gland; Tretinoin; Vitamin A Deficiency

The importance of vitamin A for host defense is undeniable and the study of its mechanisms is paramount. Of the estimated 250 million preschool children who are vitamin A-deficient (VAD), 10% will die from their increased susceptibility to infectious disease. Vitamin A supplementation was established in the 1980s as one of the most successful interventions in the developing world. Understanding how vitamin A controls immunity will help curb the mortality and morbidity associated with vitamin A deficiency and exploit the immune-enhancing capacity of vitamin A to heighten host resistance to infectious disease. The discoveries that retinoic acid (RA) imprints the homing of leukocytes to the gut and enhances the induction of regulatory T cells, highlighted a potential role for RA in mucosal tolerance. However, more recently emerging data tell of a more profound systemic impact of RA on leukocyte function and commitment. In animal models using genetic manipulation of RA signaling, we learned when and how RA controls T cell fate. Here, we review the role for RA as a critical checkpoint regulator in the differentiation of CD4(+) T cells within the immune system.

Topics: Animals; Cell Differentiation; Forkhead Transcription Factors; Humans; Immunity, Mucosal; Immunosuppressive Agents; Immunotherapy; Mice; Models, Immunological; Retinoids; Signal Transduction; T-Lymphocytes, Helper-Inducer; Tretinoin; Vitamin A; Vitamin A Deficiency

Vitamin A and its derivatives (retinoids) are critically important in the development and maintenance of multiple epithelial tissues, including skin, hair, and sebaceous glands, as shown by the detrimental effects of either vitamin A deficiency or toxicity. Thus, precise levels of retinoic acid (RA, active metabolite) are needed. These precise levels of RA are achieved by regulating several steps in the conversion of dietary vitamin A (retinol) to RA and RA catabolism. This review discusses the localization of RA synthesis to specific sites within the hair follicle and sebaceous gland, including their stem cells, during both homeostasis and disease states. It also discusses what is known about the specific roles of RA within the hair follicle and sebaceous gland. This article is part of a Special Issue entitled: Retinoid and Lipid Metabolism.

Topics: Animals; Hair Follicle; Humans; Mice; Receptors, Retinoic Acid; Sebaceous Glands; Stem Cells; Tretinoin; Vitamin A Deficiency

Vitamin A is essential for multiple functions in mammals. Without vitamin A, mammals cannot grow, reproduce, or fight off disease. Because of its numerous functions in humans, biomarkers of vitamin A status are quite diverse. Assessment of liver reserves of vitamin A is considered the gold standard because the liver is the major storage organ. However, this measure is not feasible in human studies. Alternative biomarkers of status can be classified as biological, functional, histologic, and biochemical. Historically, signs of xerophthalmia were used to determine vitamin A deficiency. Before overt clinical damage to the eye, individuals who suffer from vitamin A deficiency are plagued by night blindness and longer vision-restoration times. These types of assessments require large population-based evaluations. Therefore, surrogate biochemical measures of vitamin A status, as defined by liver reserves, have been developed. Serum retinol concentrations are a common method used to evaluate vitamin A deficiency. Serum retinol concentrations are homeostatically controlled until liver reserves are dangerously low. Therefore, other biochemical methods that respond to liver reserves in the marginal category were developed. These included dose-response tests and isotope dilution assays. Dose-response tests work on the principle that apo-retinol-binding protein builds up in the liver as liver reserves become depleted. A challenge dose of vitamin A binds to this protein, and serum concentrations increase within a few hours if liver vitamin A concentrations are low. Isotope dilution assays use stable isotopes as tracers of total body reserves of vitamin A and evaluate a wide range of liver reserves. Resources available and study objectives often dictate the choice of a biomarker.

Topics: Biomarkers; Biopsy; Humans; Liver; Milk, Human; Night Blindness; Nutritional Status; Tretinoin; Vitamin A; Vitamin A Deficiency

Retinoid acid, the bioactive metabolite of vitamin A, is a potent signaling molecule in the brains of growing and adult animals, regulates numerous gene products, and modulates neurogenesis, neuronal survival and synaptic plasticity. Vitamin A deficiency (VAD) is a global health problem, yet our knowledge of its effects on behavior and learning is still emerging. Here we review studies that have implicated retinoids in learning and memory deficits of post-embryonic and adult rodent and songbird models. Dietary vitamin A supplementation improves learning and memory in VAD rodents and can ameliorate cognitive declines associated with normal aging. Songbird studies examine the effects of retinoid signaling on vocal/auditory learning and are uniquely suited to study the behavioral effects of VAD because the neural circuitry of the song system is discrete and well understood. Similar to human speech acquisition, avian vocal learning proceeds in well-defined stages of template acquisition, rendition and maturation. Local blockade of retinoic acid production in the brain or excess dietary retinoic acid results in the failure of song maturation, yet does not affect prior song acquisition. Together these results yield significant insights into the role of vitamin A in maintaining neuronal plasticity and cognitive function in adulthood.

Topics: Animals; Behavior, Animal; Brain; Learning; Mice; Mice, Knockout; Models, Animal; Neuronal Plasticity; Neurotransmitter Agents; Songbirds; Tretinoin; Vitamin A; Vitamin A Deficiency; Vocalization, Animal

Vitamin A has been long associated with immune system competence. Vitamin A deficiency is known to compromise many aspects of both innate and adaptive immune responses. Recent advances in retinol uptake and metabolism have identified the antigen presenting cell (APC) as a central immune cell capable of vitamin A metabolism. APC are now known to express retinaldehyde dehydrogenase and secrete retinoic acid. The retinoic acid produced has both autocrine and paracrine effects. Autocrine effects include upregulation of CD1d nonclassical major histocompatibility class I-like molecule and matrix metalloproteinase-9. Paracrine effects influence multiple lymphocyte lineage cell populations. Specifically, retinoic acid increases IgA isotype class switching by B lymphocytes, enhances regulatory T cell differentiation, and directs homing of lymphocytes to mucosa. CD1d lipid antigen presentation expands natural killer T cell populations. Previously, the focus of vitamin A action in adaptive immunity was on lymphocytes, but these recent advances suggest the APC may be the central player in carrying out the immune system functions of vitamin A.

Topics: Animals; Antigen-Presenting Cells; Antigens, CD1d; B-Lymphocytes; Humans; Immunity; Immunoglobulin A; Killer Cells, Natural; Matrix Metalloproteinase 9; Retinal Dehydrogenase; T-Lymphocytes, Regulatory; Tretinoin; Up-Regulation; Vitamin A; Vitamin A Deficiency

Retinoic acid (RA) is a pleiotropic derivative of vitamin A, or retinol, which is responsible for all of the bioactivity associated with this vitamin. The teratogenic influences of vitamin A deficiency and excess RA in rodents were first observed more than 50 years ago. Efforts over the last 15-20 years have refined these observations by defining the molecular mechanisms that control RA availability and signaling during murine embryonic development. This review will discuss our current understanding of the role of RA in teratogenesis, with specific emphasis on the essential function of the RA catabolic CYP26 enzymes in preventing teratogenic consequences caused by uncontrolled distribution of RA. Particular focus will be paid to the RA-sensitive tissues of the caudal and cranial regions, the limb, and the testis, and how genetic mutation of factors controlling RA distribution have revealed important roles for RA during embryogenesis.

Topics: Animals; Congenital Abnormalities; Cytochrome P-450 Enzyme System; Embryonic Development; Extremities; Female; Humans; Male; Mice; Mice, Knockout; Neural Tube Defects; Pregnancy; Retinoic Acid 4-Hydroxylase; Teratogens; Testis; Tretinoin; Vitamin A Deficiency

Vitamin A insufficiency has profound adverse effects on embryonic development. Major advances in understanding the role of vitamin A in vertebrate heart formation have been made since the discovery that the vitamin A active form, all-trans-retinoic acid, regulates many genes, including developmental genes. Among the experimental models used, the vitamin A-deficient avian embryo has been an important tool to study the function of vitamin A during early heart formation. A cluster of retinoic acid-regulated developmental genes have been identified that participate in building the heart. In the absence of retinoic acid the embryonic heart develops abnormally leading to embryolethality.

Topics: Animals; Cadherins; Embryonic Development; Female; Heart; Mice; Pregnancy; Quail; Receptors, Retinoic Acid; Transforming Growth Factor beta2; Tretinoin; Vitamin A Deficiency

Vitamin A deficiency causes a marked reduction in the number of T and B cells in the small intestinal tissues. The vitamin A metabolite retinoic acid imprints lymphocytes with gut-homing specificity upon antigenic stimulation. In the small intestinal lamina propria, Peyer's patches, and mesenteric lymph nodes, there are dendritic cells capable of producing retinoic acid. Their capacity depends on the expression of retinal dehydrogenases (RALDH). RALDH2, encoded by Aldh1a2, is a major isoform of RALDH in the intestinal dendritic cells under specific pathogen-free conditions, and can be induced by multiple factors constitutively present or induced in the small intestinal microenvironment.

Topics: Aldehyde Oxidoreductases; Animals; Cell Movement; Dendritic Cells; Humans; Intestinal Mucosa; Intestines; Receptors, Retinoic Acid; T-Lymphocytes; Tretinoin; Vitamin A; Vitamin A Deficiency

Vitamin A (VA, retinol) is essential for normal immune system maturation, but the effect of VA(1) on antibody production, the hallmark of successful vaccination, is still not well understood. In countries where VA deficiency is a public health problem, many children worldwide are now receiving VA along with immunizations against poliovirus, measles, diphtheria, pertussis, and tetanus. The primary goal has been to provide enough VA to protect against the development of VA deficiency for a period of 4-6 months. However, it is also possible that VA might promote the vaccine antibody response. Several community studies, generally of small size, have been conducted in children supplemented with VA at the time of immunization, as promoted by the World Health Organization/UNICEF. However, only a few studies have reported differences in antibody titers or seroconversion rates due to VA. However, VA status was not directly assessed, and in some communities children were often breast fed, another strategy for preventing VA deficiency. Some of the vaccines used induced a high rate of seroconversion, even without VA. In children likely to have been VA deficient, oral polio vaccine seroconversion rate was increased by VA. In animal models, where VA status was controlled and VA deficiency confirmed, the antibody response to T-cell-dependent (TD) and polysaccharide antigens was significantly reduced, congruent with other defects in innate and adaptive immunity. Moreover, the active metabolite of VA, retinoic acid (RA) can potentiate antibody production to TD antigens in normal adult and neonatal animals. We speculate that numerous animal studies have correctly identified VA deficiency as a risk factor for low antibody production. A lack of effect of VA in human studies could be due to a low rate of VA deficiency in the populations studied or low sample numbers. The ability to detect differences in antibody response may also depend on the vaccine-adjuvant combination used. Future studies of VA supplementation and immunization should include assessment of VA status and a sufficiently large sample size. It would also be worthwhile to test the effect of neonatal VA supplementation on the response to immunization given after 6 months to 1 year of age, as VA supplementation, by preventing the onset of VA deficiency, may improve the response to immunizations given later on.

Topics: Animals; Antibody Formation; Dietary Supplements; Humans; Mass Vaccination; Tretinoin; Vitamin A; Vitamin A Deficiency; Vitamins

Within the last several years, research scientists and clinicians have been intrigued with the potential use of an active form of vitamin A, retinoic acid (RA), for the treatment and prevention of emphysema. The interest in this area can be largely attributed to the work of Massaro and Massaro (1996, 1997, 2000) in which they presented evidence that RA partially protects against and to some degree restores elastase-induced emphysema in rats. The mechanism for this protective effect of RA is in part related to elastin metabolism. RA also inhibits inflammation, an upstream event that may lead to the development of emphysema. Although there is evidence of this protective effect in young rats and a mechanistic explanation, more studies are needed in humans in order to establish a role for vitamin A in protecting against emphysema. Too many unanswered questions remain to definitively state that vitamin A protects against this disease in humans. Nevertheless, the potential for this novel approach in prevention and treatment of emphysema is an exciting area of research.

Topics: Animals; Elastin; Humans; Inflammation; Lung; Pulmonary Emphysema; Smoking; Tretinoin; Vitamin A; Vitamin A Deficiency; Vitamins

Aging of skin is an intricate biological process consisting of two types. While intrinsic or chronological aging is an inevitable process, photoaging involves the premature aging of skin occurring due to cumulative exposure to ultraviolet radiation. Chronological and photoaging both have clinically differentiable manifestations. Various natural and synthetic retinoids have been explored for the treatment of aging and many of them have shown histological and clinical improvement, but most of the studies have been carried out in patients presenting with photoaged skin. Amongst the retinoids, tretinoin possibly is the most potent and certainly the most widely investigated retinoid for photoaging therapy. Although retinoids show promise in the treatment of skin aging, irritant reactions such as burning, scaling or dermatitis associated with retinoid therapy limit their acceptance by patients. This problem is more prominent with tretinoin and tazarotene whereas other retinoids mainly represented by retinaldehyde and retinol are considerably less irritating. In order to minimize these side effects, various novel drug delivery systems have been developed. In particular, nanoparticles have shown a good potential in improving the stability, tolerability and efficacy ofretinoids like tretinoin and retinol. However, more elaborate clinical studies are required to confirm their advantage in the delivery of topical retinoids.

Topics: Adapalene; Dermatologic Agents; Dermis; Epidermis; Humans; Naphthalenes; Nicotinic Acids; Retinaldehyde; Retinoids; Skin Aging; Treatment Outcome; Tretinoin; Vitamin A Deficiency

Retinoic acid (RA) is essential for both embryonic and adult growth, activating gene transcription via specific nuclear receptors. It is generated, via a retinaldehyde intermediate, from retinol (vitamin A). RA levels require precise regulation by controlled synthesis and catabolism, and when RA concentrations deviate from normal, in either direction, abnormal growth and development occurs. This review describes: (i) how the pattern of RA metabolic enzymes controls the actions of RA; and (ii) the type of abnormalities that result when this pattern breaks down. Examples are given of RA control of the anterior/posterior axis of the hindbrain, the dorsal/ventral axis of the spinal cord, as well as certain sex-specific segments of the spinal cord, using varied animal models including mouse, quail and mosquitofish. These functions are highly sensitive to abnormal changes in RA concentration. In rodents, the control of neural patterning and differentiation are disrupted when RA concentrations are lowered, whereas inappropriately high concentrations of RA result in abnormal development of cerebellum and hindbrain nuclei. The latter parallels the malformations seen in the human embryo exposed to RA due to treatment of the mother with the acne drug Accutane (13-cis RA) and, in cases where the child survives beyond birth, a particular set of behavioural anomalies can be described. Even the adult brain may be susceptible to an imbalance of RA, particularly the hippocampus. This report shows how the properties of RA as a neural induction agent and organizer of segmentation can explain the consequences of RA depletion and overexpression.

Topics: Animals; Brain; Cell Differentiation; Embryonic and Fetal Development; Humans; Nervous System Physiological Phenomena; Neurons; Sex Characteristics; Teratogens; Tretinoin; Vitamin A; Vitamin A Deficiency

Vitamin-A is essential for growth and development of cells and tissues. In its active form, retinoic acid, it controls the regular differentiation as a ligand for retinoic acid receptors (RAR, RXR) and is involved in the integration (gap junction formation) of cell formations [Nature 37 (1994) 528; International Review of Cytology. San Diego Academic Press, 1-31]. Vitamin-A plays a substantial role, especially in the respiratory epithelium and the lung. During moderate vitamin-A-deficiency, the incidence for diseases of the respiratory tract is considerably increased and repeated respiratory infections can be influenced therapeutically by a moderate vitamin-A-supplementation [Aust. Paediatr. J. 22 (1986) 95; Lancet 338 (1991) 67]. In addition to the importance of the vitamin for the lung function, vitamin-A is also responsible for the development of many tissues and cells as well as for the embryonic lung development. Recent studies proved that the control occurs by different expressions of retinoid receptors as well as by time-dependent changes of the vitamin-A-metabolism respectively via cellular vitamin-A-binding proteins (CRBP: cytoplasmatic retinol binding protein; CRABP: cytoplasmatic retinoic acid binding protein).

Topics: Animals; Humans; Lung; Receptors, Retinoic Acid; Respiratory Mucosa; Retinol-Binding Proteins; Retinol-Binding Proteins, Cellular; Tretinoin; Vitamin A; Vitamin A Deficiency

Advances in molecular biology and retinoic acid receptor research have significantly contributed to the understanding of the role of vitamin A during vertebrate development. Examination of the function of this vitamin during very early developmental stages using the completely vitamin A-depleted avian embryo has revealed that the vitamin A requirement begins at the time of formation of the primitive heart, circulation and specification of hindbrain. The lack of vitamin A at this critical time results in gross abnormalities and early embryonic death. In rodent models, vitamin A deficiency can be targeted to later gestational windows and documents the need for vitamin A for more advanced stages of development. Major target tissues of vitamin A deficiency include the heart, central nervous system and structures derived from it, the circulatory, urogenital and respiratory systems, and the development of skull, skeleton and limbs. These abnormalities are also evident in mice mutants from retinoid receptor knockouts; they have revealed both morphological and molecular aspects of vitamin A function during development. Retinoic acid receptors (RAR) in partnership with retinoid X receptor (RXR)alpha appear to be the important retinoid receptor transcription factors regulating vitamin A function at the gene level during development via the physiologic ligand all-trans-retinoic acid. Homeostasis of retinoic acid is maintained by developmentally regulated vitamin A metabolism enzyme systems. Inadequate vitamin A nutrition during early pregnancy may account for some pediatric congenital abnormalities.

Topics: Animals; Embryonic and Fetal Development; Humans; Mice; Mice, Knockout; Models, Animal; Quail; Rats; Receptors, Retinoic Acid; Tretinoin; Vitamin A; Vitamin A Deficiency

The tissue distribution of retinoic acid (RA) throughout development is highly restricted, defined by the expression patterns of enzymes involved in RA synthesis and catabolism. Presented is a summary of recent research that examines the role of some of the enzymes involved in RA distribution, particularly those involved in RA catabolism (P450RAI). These latter enzymes protect against premature exposure to RA, and the implications of these findings are discussed.

Topics: Catalysis; Cytochrome P-450 Enzyme System; Humans; Keratolytic Agents; Mixed Function Oxygenases; Neoplasms; Retinoic Acid 4-Hydroxylase; Signal Transduction; Tretinoin; Vitamin A Deficiency

The retinoids, natural and synthetic derivatives of vitamin A, are active in cancer therapy and prevention. Their biological effects are mediated through ligand-dependent interactions with retinoid receptors that associate with specific co-regulators. A better understanding of retinoid chemopreventive mechanisms is needed. Our prior work revealed that all-trans-retinoic acid (RA) prevented tobacco-specific carcinogenic transformation of cultured human bronchial epithelial cells. RA signaled G1 arrest that permitted repair of genomic DNA damage caused by these carcinogens. RA triggered G1 arrest at least partly through proteasome-dependent degradation of cyclin D1. Proteasomal inhibitors blocked RA-mediated cyclin D1 degradation. To confirm that a specific proteolysis pathway was induced by RA-treatment, a degradation assay was established using in vitro translated cyclin D1 and cellular extracts from RA-treated or untreated human bronchial epithelial cells. Incubation of RA-treated but not the control cellular extracts with in vitro translated cyclin D1 led to cyclin degradation. This degradation depended on the PEST domain of cyclin D1, implicating ubiquitination in this retinoid degradation. Retinoid receptor selective agonists demonstrated that retinoic acid receptor (RAR)beta and retinoid X receptor (RXR) but not RARalpha- or RARgamma-dependent pathways signaled this cyclin degradation. Findings were extended to the NT2/D1 human embryonal carcinoma differentiation model where a similar pathway was activated by RA-treatment. To determine whether G1 cyclins were involved directly in bronchial preneoplasia, immunohistochemical expression profiles for cyclins D1 and E were examined. Aberrant expression of these cyclins was frequent in bronchial preneoplasia. Taken together, these findings indicate that ubiquitin-dependent proteolysis of G1 cyclins is a retinoid chemoprevention mechanism. Whether the retinoids represent the optimal agents to activate this pathway is the subject of ongoing work. These findings provide a rationale for combining the retinoids in chemoprevention trials with other agents that do not activate this proteolysis pathway. What is now known about the retinoids as cancer prevention agents will be reviewed. Emphasis is placed on retinoid effects on cell cycle progression at G1.

Topics: Animals; Anticarcinogenic Agents; Bronchi; Bronchial Diseases; Carcinoma, Embryonal; Cell Differentiation; Cell Transformation, Neoplastic; Cyclins; Cysteine Endopeptidases; Endopeptidases; Epithelial Cells; G1 Phase; Gene Expression Profiling; Gene Expression Regulation; Humans; Metaplasia; Mice; Models, Biological; Multienzyme Complexes; Neoplasms; Precancerous Conditions; Proteasome Endopeptidase Complex; Protein Processing, Post-Translational; Protein Structure, Tertiary; Receptors, Retinoic Acid; Retinoids; Tretinoin; Tumor Cells, Cultured; Ubiquitin; Vitamin A Deficiency

Retinoic acid (RA) is the bioactive metabolite of vitamin A (retinol) which acts on cells to establish or change the pattern of gene activity. Retinol is converted to RA by the action of two types of enzyme, retinol dehydrogenases and retinal dehydrogenases. In the nucleus RA acts as a ligand to activate two families of transcription factors, the RA receptors (RAR) and the retinoid X receptors (RXR) which heterodimerize and bind to the upstream sequences of RA-responsive genes. Thus, in addition to the well-established experimental paradigm of depriving animals of vitamin A to determine the role of RA in embryonic and post-embryonic development, molecular biology has provided us with two additional methodologies: knockout the enzymes or the RAR and RXR in the mouse embryo. The distribution of the enzymes and receptors, and recent experiments to determine the endogenous distribution of RA in the embryo are described here, as well as the effects on the embryo of knocking out the enzymes and receptors. In addition, recent studies using the classical vitamin A-deprivation technique are described, as they have provided novel insights into the regions of the embryo which crucially require RA, and the gene pathways involved in their development. Finally, the post-embryonic or regenerating systems in which RA plays a part are described, i.e. the regenerating limb, lung regeneration, hair cell regeneration in the ear and spinal cord regeneration in the adult.

Topics: Animals; Embryonic and Fetal Development; Female; Growth; Humans; Mice; Mice, Knockout; Pregnancy; Receptors, Retinoic Acid; Tretinoin; Vitamin A Deficiency

Topics: Animals; Diagnosis, Differential; Hearing Disorders; Humans; Neural Tube Defects; Night Blindness; Prognosis; Retinol-Binding Proteins; Tretinoin; Vitamin A Deficiency

The mechanism of the fetal embryopathology resulting from ethanol ingestion during pregnancy is not established. This review summarizes recent research on the interaction of ethanol and vitamin A in models that explore if an interaction between these two compounds might potentially be the mechanism for fetal alcohol syndrome. The rationale for this hypothesis includes the known facts that: (1) in adults, ethanol ingestion alters vitamin A metabolism and tissue distribution; (2) there are many phenotypic similarities between fetal alcohol syndrome and malformations of both vitamin A toxicity and deficiency; and (3) the vitamin A metabolite, retinoic acid (RA), is a potent mediator in embryogenesis and differentiation. One interaction that could possibly alter fetal development is that the synthesis of RA from retinol, catalyzed by alcohol dehydrogenase, might be competitively inhibited by ethanol leading to RA deficiency. Controversy over this hypothesis continues. Another model demonstrates in vivo effects of pregnant rat mother's ethanol consumption on retinol, retinyl ester, RA content, RA receptor (RAR) binding, and the levels of RAR expression in developing fetal organs. The variable responses in this model still need clarification, and specific defects resulting from specific RAR changes have not yet been identified. In a quail embryo model, ethanol treatment mimics vitamin A deficiency, and RA appears to prevent the adverse effects of ethanol. Finally, RA and ethanol reverse or block each other's effects in studies on isolated neuroblastoma cells. Taken together, these experiments show definite interactions between ethanol and vitamin A. Further studies are needed to determine if any of these mechanisms significantly contribute to prenatal ethanol consumption embryopathy; but, clearly this hypothesis is gaining experimental support.

Topics: Adult; Animals; Ethanol; Female; Fetal Alcohol Spectrum Disorders; Humans; Pregnancy; Rats; Tretinoin; Vitamin A Deficiency

A fuller understanding of the fundamental mechanisms involved in tumor initiation, growth, and metastasis will enable us to develop innovative approaches to detection and treatment that will improve the poor survival of patients with lung cancer. Current information suggests that certain individuals may be predisposed to developing lung cancer and that lung cancers, like other solid tumors, are characterized by the activation of oncogenes, the expression of growth factor loops, and the inactivation of tumor suppressor genes. Within the next decade, it is likely that genetic abnormalities will be used to identify individuals at risk for lung cancer, to select patients for adjuvant therapy, and to develop novel forms of treatment.

Topics: Animals; Carcinogens; Carcinoma, Non-Small-Cell Lung; Clinical Trials as Topic; Codon; Genes, Tumor Suppressor; Humans; Lung Neoplasms; Mutation; Oncogenes; Retrospective Studies; Smoking; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Animals; Epithelium; Humans; Receptors, Retinoic Acid; Skin Neoplasms; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Acyltransferases; Animals; Carboxylic Ester Hydrolases; Carrier Proteins; Humans; Hypervitaminosis A; Lipid Metabolism; Liver; Liver Cirrhosis; Receptors, Retinoic Acid; Retinol-Binding Proteins; Tretinoin; Vitamin A Deficiency

We discuss here both previously published data and our current experiments which suggest that the vitamin A derivative, retinoic acid (RA), may play a role in the development of the vertebrate central nervous system (CNS). This evidence comes from the following: both an excess and a deficiency of vitamin A causes embryonic defects of the CNS; RA has been detected endogenously in the CNS; RA stimulates neurite outgrowth; the retinoic acid receptors have been detected with interesting distributions in the CNS; the binding protein for retinol, namely cellular retinol binding protein (CRBP) is found in the radial glia of the ventral floor plate; the binding protein for RA, namely, cellular retinoic acid binding protein (CRABP) is found in particular sets of axons in the developing spinal cord, in particular rhombomeres in the developing hindbrain and in the neural crest. Some hypotheses for the possible role of RA in various aspects of CNS development are discussed.

Topics: Animals; Central Nervous System; Chick Embryo; Morphogenesis; Neural Crest; Retinol-Binding Proteins; Retinol-Binding Proteins, Cellular; Tretinoin; Vitamin A Deficiency

Vitamin A, an unsaturated 20 carbon cyclic alcohol, has a variety of physiological functions including a role in vision, reproduction, growth and maintenance of epithelial and bone structures. The main sources of vitamin A are from preformed vitamin A in animal foods or from -carotene in green plants. Carotene is cleaved in the intestinal mucosa to retinol, which is transported in chylomicrons mainly to the liver which is the major storage site of vitamin A, where stores are mainly of retinyl palmitate. Utilization of vitamin A appears to be a highly regulated process; retinol is transported in the serum bound to a specific binding protein. There also may be some control of the level of retinol in cells possibly through membrane receptors or of excretion from the cell. Intracellular cytosolic retinol binding proteins transport retinol to the nucleus where specific receptors for retinol have been found. Intracellular binding proteins for retinoic acid and retinal, metabolites of retinol have also been found, and an interstitial protein transporting retinol is present in the eye. Vitamin A deficiency causes cessation of growth, night blindness, and renders the organism more susceptible to infection, and vitamin A supplementation has been shown to enhance immune response. Epidemiological studies have shown low vitamin A and carotene to be correlated with incidence of cancer. Excess vitamin A intake results in hypervitaminosis with severe detrimental effects. Vitamin A requirements appear to be met in most developed countries although in the populations at greatest risk, newborns and pregnant and nursing women, cases of deficiency are observed. However, in large areas of the world vitamin A deficiency is endemic, causing widespread blindness and mortality.

Topics: Biological Transport, Active; Carotenoids; Diet; Eye; Humans; Intestinal Absorption; Liver; Nutritional Requirements; Tretinoin; Vitamin A; Vitamin A Deficiency

Retinoids have long been associated with wound healing, but objective data, until recently, have been scarce. Vitamin A deficiency retards repair. Secondly, retinoids restore steroid-retarded repair toward normal. Because vitamin A tends to suppress fibroblasts in cell culture and stimulate steroid-treated macrophages to initiate reparative behavior in tissue, we favor the hypothesis that retinoids are particularly important in macrophagic inflammation, which plays a central role in the control of wound healing. Probably all patients who take anti-inflammatory steroids should control their retinoid intake, but how they should control it is as yet unknown.

Topics: Animals; Drug Interactions; Fibroblasts; Humans; Retinoids; Skin Physiological Phenomena; Steroids; Tretinoin; Vitamin A; Vitamin A Deficiency; Wound Healing

Topics: Adolescent; Adult; Aged; Child; Drug Administration Schedule; Etretinate; Female; Humans; Isotretinoin; Retinoids; Skin Diseases; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Animals; Cell Differentiation; Humans; Liver; Male; Neoplasms; Nutritional Requirements; Retinoids; Retinol-Binding Proteins; Skin Diseases; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Animals; Carrier Proteins; Cell Differentiation; Chemical Phenomena; Chemistry; Epithelium; Female; Growth; Humans; Isomerism; Male; Neoplasms; Pregnancy; Receptors, Retinoic Acid; Reproduction; Retinal Pigments; Retinaldehyde; Retinol-Binding Proteins; Skin Diseases; Structure-Activity Relationship; Tretinoin; Vitamin A; Vitamin A Deficiency

Vitamin A refers collectively to a number of compounds, the most biologically active of which is retinol. Named after its earliest discovered function, visual excitation, retinol is also needed for growth and reproduction, and differentiation of epithelial tissue. A closely related form, retinoic acid, has selective biologic activity for normal growth and epithelial differentiation. The term retinoids has been used in recent years to include both natural and synthetic analogs of vitamin A. Carotene and carotenoids refer to vitamin A precursors (pro-vitamins) of vegetable origin. Following a brief historical account of vitamin A and description of dietary information this article focuses on the chemistry and physiology of retinoids. The biological roles of vitamin A compounds are elaborated, as well as the clinical significance of vitamin A deficiency and excess. Discussion of retinoids in therapy includes their use as anti-cancer agents and as therapeutics for dermatological disorders. Finally, a review of analytical techniques for laboratory assessment of vitamin A is presented.

Topics: Adult; Animals; Antineoplastic Agents; Carcinogens; Carotenoids; Chemical and Drug Induced Liver Injury; Chemical Phenomena; Chemistry; Female; Humans; Hypervitaminosis A; Infant; Male; Pregnancy; Rhodopsin; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Animals; Male; Rats; Retinol-Binding Proteins; Testis; Testosterone; Tretinoin; Vitamin A; Vitamin A Deficiency

The Vitamin A. derivatives known as Retinoids, are among the most exciting pharmacological agents used in the last few years. They strongly influence the keratinizing epithelia, have an antipromoting effect upon experimentally induced tumors and have immunomodulatory activities. Teratogenicity seems to be the most worrisome effect of the retinoids. However, it is reasonable to assume their release for physicians prescription in the near future. This review is intended to provide a general background for their correct use by dermatologists.

Topics: Animals; Carcinogens; Darier Disease; Humans; Ichthyosis; Mice; Neoplasms; Psoriasis; Rats; Skin Diseases; T-Lymphocytes; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Animals; Carcinogens; Cocarcinogenesis; Cyclamates; Cyclophosphamide; Humans; Hyperplasia; Mice; Neoplasms, Experimental; Rats; Saccharin; Schistosomiasis; Time Factors; Tretinoin; Tryptophan; Urinary Bladder; Urinary Bladder Neoplasms; Urinary Calculi; Vitamin A Deficiency

A survey is given concerning recent realizations of importance and metabolism of vitamin A in man and experimental animals. The transport of vitamin A from the small intestine takes place in form of esters by inclusion in chylomicrons and lipoproteins. A higher content in the blood plasma over a longer period evokes toxic effects. In the liver one part of the vitamin A in form of an ester with binding to lipoproteins is accumulated, one part is - according to the need - associated with a vitamin-A-binding protein, which forms complexes with prealbumin molecules and transports the vitamin to the various places of effect. With the help of the receptor proteins the vitamin A is included in the cells, in which it is effective and is transported into the vitamin-A-aldehyde and the vitamin-A-acid. The vitamin A and the vitamin-A-acid are transported into the cell nuclei with the help of receptor-proteins and play a role in the differentiation of the tissues and in the regulation of the synthesis of RNA. When a deficit or an abundance of vitamin A is present in the early phase of pregnancy malformations in the fetuses appear. The vitamin A and structure-similar connections have an inhibiting effect on the development of tumours.

Topics: Animals; Biological Transport; Birds; Carotenoids; Carrier Proteins; Chylomicrons; Congenital Abnormalities; Female; Fishes; Humans; Lipoproteins; Lung Neoplasms; Male; Mammals; Pregnancy; Pregnancy Trimester, First; Retinaldehyde; RNA; Smoking; Tretinoin; Vitamin A; Vitamin A Deficiency

The objective of the present study was to determine marginal vitamin A deficiency (VAD) by testing the hydrolysis of retinoyl glucuronide (RAG) to retinoic acid (RA) in children. Previous studies in rats showed that hydrolysis occurred when rats were vitamin A deficient. Children (n 61) aged 3-18 years, were divided into two groups, I and II. Blood was collected from the children in Group I (n 19) who were not dosed with RAG. Children in Group II (n 42) were administered all-trans retinoyl glucuronide (RAG) orally, and blood was collected 4 h after the dose. All serum samples were analysed for retinoids and carotenoids. RA was detected in serum only when serum retinol was < 0.85 micromol/l. Thus, hydrolysis of RAG to RA occurred in children with VAD or marginal VAD. Serum retinol was < 0.35 micromol/l in twenty-one children, 0.35-0.7 micromol/l in twenty-three children, 0.7-0.9 micromol/l in eleven children and >1 micromol/l in six children. Mean serum retinol in sixty-one children was 0.522 (sd 0.315) micromol/l. Mean beta-carotene (0.016 (sd 0.015) micromol/l) was far below normal compared to the level of lutein (0.176 (sd 0.10) micromol/l) in sixty-one children. A low beta-carotene level might be due to a low intake of carotene but high demand for vitamin A. The RAG hydrolysis test may prove to be a useful approach for the determination of marginal VAD with no clinical or subclinical signs of VAD. High prevalence of VAD amongst certain communities in Assam cannot be ruled out.

Topics: Adolescent; beta Carotene; Child; Child, Preschool; Chromatography, High Pressure Liquid; Developing Countries; Female; Humans; India; Linear Models; Lutein; Male; Nutritional Status; Spectrophotometry; Tretinoin; Vitamin A Deficiency

Vitamin A plays a prominent role for maintaining optimal bone status, but its impact upon the bone in response to vitamin A deficiency is not well defined. The purpose of this study was to evaluate how replenishing vitamin A by either whole food cod liver oil (COD) or the active metabolite of vitamin A, retinoic acid (RA), altered bone thickness of vitamin A-deficient (VAD) rats. Weanling rats were administered a control diet (CTRL) or VAD diet for 9 weeks. This was followed by four weeks of treatment in which the VAD group was divided into the following 4 subgroups: (1) VAD (9 weeks)-VAD (4 weeks); (2) VAD-CTRL; (3) VAD-COD; and (4) VAD-RA. Compared to controls, VAD rats had thicker bones which showed marked dysplasia. VAD-rats treated with COD produced a thinner bone that was not significantly different from that of untreated rats. In contrast, RA did not significantly change the thicker bone, and also had significantly greater periosteal and endosteal osteoblast numbers compared to VAD-COD. Active osteoclasts were not detected in VAD rats, nor during the treatment period. These findings suggest that the abnormal bone thickness in VAD rats appears to be more effectively restored to bone thickness of untreated control rats when treated with COD.

Topics: Animals; Cod Liver Oil; Rats; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Animals; Chemokine CCL5; Conjunctiva; Cornea; Dry Eye Syndromes; Eye; Female; Goblet Cells; Homeostasis; Immunity, Innate; Inflammation; Interleukin-12 Subunit p35; Interleukin-1beta; Lacrimal Apparatus; Mice; Mice, Inbred C57BL; Myeloid Cells; Receptors, Retinoic Acid; Retinoids; Signal Transduction; Tretinoin; Vitamin A; Vitamin A Deficiency

Vitamin A (VA) deficiency remains prevalent in resource limited areas. Using Citrobacter rodentium infection in mice as a model for diarrheal diseases, previous reports showed reduced pathogen clearance and survival due to vitamin A deficient (VAD) status. To characterize the impact of preexisting VA deficiency on gene expression patterns in the intestines, and to discover novel target genes in VA-related biological pathways, VA deficiency in mice were induced by diet. Total mRNAs were extracted from small intestine (SI) and colon, and sequenced. Differentially Expressed Gene (DEG), Gene Ontology (GO) enrichment, and co-expression network analyses were performed. DEGs compared between VAS and VAD groups detected 49 SI and 94 colon genes. By GO information, SI DEGs were significantly enriched in categories relevant to retinoid metabolic process, molecule binding, and immune function. Three co-expression modules showed significant correlation with VA status in SI; these modules contained four known retinoic acid targets. In addition, other SI genes of interest (e.g., Mbl2, Cxcl14, and Nr0b2) in these modules were suggested as new candidate genes regulated by VA. Furthermore, our analysis showed that markers of two cell types in SI, mast cells and Tuft cells, were significantly altered by VA status. In colon, "cell division" was the only enriched category and was negatively associated with VA. Thus, these data suggested that SI and colon have distinct networks under the regulation of dietary VA, and that preexisting VA deficiency could have a significant impact on the host response to a variety of disease conditions.

Topics: Animals; Citrobacter rodentium; Colon; Enterobacteriaceae Infections; Gene Expression Profiling; Gene Ontology; Intestine, Small; Mice; Mice, Inbred C57BL; RNA-Seq; RNA, Messenger; Transcriptome; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Adoptive Transfer; Animals; Cells, Cultured; Disease Resistance; Lymphocyte Activation; Lymphocytic Choriomeningitis; Lymphocytic choriomeningitis virus; Mice, Inbred C57BL; Mice, Knockout; Oncogene Protein v-akt; Receptors, Antigen, T-Cell; Signal Transduction; T-Lymphocytes; Tretinoin; Vitamin A Deficiency

Topics: Cell Differentiation; Humans; Lymphotoxin-alpha; Tretinoin; Vitamin A; Vitamin A Deficiency

This study presents two new concepts and definitions to the medical literature. One of those is "endogenous retinoic acid theory" and the other "retinoic acid depletion syndrome". A new classification will be provided for the immune system: "retinoic acid-dependent component" and "retinoic acid non-dependent component". If this theory is verified, all the diseases where the retinoic acid metabolism is defective and retinoic acid levels are low will be identified and new approaches will be developed fortreating such diseases. When the need for retinoic acids increases, such as acute infection, high fever, severe catabolic process, or chronic antigenic stimulation, cytochrome oxidase enzymes are inhibited by drugs or internal mechanisms. Metabolism and excretion of retinoic acids stored in the liver are prevented. In this way, retinoic acid levels in the blood are raised to therapeutic levels. This is called "Endogenous Retinoic Acid Theory". Retinoic acids also manage their metabolism through feedback mechanisms. Despite compensatory mechanisms, causes such as high fever, serious catabolic process and excessively large viral genome (SARS-CoV-2), excessive use of RIG-I and Type I interferon synthesis pathway using retinoic acid causes emptying of retinoic acid stores. As a result, the RIG-I pathway becomes ineffective, Type I IFN synthesis stops, and the congenital immune system collapses. Then the immune mechanism passes to TLR3, TLR7, TLR8, TLR9, MDA5 and UPS pathways in the monocyte, macrophage, neutrophil and dendritic cells of the adaptive immune defense system that do not require retinoic acid. This leads to excessive TNFα and cytokine discharge from the pathway. With the depletion of retinoic acid stores as a result of this overuse, the immune defense mechanism switches from the congenital immune system to the adaptive immune system, where retinoic acids cannot be used. As a result of this depletion of retinoic acids, the shift of the immune system to the NFκB arm, which causes excessive cytokine release, is called "retinoic acid depletion syndrome". COVID-19 and previously defined sepsis, SIRS and ARDS are each retinoic acid depletion syndrome. We claim that retinoic acid metabolism is defective in most inflammatory diseases, particularly COVID-19 (cytokine storm) sepsis, SIRS and ARDS. Finding a solution to this mechanism will bring a new perspective and treatment approach to such diseases.

Topics: Autoimmunity; Carotenoids; COVID-19; DEAD Box Protein 58; Humans; Immune System; Interferon Type I; Interferons; Liver; Models, Theoretical; Nervous System; Receptors, Immunologic; Syndrome; Tretinoin; Viral Load; Vitamin A; Vitamin A Deficiency; Zinc

Activation of hepatic stellate cells (HSCs) is the effector factor of hepatic fibrosis and hepatocellular carcinoma (HCC) development. Accumulating evidence suggests that retinoic acids (RAs), derivatives of vitamin A, contribute to prevention of liver fibrosis and carcinogenesis, however, regulatory mechanisms of RAs still remain exclusive. To elucidate RA signaling pathway, we previously performed a genome-wide screening of RA-responsive genes by in silico analysis of RA-response elements, and identified 26 RA-responsive genes. We found that thioredoxin interacting protein (TXNIP), which inhibits antioxidant activity of thioredoxin (TRX), was downregulated by all-trans retinoic acid (ATRA). In the present study, we demonstrate that ATRA ameliorates activation of HSCs through TXNIP suppression. HSC activation was attenuated by TXNIP downregulation, whereas potentiated by TXNIP upregulation, indicating that TXNIP plays a crucial role in activation of HSCs. Notably, we showed that TXNIP-mediated HSC activation was suppressed by antioxidant N-acetylcysteine. In addition, ATRA treatment or downregulation of TXNIP clearly declined oxidative stress levels in activated HSCs. These data suggest that ATRA plays a key role in inhibition of HSC activation via suppressing TXNIP expression, which reduces oxidative stress levels.

Topics: Animals; Antioxidants; Carrier Proteins; Cell Line; Disease Models, Animal; Dose-Response Relationship, Drug; Gene Expression Regulation; Hepatic Stellate Cells; Humans; Mice, Inbred C57BL; Oxidative Stress; RNA Interference; Signal Transduction; Thioredoxins; Transfection; Tretinoin; Vitamin A Deficiency

Our previous studies demonstrated that vitamin A deficiency (VAD) can impair the postnatal cognitive function of rats by damaging the hippocampus. The present study examined the effects of retinoic acid (RA) on apoptosis induced by hypoxic-ischemic damage in vivo and in vitro, and investigated the possible signaling pathway involved in the neuroprotective anti-apoptotic effects of RA. Flow cytometry, immunofluorescence staining and behavioral tests were used to evaluate the neuroprotective and anti-apoptotic effects of RA. The protein and mRNA levels of RARα, PI3K, Akt, Bad, caspase-3, caspase-8, Bcl-2, Bax, and Bid were measured with western blotting and real-time PCR, respectively. We found impairments in learning and spatial memory in VAD group compared with vitamin A normal (VAN) and vitamin A supplemented (VAS) group. Additionally, we showed that hippocampal apoptosis was weaker in the VAN group than that in VAD group. Relative to the VAD group, the VAN group also had increased mRNA and protein levels of RARα and PI3K, and upregulated phosphorylated Akt/Bad levels in vivo. In vitro, excessively low or high RA signaling promoted apoptosis. Furthermore, the effects on apoptosis involved the mitochondrial membrane potential (MMP). These data support the idea that sustained VAD following hypoxic-ischemic brain damage (HIBD) inhibits RARα, which downregulates the PI3K/Akt/Bad and Bcl-2/Bax pathways and upregulates the caspase-8/Bid pathway to influence the MMP, ultimately producing deficits in learning and spatial memory in adolescence. This suggests that clinical interventions for HIBD should include suitable doses of VA.

Topics: Animals; Apoptosis; bcl-2-Associated X Protein; Caspases; Cells, Cultured; Dietary Supplements; Female; Glucose; Hippocampus; Hypoxia-Ischemia, Brain; Learning; Membrane Potential, Mitochondrial; Mitochondria; Neurons; Oxygen; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins c-akt; Rats, Sprague-Dawley; Retinoic Acid Receptor alpha; RNA, Messenger; Signal Transduction; Spatial Memory; Tretinoin; Vitamin A; Vitamin A Deficiency

This study investigated the prenatal marginal vitamin A deficiency (mVAD)-related impairment in learning and memory and the interactions between RARα, Src and NR1. Learning and memory were assessed in adult rats that were exposed to prenatal mVAD with Morris water maze. The average escape time was longer in mVAD rats, and they passed the hidden platform fewer times during the memory retention test than normal vitamin A intake (VAN) rats. The mRNA and protein levels of RARα, Src and NR1 in mVAD rats were significantly lower than those in VAN rats. RARα and Src, but not NR1, were in the same protein complex. RA treatment-induced increase in RARα, Src and NR1 expressions in mVAD neurons was much lower than that in VAN neurons. Overexpression of RARα gene in VAN neurons induced an increase in RARα, Src and NR1 expressions, while silencing of RARα gene induced a decrease in expressions of RARα and Src, but not that of of NR1. In mVAD neurons, however, overexpression of RARα did not induce an increase in NR1 expression, while silencing of RARα gene had no effect on Src and NR1 expressions. Furthermore, inhibition of Src was associated with a decrease in NR1 expression but not that of RARα. Prenatal mVAD was associated with impaired learning and memory in adult rats. It is possible that mVAD-related decrease in RARα led to a decrease in Src expression, which in turn down-regulated NR1 expression and Ca

Topics: Animals; Animals, Newborn; Cells, Cultured; Female; Gene Expression Regulation, Developmental; HEK293 Cells; Hippocampus; Humans; Learning Disabilities; Male; Maternal Nutritional Physiological Phenomena; Memory Disorders; Nerve Tissue Proteins; Neurons; Pregnancy; Proto-Oncogene Proteins pp60(c-src); Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Retinoic Acid Receptor alpha; RNA Interference; Severity of Illness Index; Tretinoin; Vitamin A; Vitamin A Deficiency

The present study extends an earlier report that retinoic acid (RA) down-regulates IgE Ab synthesis in vitro. Here, we show the suppressive activity of RA on IgE production in vivo and its underlying mechanisms. We found that RA down-regulated IgE class switching recombination (CSR) mainly through RA receptor α (RARα). Additionally, RA inhibited histone acetylation of germ-line ε (GL ε) promoter, leading to suppression of IgE CSR. Consistently, serum IgE levels were substantially elevated in vitamin A-deficient (VAD) mice and this was more dramatic in VAD-lecithin:retinol acyltransferase deficient (LRAT

Topics: Acyltransferases; Animals; Chymases; Food Hypersensitivity; Immunoglobulin Class Switching; Immunoglobulin E; Interleukin-4; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Retinoic Acid Receptor alpha; T-Lymphocytes, Regulatory; Tretinoin; Vitamin A; Vitamin A Deficiency

Retinoic acid (RA) signaling is crucial for spermatogonial differentiation, which is a key step for spermatogenesis. We explored the mechanisms underlying spermatogonial differentiation by targeting expression of a dominant-negative mutant of retinoic acid receptor α (RARα) specifically to the germ cells of transgenic mice to subvert the activity of endogenous receptors. Here we show that: (1) inhibition of retinoid signaling in germ cells completely blocked spermatogonial differentiation identical to vitamin A-deficient (VAD) mice; (2) the blockage of spermatogonial differentiation by impaired retinoid signaling resulted from an arrest of entry of the undifferentiated spermatogonia into S phase; and (3) retinoid signaling regulated spermatogonial differentiation through controlling expression of its direct target genes, including replication-dependent core histone genes. Taken together, our results demonstrate that the action of retinoid signaling on spermatogonial differentiation in vivo is direct through the spermatogonia itself, and provide the first evidence that this is mediated by regulation of expression of replication-dependent core histone genes.

Topics: Animals; Cell Differentiation; Histones; Male; Mice; Mice, Transgenic; Retinoic Acid Receptor alpha; S Phase Cell Cycle Checkpoints; Signal Transduction; Spermatogenesis; Spermatogonia; Testis; Tretinoin; Vitamin A Deficiency

Changes in diet and microbiota have determining effects on the function of the mucosal immune system. For example, the active metabolite of vitamin A, retinoic acid (RA), has been described to maintain homeostasis in the intestine by its influence on both lymphocytes and myeloid cells. Additionally, innate lymphoid cells (ILCs), important producers of cytokines necessary for intestinal homeostasis, are also influenced by vitamin A in the small intestines. In this study, we show a reduction of both NCR(-) and NCR(+) ILC3 subsets in the small intestine of mice raised on a vitamin A-deficient diet. Additionally, the percentages of IL-22-producing ILCs were reduced in the absence of dietary vitamin A. Conversely, mice receiving additional RA had a specific increase in the NCR(-) ILC3 subset, which contains the lymphoid tissue inducer cells. The dependence of lymphoid tissue inducer cells on vitamin A was furthermore illustrated by impaired development of enteric lymphoid tissues in vitamin A-deficient mice. These effects were a direct consequence of ILC-intrinsic RA signaling, because retinoic acid-related orphan receptor γt-Cre × RARα-DN mice had reduced numbers of NCR(-) and NCR(+) ILC3 subsets within the small intestine. However, lymphoid tissue inducer cells were not affected in these mice nor was the formation of enteric lymphoid tissue, demonstrating that the onset of RA signaling might take place before retinoic acid-related orphan receptor γt is expressed on lymphoid tissue inducer cells. Taken together, our data show an important role for vitamin A in controlling innate lymphoid cells and, consequently, postnatal formed lymphoid tissues within the small intestines.

Topics: Animals; Antigens, Ly; Immunity, Innate; Intestine, Small; Lymphocyte Subsets; Lymphoid Tissue; Mice; Natural Cytotoxicity Triggering Receptor 1; Nuclear Receptor Subfamily 1, Group F, Member 3; Signal Transduction; Tretinoin; Vitamin A; Vitamin A Deficiency

Ras signaling is tightly regulated during neural stem cell (NSC) differentiation, and defects in this pathway result in aberrant brain development. However, the mechanism regulating Ras signaling during NSC differentiation was unknown. Here, we show that stabilized HRas specifically induces neuronal differentiation of NSCs. Lentivirus-mediated HRas overexpression and knockdown resulted in stimulation and inhibition, respectively, of NSC differentiation into neuron in the ex vivo embryo. Retinoic acid, an active metabolite of vitamin A, promoted neuronal differentiation of NSCs by stabilizing HRas, and HRas knockdown blocked the retinoic acid effect. Vitamin-A-deficient mice displayed abnormal brain development with reduced HRas levels and a reduced thickness of the postmitotic region containing differentiated neurons. All of these abnormal phenotypes were rescued with the restoration of HRas protein levels achieved upon feeding with a retinoic-acid-supplemented diet. In summary, this study shows that retinoic acid stabilizes HRas protein during neurogenesis, and that this is required for NSC differentiation into neurons and murine brain development.

Topics: Animals; Brain; Cell Differentiation; Embryo, Mammalian; Gene Expression Profiling; Gene Expression Regulation, Developmental; HEK293 Cells; Humans; Mice, Inbred C57BL; Neural Stem Cells; Neurons; Protein Stability; Proto-Oncogene Proteins p21(ras); Tretinoin; Vitamin A Deficiency

The aim of this study was to compare if lycopene also possesses pro-vitamin A (VA) activity comparable to known VA derivatives.. We used a transgenic retinoic acid response element reporter mouse model (n = 8, per group) for this study, and after the initial wash out of VA using a vitamin A deficient diet (VAD) for 18 weeks, the animals were supplemented further with (a) VAD-fed mice, (b) VAD-fed mice plus retinol (20 mg/kg bw), (c) VAD-fed mice plus β-carotene (40 mg/kg bw), and (d) VAD-fed mice plus lycopene (40 mg/kg bw). Using ex vivo scanning and gene expression analysis of retinoid target and VA marker gene analysis in various organs of these supplemented mice (b, c, d), we found increased luciferase activity and normalized marker and target gene analysis compared to group a.. Lycopene can restore VA deficiency and compensate VA for retinoic acid receptor (RAR)-mediated signaling as the major function of VA in the mammalian organism. Lycopene administration can initiate upregulation of RAR-mediated signaling in various organs in VAD-fed animals via potential novel bioactive lycopene metabolites. This indicates that lycopene possesses partial pro-VA activity in mice transmitted via RAR-mediated signaling.

Topics: Animals; Carotenoids; Carrier Proteins; Diet; Dietary Supplements; Liver; Luciferases; Lycopene; Mice; Mice, Transgenic; Receptors, Retinoic Acid; Retinoids; RNA, Messenger; Signal Transduction; Transcriptional Activation; Tretinoin; Vitamin A Deficiency

Retinoic acid, the bioactive metabolite of beta-carotene or vitamin A, plays a pleiotropic, multifunctional role in vertebrate development. Studies in rodents revealed that a diet deficient in vitamin A results in a complex neonatal syndrome (the VAD syndrome), manifested in many organs. In humans, the function of retinoic acid (RA) extends into adulthood, where it has important roles in fertility, vision, and suppression of neoplastic growth. In recent years, it has also been suggested that retinoic acid might potentially act as a therapeutically relevant drug in attenuating or even preventing neurodegenerative diseases such as Alzheimer's disease (AD). Here, we report that VAD leads to an increase in A-beta peptide levels while only minor effects were observed on expression levels of the amyloid precursor protein (APP) processing proteinases in wild type mice. In line with these findings, rescue of hypovitaminosis reduced A-beta amount to baseline and induced sApp-alpha secretion in combination with an increase of alpha-secretase Adam10. By comparing retinoic acid treatment starting from a full nutrition status and a "VAD" situation in human neuroblastoma cells, we show that while intensities of differential gene expression were higher in replenished cells, a large overlap in AD-related, regulated genes was observed. Our data suggest that hypovitaminosis A can contribute to onset or progression of AD by increasing synthesis of A-beta peptides and that several AD-related genes such as ADAM10 or BDNF are regulated by retinoic acid. We suggest that dietary supplementation with retinoic acid derivatives is likely to have a beneficial effect on AD-pathology in individuals with hypovitaminosis and patients with normal vitamin A status.

Topics: Acitretin; ADAM10 Protein; Amyloid beta-Peptides; Amyloid beta-Protein Precursor; Animals; Animals, Newborn; Cells, Cultured; Cerebral Cortex; Disease Models, Animal; Female; Gene Regulatory Networks; Humans; Keratolytic Agents; Mice; Neuroblastoma; Neurons; Peptide Fragments; Presenilin-2; Rats, Wistar; Tretinoin; Vitamin A Deficiency

Vitamin A deficiency is known to affect 20 million pregnant women worldwide. However, the prenatal effects of maternal vitamin A deficiency on pancreas development have not been clearly determined. The present study examined how maternal vitamin A deficiency affects fetal islet development. Vitamin A-deficient mice were generated by feeding female mice with a chemically defined diet lacking vitamin A prior to mating as well as during pregnancy. We found that maternal vitamin A deficiency during pregnancy affected fetal pancreas development. Although the exocrine differentiation appeared normal, development of islet tissue was impaired. In the pancreas of neonatal mice, only a few endocrine cell clusters were formed, and these cell clusters lacked capillary endothelial cells. To further determine how vitamin A metabolites, such as retinoic acid, regulate vascularized islet development, ex vivo culture of embryonic pancreas either in the presence of 4-diethylaminobenzaldehyde (DEAB; an inhibitor of retinaldehyde dehydrogenase), all-trans retinoic acid (atRA) or retinoic acid receptor agonist (E)-4-[2-(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-naphthylenyl)-1-propenyl] benzoic acid (TTNPB) was carried out. We found that the addition of DEAB blocked vascularization and suppressed β-cell differentiation. Conversely, atRA or TTNPB promoted β-cell differentiation accompanied by enhanced expression of vascular basement component, laminin. We further demonstrated that atRA regulated vascularization via upregulating vascular endothelial growth factor-A (VEGF-A) secretion in embryonic pancreas and treatment with VEGF-A was able to partially rescue vascularization and β-cell differentiation in DEAB-treated embryonic pancreas cultures. The findings explain why maternal vitamin A deficiency affects fetal islet development and support an essential role of retinoid signaling in regulating vascularized islet development.

Topics: Animals; Animals, Newborn; Benzaldehydes; Benzoates; Cell Differentiation; Embryo, Mammalian; Enzyme Inhibitors; Female; Fetal Development; Insulin-Secreting Cells; Islets of Langerhans; Maternal Nutritional Physiological Phenomena; Mice, Inbred C57BL; Mice, Transgenic; Neovascularization, Physiologic; Pregnancy; Random Allocation; Receptors, Retinoic Acid; Retinal Dehydrogenase; Retinoids; Tissue Culture Techniques; Tretinoin; Vitamin A Deficiency

Topics: Allografts; Animals; CD4 Antigens; Graft vs Host Disease; Hematopoietic Stem Cell Transplantation; Mice; Receptors, Retinoic Acid; T-Lymphocytes; Tissue Donors; Tretinoin; Vitamin A Deficiency

Chronic intestinal inflammation accompanies familial adenomatous polyposis (FAP) and is a major risk factor for colorectal cancer in patients with this disease, but the cause of such inflammation is unknown. Because retinoic acid (RA) plays a critical role in maintaining immune homeostasis in the intestine, we hypothesized that altered RA metabolism contributes to inflammation and tumorigenesis in FAP. To assess this hypothesis, we analyzed RA metabolism in the intestines of patients with FAP as well as APC

Topics: Adenoma; Adenomatous Polyposis Coli; Animals; Antineoplastic Agents; Cell Transformation, Neoplastic; Colorectal Neoplasms; Dendritic Cells; Enterocolitis; Genes, APC; Humans; Mice; Phenotype; Th17 Cells; Tretinoin; Tumor Burden; Vitamin A; Vitamin A Deficiency

Cellular uptake of vitamin A (retinol) is essential for many biological functions. The Stra6 protein binds the serum retinol-binding protein, RBP4, and acts in conjunction with the enzyme lecithin:retinol acyltransferase to facilitate retinol uptake in some cell types. We show that in embryonic stem (ES) cells and in some tissues, the Stra6 gene encodes two distinct mRNAs transcribed from two different promoters. Whereas both are all-trans-retinoic acid (RA)-responsive in ES cells, the downstream promoter contains a half-site RA response element (RARE) and drives an ∼ 13-fold, RA-associated increase in luciferase reporter activity. We employed CRISPR-Cas9 genome editing to show that the endogenous RARE is required for RA-induced transcription of both Stra6 isoforms. We further demonstrate that in ES cells, 1) both RARγ and RXRα are present at the Stra6 RARE; 2) RA increases co-activator p300 (KAT3B) binding and histone H3 Lys-27 acetylation at both promoters; 3) RA decreases Suz12 levels and histone H3 Lys-27 trimethylation epigenetic marks at both promoters; and 4) these epigenetic changes are diminished in the absence of RARγ. In the brains of WT mice, both the longer and the shorter Stra6 transcript (Stra6L and Stra6S, respectively) are highly expressed, whereas these transcripts are found only at low levels in RARγ(-/-) mice. In the brains of vitamin A-deficient mice, both Stra6L and Stra6S levels are decreased. In contrast, in the vitamin A-deficient kidneys, the Stra6L levels are greatly increased, whereas Stra6S levels are decreased. Our data show that kidneys respond to retinol deficiency by differential Stra6 promoter usage, which may play a role in the retention of retinol when vitamin A is low.

Topics: Animals; Base Sequence; Blotting, Western; Brain; Cells, Cultured; Chromatin Immunoprecipitation; Embryonic Stem Cells; Epigenesis, Genetic; Gene Expression Regulation; Membrane Proteins; Mice; Mice, Inbred C57BL; Mice, Knockout; Molecular Sequence Data; Promoter Regions, Genetic; Protein Isoforms; Real-Time Polymerase Chain Reaction; Receptors, Retinoic Acid; Response Elements; Retinoic Acid Receptor gamma; Reverse Transcriptase Polymerase Chain Reaction; RNA Splicing; RNA, Messenger; Transcription Initiation Site; Tretinoin; Vitamin A Deficiency

Stem cells ensure tissue homeostasis through the production of differentiating and self-renewing progeny. In some tissues, this is achieved by the function of a definitive stem cell niche. However, the mechanisms that operate in mouse spermatogenesis are unknown because undifferentiated spermatogonia (Aundiff) are motile and intermingle with differentiating cells in an 'open' niche environment of seminiferous tubules. Aundiff include glial cell line-derived neurotrophic factor receptor α1 (GFRα1)(+) and neurogenin 3 (NGN3)(+) subpopulations, both of which retain the ability to self-renew. However, whereas GFRα1(+) cells comprise the homeostatic stem cell pool, NGN3(+) cells show a higher probability to differentiate into KIT(+) spermatogonia by as yet unknown mechanisms. In the present study, by combining fate analysis of pulse-labeled cells and a model of vitamin A deficiency, we demonstrate that retinoic acid (RA), which may periodically increase in concentration in the tubules during the seminiferous epithelial cycle, induced only NGN3(+) cells to differentiate. Comparison of gene expression revealed that retinoic acid receptor γ (Rarg) was predominantly expressed in NGN3(+) cells, but not in GFRα1(+) cells, whereas the expression levels of many other RA response-related genes were similar in the two populations. Ectopic expression of RARγ was sufficient to induce GFRα1(+) cells to directly differentiate to KIT(+) cells without transiting the NGN3(+) state. Therefore, RARγ plays key roles in the differentiation competence of NGN3(+) cells. We propose a novel mechanism of stem cell fate selection in an open niche environment whereby undifferentiated cells show heterogeneous competence to differentiate in response to ubiquitously distributed differentiation-inducing signals.

Topics: Animals; Basic Helix-Loop-Helix Transcription Factors; Cell Differentiation; Fluorescent Antibody Technique; Glial Cell Line-Derived Neurotrophic Factor Receptors; In Situ Hybridization; Male; Mice; Mice, Transgenic; Nerve Tissue Proteins; Real-Time Polymerase Chain Reaction; Receptors, Retinoic Acid; Retinoic Acid Receptor gamma; Spermatogenesis; Stem Cells; Tretinoin; Vitamin A Deficiency

We report the ocular and systemic manifestations of vitamin A deficiency in a child with a complicated medical history including autism and a restricted diet, living in a developed country. This child had significant vitamin A deficiency despite being under long-term medical care, yet the diagnosis was not considered until he had an ophthalmology review for visual deterioration.

Topics: Autistic Disorder; Child; Child Nutritional Physiological Phenomena; Diet; Feeding and Eating Disorders of Childhood; Feeding Behavior; Humans; Keratolytic Agents; Male; Malnutrition; Treatment Outcome; Tretinoin; Vision Disorders; Vitamin A; Vitamin A Deficiency; Xerophthalmia

Patients with alcoholic liver disease (ALD) have vitamin A (VA) deficiency and an enhanced immune response associated with disease severity. All-trans retinoic acid (ATRA), a VA-active metabolite, has anti-inflammatory effects and its deficiency could contribute to the exacerbated proinflammatory reaction. The aim of this study was to investigate the effects of ATRA/VA deficiency and supplementation on the monocyte response in ALD.. Vitamin A and ATRA plasma levels were quantified in ALD patients and healthy subjects (HS). The in vitro effect of ATRA on lipopolysaccharide (LPS)-induced TNF-α production by human peripheral blood mononuclear cells (PBMC) was assessed by ELISA and RT-PCR. The activation pattern of peritoneal macrophages (PerMΦ) and circulating monocytes isolated from VA-deficient mice and ALD patients, respectively, was evaluated by flow cytometry, quantification of TNF-α and NO2 production.. Alcoholic liver disease patients (n = 85) showed plasmatic VA deficiency that was correlated with scores of severity and with the hepatic venous pressure gradient. ATRA levels correlated significantly with VA levels. In vitro, ATRA pretreatment decreased the overproduction of TNF-α by LPS-stimulated PBMC of ALD patients. In vivo, VA deficiency in mice was associated with increased activation of PerMΦ, while oral ATRA supplementation normalized it.. For the first time, we show that VA/ATRA deficiencies in ALD patients are associated with disease severity. Furthermore, our data strongly suggest that the VA deficiency observed in ALD patients might participate in the pathophysiology of the disease by priming immune cells, and that ATRA supplementation could downregulate the deleterious proinflammatory state in cirrhosis and might thus be of therapeutic use.

Topics: Adult; Aged; Animals; Case-Control Studies; Cells, Cultured; Disease Models, Animal; Down-Regulation; Female; Humans; Lipopolysaccharides; Liver Cirrhosis, Alcoholic; Macrophage Activation; Macrophages, Peritoneal; Male; Mice; Mice, Inbred C57BL; Middle Aged; Monocytes; RNA, Messenger; Tretinoin; Tumor Necrosis Factor-alpha; Vitamin A; Vitamin A Deficiency

Hearing loss is a substantial public health problem with profound social and economic consequences in the developing world. The World Health Organization (WHO) estimates that there are 360 million people living with disabling hearing loss globally, and 80% of these individuals are from low- and middle-income countries. The epidemiology of hearing impairment remains poorly defined in most impoverished societies. Middle ear infections in childhood are a key determinant; however, congenital anomalies may also comprise an important etiology and may arise from gestational malnutrition. While evidence exists that preventable vitamin A deficiency exacerbates the severity of ear infections and, consequently, hearing loss, antenatal vitamin A deficiency during sensitive periods of fetal development may represent an etiologically distinct and virtually unexplored causal pathway. Evidence from multiple animal systems clearly shows that fetal inner ear development requires adequate vitamin A nutriture to proceed normally. Inner ear malformations occur in experimentally imposed maternal vitamin A deficiency in multiple species in a dose-response manner. These anomalies are likely due to the loss of retinoic acid-dependent regulation of both hindbrain development and otic morphogenic processes. Based on in vivo evidence in experimental animals, we hypothesize that preventable gestational vitamin A deficiency, especially during early stages of fetal development, may predispose offspring to inner ear malformations and sensorineural hearing loss. As vitamin A deficiency affects an estimated 20 million pregnant women globally, we hypothesize that, in undernourished settings, routine provision of supplemental vitamin A at the recommended allowance throughout pregnancy may promote normal inner ear development and reduce risk of an as yet unknown fraction of sensorineural hearing loss. If our hypothesis proves correct, gestational vitamin A deficiency would represent a potentially preventable etiology of sensorineural hearing loss of substantial public health significance.

Topics: Developing Countries; Dietary Supplements; Ear, Inner; Female; Hearing Loss, Sensorineural; Humans; Models, Biological; Pregnancy; Tretinoin; Vitamin A; Vitamin A Deficiency

The vitamin A (VA) metabolite retinoic acid (RA) affects the properties of T cells and dendritic cells (DCs). In VA-deficient mice, we observed that mesenteric lymph node (MLN)-DCs induce a distinct inflammatory T helper type 2 (Th2)-cell subset that particularly produces high levels of interleukin (IL)-13 and tumor necrosis factor-α (TNF-α). This subset expressed homing receptors for skin and inflammatory sites, and was mainly induced by B220(-)CD8α(-)CD11b(+)CD103(-) MLN-DCs in an IL-6- and OX40 ligand-dependent manner, whereas RA inhibited this induction. The corresponding MLN-DC subset of VA-sufficient mice induced a similar T-cell subset in the presence of RA receptor antagonists. IL-6 induced this subset differentiation from naive CD4(+) T cells upon activation with antibodies against CD3 and CD28. Transforming growth factor-β inhibited this induction, and reciprocally enhanced Th17 induction. Treatment with an agonistic anti-OX40 antibody and normal MLN-DCs enhanced the induction of general inflammatory Th2 cells. In VA-deficient mice, proximal colon epithelial cells produced TNF-α that may have enhanced OX40 ligand expression in MLN-DCs. The repeated oral administrations of a T cell-dependent antigen primed VA-deficient mice for IL-13-dependent strong immunoglobulin G1 (IgG1) responses and IgE responses that caused skin allergy. These results suggest that RA inhibits allergic responses to oral antigens by preventing MLN-DCs from inducing IL-13-producing inflammatory Th2 cells.

Topics: Administration, Oral; Animals; Antigens; CD40 Ligand; Cell Differentiation; Colon; Cytokines; Dendritic Cells; Immunoglobulin Isotypes; Immunophenotyping; Inflammation; Inflammation Mediators; Interleukin-13; Intestinal Mucosa; Lymph Nodes; Mesentery; Mice; Phenotype; Receptors, Retinoic Acid; Signal Transduction; T-Lymphocyte Subsets; Th17 Cells; Th2 Cells; Tretinoin; Tumor Necrosis Factor-alpha; Vitamin A Deficiency

There is increasing evidence that vitamin A deficiency in utero correlates with abnormal airway smooth muscle (SM) function in postnatal life. The bioactive vitamin A metabolite retinoic acid (RA) is essential for formation of the lung primordium; however, little is known about the impact of early fetal RA deficiency on postnatal lung structure and function. Here, we provide evidence that during murine lung development, endogenous RA has a key role in restricting the airway SM differentiation program during airway formation. Using murine models of pharmacological, genetic, and dietary vitamin A/RA deficiency, we found that disruption of RA signaling during embryonic development consistently resulted in an altered airway SM phenotype with markedly increased expression of SM markers. The aberrant phenotype persisted postnatally regardless of the adult vitamin A status and manifested as structural changes in the bronchial SM and hyperresponsiveness of the airway without evidence of inflammation. Our data reveal a role for endogenous RA signaling in restricting SM differentiation and preventing precocious and excessive SM differentiation when airways are forming.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoconstrictor Agents; Cell Differentiation; Diet; Disease Models, Animal; Female; Lung; Methacholine Chloride; Mice; Mice, Knockout; Muscle, Smooth; Oligonucleotide Array Sequence Analysis; Phenotype; Pregnancy; Signal Transduction; Tretinoin; Vitamin A; Vitamin A Deficiency

Vitamin A deficiency (VAD) is a leading cause of pediatric morbidity and mortality due to infectious diseases. Recent pre-clinical studies have revealed that VAD impairs mucosal IgA-producing antibody forming cell (AFC) responses toward a paramyxovirus vaccine in the upper respiratory tract (URT), thus impeding a first line of defense at the pathogen's point-of-entry. The studies described here tested the hypothesis that VAD may also impair immune responses after FluMist vaccinations. Results show that (i) IgA-producing antibody forming cells (AFCs) are significantly reduced following FluMist vaccination in VAD mice, and (ii) oral doses of either retinyl palmitate or retinoic acid administered on days 0, 3, and 7 relative to vaccination rescue the response. Data encourage the conduct of clinical studies to determine if there are FluMist vaccine weaknesses in human VAD populations and to test corrective supplementation strategies. Improvements in vaccine efficacy may ultimately reduce the morbidity and mortality caused by influenza virus worldwide.

Topics: Administration, Intranasal; Animals; Antibody-Producing Cells; Diterpenes; Female; Immunity, Mucosal; Immunoglobulin A; Influenza Vaccines; Mice; Mice, Inbred C57BL; Pregnancy; Retinyl Esters; Tretinoin; Vaccination; Vitamin A; Vitamin A Deficiency

Retinoic acid (RA) is an active metabolite of vitamin A and plays important roles in embryonic development. CYP26 enzymes degrade RA and have specific expression patterns that produce a RA gradient, which regulates the patterning of various structures in the embryo. However, it has not been addressed whether a RA gradient also exists and functions in organs after birth. We found localized RA activities in the diaphyseal portion of the growth plate cartilage were associated with the specific expression of Cyp26b1 in the epiphyseal portion in juvenile mice. To disturb the distribution of RA, we generated mice lacking Cyp26b1 specifically in chondrocytes (Cyp26b1(Δchon) cKO). These mice showed reduced skeletal growth in the juvenile stage. Additionally, their growth plate cartilage showed decreased proliferation rates of proliferative chondrocytes, which was associated with a reduced height in the zone of proliferative chondrocytes, and closed focally by four weeks of age, while wild-type mouse growth plates never closed. Feeding the Cyp26b1 cKO mice a vitamin A-deficient diet partially reversed these abnormalities of the growth plate cartilage. These results collectively suggest that Cyp26b1 in the growth plate regulates the proliferation rates of chondrocytes and is responsible for the normal function of the growth plate and growing bones in juvenile mice, probably by limiting the RA distribution in the growth plate proliferating zone.

Topics: Animals; Bone Development; Cell Proliferation; Chondrocytes; Cytochrome P-450 Enzyme System; Growth Plate; Mice; Mice, Knockout; Mice, Transgenic; Retinoic Acid 4-Hydroxylase; Tretinoin; Vitamin A Deficiency

The vitamin A metabolite retinoic acid is important for the function of the adaptive immune system, but the mechanism is not completely understood. Here we show that vitamin A is essential for the generation of Notch-dependent CD8(-) dendritic cells (DCs) in the spleen. We observed that CD8(-) CD4(-) (double negative (DN)) and CD4(+) DCs, but not CD8(+) DCs, express vitamin A regulated genes. To determine whether vitamin A levels influence splenic DC development, we generated mice that were fed a vitamin A-deficient diet. We detected a specific reduction of CD4(+) and DN DCs in the spleens of mice fed a vitamin A-deficient diet, while pre-DC numbers in both spleen and bone marrow were not affected. Vitamin A was specifically necessary for the development of RelB(high) , Notch-dependent CD4(+) , and DN DCs. In addition, DN DCs showed reduced proliferation during vitamin A deficiency. In contrast, mice that had received a diet with increased amounts of retinoic acid showed a significant expansion of Notch-dependent DN DCs. These data demonstrate that vitamin A stimulates the development of Notch-dependent splenic DCs and indicate that inefficient generation of DCs may contribute to the immune deficits observed during vitamin A deficiency.

Topics: Animals; CD4 Antigens; CD8 Antigens; Cell Differentiation; Cell Lineage; Cell Proliferation; Dendritic Cells; Feeding Behavior; Female; Mice; Mice, Inbred C57BL; Pregnancy; Receptors, Notch; Spleen; Transcription Factor RelB; Tretinoin; Vitamin A Deficiency

The retinaldehyde dehydrogenase 3 (Raldh3) gene encodes a major retinoic acid synthesizing enzyme and is highly expressed in the inner ear during embryogenesis. We found that mice deficient in Raldh3 bear severe impairment in vestibular functions. These mutant mice exhibited spontaneous circling/tilted behaviors and performed poorly in several vestibular-motor function tests. In addition, video-oculography revealed a complete loss of the maculo-ocular reflex and a significant reduction in the horizontal angular vestibulo-ocular reflex, indicating that detection of both linear acceleration and angular rotation were compromised in the mutants. Consistent with these behavioral and functional deficiencies, morphological anomalies, characterized by a smaller vestibular organ with thinner semicircular canals and a significant reduction in the number of otoconia in the saccule and the utricle, were consistently observed in the Raldh3 mutants. The loss of otoconia in the mutants may be attributed, at least in part, to significantly reduced expression of Otop1, which encodes a protein known to be involved in calcium regulation in the otolithic organs. Our data thus reveal a previously unrecognized role of Raldh3 in structural and functional development of the vestibular end organs.

Topics: Aldehyde Dehydrogenase 1 Family; Analysis of Variance; Animals; Behavioral Symptoms; Embryo, Mammalian; Eye Movements; Female; Gene Expression Regulation, Developmental; Imaging, Three-Dimensional; Isoenzymes; Male; Membrane Proteins; Mice; Mice, Knockout; Microscopy, Electron, Transmission; Motor Activity; Mutation; Otolithic Membrane; Pregnancy; Prenatal Exposure Delayed Effects; Reflex, Vestibulo-Ocular; Retinal Dehydrogenase; Swimming; Tretinoin; Vestibular Function Tests; Vestibule, Labyrinth; Video Recording; Vitamin A Deficiency; Walking

We have recently shown that a combination of vitamin A (VA) and retinoic acid (RA) in a 10:1 molar ratio (VARA) synergistically increases lung retinoid content in newborn rodents, more than either VA or RA alone in equimolar amounts. We hypothesized that the increase in lung retinoids would reduce oxidative stress and proinflammatory cytokines, resulting in attenuation of alveolar simplification and abnormal lung function in hyperoxia-exposed newborn mice. Newborn C57BL/6 mice were exposed to 85% O₂ (hyperoxia) or air (normoxia) for 7 or 14 days from birth and given vehicle or VARA every other day. Lung retinol content was measured by HPLC, function was assessed by flexiVent, and development was evaluated by radial alveolar counts, mean linear intercept, and secondary septal crest density. Mediators of oxidative stress, inflammation, and alveolar development were evaluated in lung homogenates. We observed that VARA increased lung retinol stores and attenuated hyperoxia-induced alveolar simplification while increasing lung compliance and lowering resistance. VARA attenuated hyperoxia-induced increases in DNA damage and protein oxidation accompanied with a reduction in nuclear factor (erythroid-derived 2)-like 2 protein but did not alter malondialdehyde adducts, nitrotyrosine, or myeloperoxidase concentrations. Interferon-γ and macrophage inflammatory protein-2α mRNA and protein increased with hyperoxia, and this increase was attenuated by VARA. Our study suggests that the VARA combination may be a potential therapeutic strategy in conditions characterized by VA deficiency and hyperoxia-induced lung injury during lung development, such as bronchopulmonary dysplasia in preterm infants.

Topics: Animals; Animals, Newborn; Female; Hyperoxia; Inflammation Mediators; Lung; Lung Compliance; Mice; Mice, Inbred C57BL; Oxidative Stress; Platelet-Derived Growth Factor; Pulmonary Alveoli; RNA, Messenger; Tenascin; Tretinoin; Vitamin A; Vitamin A Deficiency

Vitamin A and its metabolite retinoic acid influence various aspects of immunity. Although the capacity of vitamin A to condition intestinal CD103(+) DCs to imprint tissue-specific homing programs onto activated lymphocytes is well documented, it is unclear whether vitamin A also regulates DC populations in other tissues. A study published in this issue of the European Journal of Immunology, Beijer et al. [Eur. J. Immunol. 2013. 43: 1608-1616] now demonstrates that vitamin A exerts profound effects on the subset composition of splenic DCs. By resolving that splenic ESAM(hi) CD11b(hi) DCs are preferentially responsive to regulation by vitamin A, these novel insights not only further support the notion that ESAM expression marks two distinct lineages of splenic CD11b(hi) DCs, but also provide an important extension to our understanding of how vitamin A influences the immune system.

Topics: Animals; Dendritic Cells; Female; Pregnancy; Tretinoin; Vitamin A Deficiency

The aim of this study was to explore the involvement of retinoids in the hypoactivity and hyporeactivity to stress of the hypothalamic-pituitary-adrenal (HPA) axis in LOU/C rats. We measured the effects of vitamin A deficiency administered or not with retinoic acid (RA) on plasma corticosterone in standard conditions and in response to restraint stress and on hypothalamic and hippocampal expression of corticosteroid receptors, corticotropin-releasing hormone and 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) in LOU/C rats. Interestingly, under control conditions, we measured a higher plasma concentration of retinol in LOU/C than in Wistar rats, which could contribute to the lower basal activity of the HPA axis in LOU/C rats. Vitamin A deficiency induced an increased HPA axis activity in LOU/C rats, normalized by RA administration. Compared with LOU/C control rats, vitamin A-deficient rats showed a delayed and heightened corticosterone response to restraint stress. The expression of corticosteroid receptors was strongly decreased by vitamin A deficiency in the hippocampus, which could contribute to a less efficient feedback by corticosterone on HPA axis tone. The expression of 11β-HSD1 was increased by vitamin A deficiency in the hypothalamus (+62.5%) as in the hippocampus (+104.7%), which could lead to a higher production of corticosterone locally and contribute to alteration of the hippocampus. RA supplementation treatment restored corticosterone concentrations and 11β-HSD1 expression to control levels. The high vitamin A status of LOU/C rats could contribute to their low HPA axis activity/reactivity and to a protective effect against 11β-HSD1-mediated deleterious action on cognitive performances during ageing.

Topics: 11-beta-Hydroxysteroid Dehydrogenase Type 1; Animals; Corticosterone; Corticotropin-Releasing Hormone; Hypothalamo-Hypophyseal System; Male; Pituitary-Adrenal System; Rats; Rats, Inbred Strains; Rats, Wistar; Stress, Physiological; Stress, Psychological; Tretinoin; Vitamin A; Vitamin A Deficiency

We investigated the effect of β-carotene (bC) supplementation during pregnancy in a mouse model of severe vitamin A deficiency, i.e. Lrat-/-Rbp-/- dams maintained on a vitamin A-deficient diet during gestation. bC, a provitamin A carotenoid, can be enzymatically cleaved to form vitamin A for use by the developing embryo. We found that an acute supplementation (13.5 days post coitum, dpc) of bC to Lrat-/-Rbp-/- dams on a vitamin A-deficient diet activated transcriptional mechanisms in the developing tissues to maximize the utilization of bC provided to the dams. Nevertheless, these regulatory mechanisms are inefficient under this regimen, as the embryonic phenotype was not improved. We further investigated the effect of a repeated supplementation of bC during a crucial developmental period (6.5-9.5 dpc) on the above-mentioned mouse model. This treatment improved the embryonic abnormalities, as 40% of the embryos showed a normal phenotype. In addition, analysis of retinoic acid-responsive genes, such as Cyp26a1 in these embryos suggests that bC cleavage results in the production of retinoic acid which then can be used by the embryo. Taken together, these in vivo studies show that bC can be used as a source of vitamin A for severely vitamin A-deficient mammalian embryos.

Topics: Animals; beta Carotene; Cytochrome P-450 Enzyme System; Disease Models, Animal; Female; Gene Expression Regulation, Developmental; Maternal-Fetal Exchange; Mice; Phenotype; Pregnancy; Retinoic Acid 4-Hydroxylase; Retinoids; Severity of Illness Index; Tretinoin; Vitamin A Deficiency

The blood testis-barrier (BTB) is essential for maintaining homeostasis in the seminiferous epithelium. Although many studies have reported that vitamin A (VA) is required for the maintenance of spermatogenesis, the relationships between the BTB, spermatogenesis and VA have not been elucidated. In this study, we analyzed BTB assembly and spermatogenesis in the testes of mice fed the VA-deficient (VAD) diet from the prepubertal period to adulthood. During the prepubertal period, no changes were observed in the initiation and progression of the first spermatogenic wave in mice fed the VAD diet. However, the numbers of preleptotene/leptotene spermatocytes derived from the second spermatogenic wave onwards were decreased, and initial BTB formation was also delayed, as evidenced by the decreased expression of mRNAs encoding BTB components and VA signaling molecules. From 60 days postpartum, mice fed the VAD diet exhibited apoptosis of germ cells, arrest of meiosis, disruption of the BTB, and dramatically decreased testis size. Furthermore, vacuolization and calcification were observed in the seminiferous epithelium of adult mice fed the VAD diet. Re-initiation of spermatogenesis by VA replenishment in adult mice fed the VAD diet rescued BTB assembly after when the second spermatogenic wave initiated from the arrested spermatogonia reached the preleptotene/leptotene spermatocytes. These results suggested that BTB integrity was regulated by VA metabolism with meiotic progression and that the impermeable BTB was required for persistent spermatogenesis rather than meiotic initiation. In conclusion, consumption of the VAD diet led to critical defects in spermatogenesis progression and altered the dynamics of BTB assembly.

Topics: Animals; Apoptosis; Biomarkers; Blood-Testis Barrier; Calcinosis; Diet; Disease Models, Animal; Epididymis; Female; Gene Expression Regulation, Developmental; Infertility, Male; Male; Metaplasia; Mice; Mice, Inbred C57BL; Organ Size; Spermatogenesis; Spermatogonia; Testis; Tretinoin; Vacuoles; Vitamin A Deficiency

Vitamin A is essential for lung development and pulmonary cell differentiation. Its deficiency leads to altered lung structure and function and to basement membrane architecture and composition disturbances. Previously, we showed that lack of retinoids thickens the alveolar basement membrane and increases collagen IV, which are reversed by retinoic acid, the main biologically active vitamin A form. This study analyzed how vitamin A deficiency affects the subunit composition of collagen IV and laminin of lung basement membranes and pulmonary matrix metalloproteinase content, plus the recovering effect of all-trans-retinoic acid. Male weanling pups were fed a retinol-adequate/-deficient diet until 60 days old. A subgroup of vitamin-A-deficient pups received daily intraperitoneal all-trans-retinoic acid injections for 10 days. Collagen IV and laminin chain composition were modified in vitamin-A-deficient rats. The protein and mRNA contents of chains α1(IV), α3(IV) and α4(IV) increased; those of chains α2(IV) and α5(IV) remained unchanged; and the protein and mRNA contents of laminin chains α5, β1 and γ1 decreased. The mRNA of laminin chains α2 and α4 also decreased. Matrix metalloproteinases 2 and 9 decreased, but the tissue inhibitors of metalloproteinases 1 and 2 did not change. Treating vitamin-A-deficient rats with retinoic acid reversed all alterations, but laminin chains α2, α4 and α5 and matrix metalloproteinase 2 remained low. In conclusion, vitamin A deficiency alters the subunit composition of collagen IV and laminin and the lung's proteolytic potential, which are partly reverted by retinoic acid. These alterations could contribute to impaired lung function and predispose to pulmonary disease.

Topics: Animals; Basement Membrane; Collagen Type IV; Female; Gene Expression; Laminin; Lung; Male; Matrix Metalloproteinases; Rats; Rats, Wistar; Tissue Inhibitor of Metalloproteinases; Tretinoin; Vitamin A; Vitamin A Deficiency

Vitamin A (VA) plays an important role in post-natal lung development and maturation. Previously, we have reported that a supplemental dose of VA combined with 10% of all-trans-retinoic acid (VARA) synergistically increases retinol uptake and retinyl ester (RE) storage in neonatal rat lung, while up-regulating several retinoid homeostatic genes including lecithin:retinol acyltransferase (LRAT) and the retinol-binding protein receptor, stimulated by retinoic acid 6 (STRA6). However, whether inflammation has an impact on the expression of these genes and thus compromises the ability of VARA to increase lung RE content is not clear. Neonatal rats, 7- to 8-d-old, were treated with VARA either concurrently with lipopolysaccharide (LPS; Expt 1) or 12 h after LPS administration (Expt 2); in both studies, lung tissue was collected 6 h after VARA treatment, when RE formation is maximal. Inflammation was confirmed by increased IL-6 and chemokine (C–C motif) ligand 2 (CCL2) gene expression in lung at 6 h and C-reactive protein in plasma at 18 h. In both studies, LPS-induced inflammation only slightly reduced, but did not prevent the VARA-induced increase in lung RE. Quantitative RT-PCR showed that co-administration of LPS with VARA slightly attenuated the VARA-induced increase of LRAT mRNA, but not of STRA6 or cytochrome P450 26B1, the predominant RA hydroxylase in lung. By 18 h post-LPS, expression had subsided and none of these genes differed from the level in the control group. Overall, the present results suggest that retinoid homeostatic gene expression is reduced modestly, if at all, by acute LPS-induced inflammation and that VARA is still effective in increasing lung RE under conditions of moderate inflammation.

Topics: Acyltransferases; Animals; Animals, Newborn; C-Reactive Protein; Chemokine CCL2; Cytochrome P-450 Enzyme System; Esters; Gene Expression; Inflammation; Interleukin-6; Lipopolysaccharides; Lung; Membrane Proteins; Rats; Receptors, Cell Surface; Retinoids; RNA, Messenger; Tretinoin; Vitamin A; Vitamin A Deficiency

Primary cultured retinal pigment epithelial (RPE) cells can convert T cells into T regulatory cells (Tregs) through inhibitory factor(s) including transforming growth factor β (TGFβ) in vitro. Retinoic acid (RA) enhances induction of CD4(+) Tregs in the presence of TGFβ. We investigated whether RA produced by RPE cells can promote generation of Tregs. We found that in vitro, RA-treated T cells expressed high levels of Foxp3 in the presence of recombinant TGFβ. In GeneChip analysis, cultured RPE cells constitutively expressed RA-associated molecules such as RA-binding proteins, enzymes, and receptors. RPE from normal mice, but not vitamin A-deficient mice, contained significant levels of TGFβ. RPE-induced Tregs from vitamin A-deficient mice failed to suppress activation of target T cells. Only a few Foxp3(+) T cells were found in intraocular cells from vitamin A-deficient experimental autoimmune uveitis (EAU) mice, whereas expression was higher in cells from normal EAU mice. RA receptor antagonist-pretreated or RA-binding protein-siRNA-transfected RPE cells failed to convert CD4(+) T cells into Tregs. Our data support the hypothesis that RPE cells produce RA, thereby enabling bystander T cells to be converted into Tregs through TGFβ promotion, which can then participate in the establishment of immune tolerance in the eye.

Topics: Animals; Autoimmune Diseases; CD4-Positive T-Lymphocytes; Cell Proliferation; Cells, Cultured; Coculture Techniques; Enzyme-Linked Immunosorbent Assay; Female; Flow Cytometry; Forkhead Transcription Factors; Lymphocyte Activation; Mice; Mice, Inbred C57BL; Oligonucleotide Array Sequence Analysis; Pregnancy; Real-Time Polymerase Chain Reaction; Receptors, Retinoic Acid; Retinal Pigment Epithelium; RNA, Messenger; RNA, Small Interfering; T-Lymphocytes, Regulatory; Transfection; Transforming Growth Factor beta; Tretinoin; Uveitis; Vitamin A; Vitamin A Deficiency

Fetal alcohol spectrum disorder (FASD) is an umbrella term used to describe the craniofacial dysmorphic features, malformations, and disturbances in growth, neurodevelopment and behavior occurring in individuals prenatally exposed to alcohol. Fetal alcohol syndrome (FAS) represents the severe end of this spectrum. Many pathophysiological mechanisms have hitherto been proposed to account for the disrupted growth and morphogenesis seen in FAS. These include impaired cholesterol-modification of the Sonic hedgehog morphogen, retinoic acid deficiency, lipoperoxidative damage due to alcohol-induced reactive oxygen species combined with reduced antioxidant defences, and malfunctioning cell adhesion molecules. In this report, we propose a completely novel concept regarding the pathogenesis of FAS. Based on our observation that transferrin isoelectric focusing (TIEF) - the most widely used screening tool for congenital disorders of glycosylation (CDG) - was transiently abnormal in a newborn with FAS and a confirmed maternal history of gestational alcohol abuse, we came to believe that FAS exemplifies a congenital disorder of glycosylation secondary to alcohol-inflicted disruption of (N-linked) protein glycosylation. Various pieces of evidence were found in the literature to substantiate this hypothesis. This observation implies, among others, that one might need to consider the possibility of maternal alcohol consumption in newborns with transient glycosylation abnormalities. We also present an integrated pathophysiological model of FAS, which incorporates all existing theories mentioned above as well as our novel concept. This model highlights the pivotal role of disrupted isoprenoid metabolism in the origination of FAS.

Topics: Alcohol Drinking; Alcoholism; Antioxidants; Cholesterol; Dolichols; False Positive Reactions; Female; Fetal Alcohol Spectrum Disorders; Glycosylation; Hedgehog Proteins; Humans; Infant; Infant, Newborn; Isoelectric Focusing; Male; Models, Theoretical; Pregnancy; Reactive Oxygen Species; Transferrin; Tretinoin; Vitamin A Deficiency

Vitamin A (VA) and its active form, retinoic acid (RA), are regulators of skeletal development. In the present study, we investigated if maternal VA intake during pregnancy and lactation, as well as direct oral supplementation of neonates with VA + RA (VARA) in early life, alters neonatal bone formation and chondrocyte gene expression. Offspring of dams fed 3 levels of VA (marginal, adequate, and supplemented) for 10 wk were studied at birth (P0) and postnatal day 7 (P7). One-half of the newborns received an oral supplement of VARA on P1, P4, and P7. Tissues were collected on P0 and 6 h after the last dose on P7. Pup plasma and liver retinol concentrations were increased by both maternal VA intake and VARA (P < 0.01). Although maternal VA did not affect bone mineralization as assessed by von Kossa staining, newborn femur length was increased with maternal VA (P < 0.05). VARA supplementation of neonates increased the length of the hypertrophic zone only in VA-marginal pups, close to that in neonates from VA-adequate dams, suggesting VARA caused a catching up of growth that was limited by low maternal VA intake. Maternal diet did not alter type X nor type II collagen mRNA. However, VARA-treated pups from VA-supplemented dams had reduced mRNA for aggrecan, a major component of cartilage matrix, and increased mRNA for matrix metalloproteinase (MMP)13, which catalyzes the degradation of aggrecan and collagens. These results suggest that moderately high maternal VA intake combined with neonatal VARA supplementation can reduce the ratio of aggrecan:MMP, which may unfavorably alter early bone development.

Topics: Aggrecans; Animals; Animals, Newborn; Chondrocytes; Dietary Supplements; Female; Femur; Gene Expression Regulation, Developmental; Growth Plate; Lactation; Liver; Maternal Nutritional Physiological Phenomena; Matrix Metalloproteinase 13; Osteogenesis; Pregnancy; Rats; Rats, Sprague-Dawley; Tretinoin; Vitamin A; Vitamin A Deficiency; Weaning

Iron bioavailability seems to be regulated by vitamin A (VA) but the molecular events involved in this mechanism are not well understood. It is also known that retinoids mediate most of their function via interaction with retinoid receptors, which act as ligand-activated transcription factors controlling the expression of a number of target genes. Here, we evaluated the VA effects on the modulation of the levels of mRNA encoding proteins involved in the iron bioavailability, whether in the intestinal absorption process or in the liver iron metabolism. The expression of genes involved in iron intestinal absorption (divalent metal transporter 1, duodenal cytochrome B, ferroportin 1 FPN1, and ferritin) were evaluated in vitro by treating Caco-2 cells with retinoic acid or in vivo by observing the effects of vitamin A deficiency (VAD) in BALB/C mice. Liver hepcidin and ferritin mRNA levels were upregulated by VAD; however, this condition did not promote any change on the expression of those genes that participate in the iron absorption. Moreover, data from the in vitro analysis showed that VA induced FPN1 gene expression by a hepcidin-independent manner. Therefore, the in vivo results support the idea that VAD may not affect iron absorption but would rather affect iron mobilization mechanisms. On the other hand, our results using Caco-2 cells raises the possibility that VA addition to intestinal epithelium may improve iron absorption through the induction of FPN1 gene expression.

Topics: Animals; Antimicrobial Cationic Peptides; Blotting, Western; Caco-2 Cells; Cation Transport Proteins; Dietary Supplements; Diterpenes; Drug Evaluation, Preclinical; Duodenum; Ferritins; Gene Expression Regulation; Hepcidins; Humans; Intestinal Absorption; Intestinal Mucosa; Iron; Liver; Male; Mice; Mice, Inbred BALB C; Retinyl Esters; RNA, Messenger; Tretinoin; Vitamin A; Vitamin A Deficiency

Retinaldehyde dehydrogenases (RALDH) catalyze the synthesis of the regulatory factor retinoic acid (RA). Cultured astrocytes express several of the RALDH enzyme family, and it has been assumed that this can be extrapolated to astrocytes in vivo. However, this study finds that few astrocytes in the rodent brain express detectable RALDH enzymes, and only when these cells are grown in culture are these enzymes upregulated. Factors controlling the expression of the RALDHs in cultured astrocytes were explored to determine possible reasons for differences between in vitro versus in vivo expression. Retinoids were found to feedback to suppress several of the RALDHs, and physiological levels of retinoids may be one route by which astrocytic RALDHs are maintained at low levels. In the case of RALDH2, in vivo reduction of vitamin A levels in rats resulted in an increase in astrocyte RALDH2 expression in the hippocampus. Other factors though are likely to control RALDH expression. A shift in astrocytic RALDH subcellular localization is a potential mechanism for regulating RA signaling. Under conditions of vitamin A deficiency, RALDH2 protein moved from the cytoplasm to the nucleus where it may synthesize RA at the site of the nuclear RA receptors. Similarly, in conditions of oxidative stress RALDH1 and RALDH2 moved from the cytoplasm to a predominantly nuclear position. Thus, the RALDHs have been revealed to be dynamic in their expression in astrocytes where they may maintain retinoid homeostasis in the brain.

Topics: Aldehyde Dehydrogenase 1 Family; Animals; Animals, Newborn; Astrocytes; Brain; Cells, Cultured; Humans; Male; Mice; Mice, Transgenic; Middle Aged; Rats; Rats, Sprague-Dawley; Retinal Dehydrogenase; Tretinoin; Vitamin A Deficiency

Vitamin A deficiency causes impaired vision and blindness in millions of children around the world. Previous studies in zebrafish have demonstrated that retinoic acid (RA), the acid form of vitamin A, plays a vital role in early eye development. The objective of this study was to describe the effects of early RA deficiency by treating zebrafish with diethylaminobenzaldehyde (DEAB), a potent inhibitor of the enzyme retinaldehyde dehydrogenase (RALDH) that converts retinal to RA. Zebrafish embryos were treated for 2 h beginning at 9 h postfertilization. Gross morphology and retinal development were examined at regular intervals for 5 days after treatment. The optokinetic reflex (OKR) test, visual background adaptation (VBA) test, and the electroretinogram (ERG) were performed to assess visual function and behavior. Early treatment of zebrafish embryos with 100 μM DEAB (9 h) resulted in reduced eye size, and this microphthalmia persisted through larval development. Retinal histology revealed that DEAB eyes had significant developmental abnormalities but had relatively normal retinal lamination by 5.5 days postfertilization. However, the fish showed neither an OKR nor a VBA response. Further, the retina did not respond to light as measured by the ERG. We conclude that early deficiency of RA during eye development causes microphthalmia as well as other visual defects, and that timing of the RA deficiency is critical to the developmental outcome.

Topics: Adaptation, Ocular; Animals; Behavior, Animal; Electroretinography; Embryo, Nonmammalian; Eye; Larva; Microphthalmos; Nystagmus, Optokinetic; p-Aminoazobenzene; Phenotype; Reflex; Tretinoin; Vitamin A Deficiency; Zebrafish

α(4) and β(7) integrins, such as α(4)β(1), α(4)β(7), and α(E)β(7), are major integrins required for migration of leukocytes into mucosal tissues. The mechanisms responsible for coordinated expression of these three integrins have been poorly elucidated to date. We report that expression of the Itg-α(4) subunit by both CD4(+) and CD8(+) T cells requires the retinoic acid signal. In contrast, transcription of Itg-α(E) genes is induced by the transforming growth factor-β1 (TGFβ1) signal. Expression of Itg-β(7) is constitutive but can be further increased by TGFβ1. Consistently, expression of α(4)-containing integrins is severely suppressed in vitamin A deficiency with a compensatory increase of α(E)β(7), whereas expression of Itg-α(E) and Itg-β(7) is decreased in TGFβ-signal deficiency with a compensatory increase in α(4)β(1). The retinoic acid-mediated regulation of α(4) integrins is required for specific migration of T cells in vitro and in vivo. These results provide central regulatory mechanisms for coordinated expression of the major mucosal integrins.

Topics: Animals; Base Sequence; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Cells, Cultured; Flow Cytometry; Forkhead Transcription Factors; Gene Expression Regulation; Humans; Immunoprecipitation; Integrin alpha4; Integrins; Lymphocyte Activation; Mice; Mucous Membrane; Oligonucleotide Array Sequence Analysis; Signal Transduction; T-Lymphocytes; Transforming Growth Factor beta1; Tretinoin; Vitamin A Deficiency

The vitamin A metabolite retinoic acid (RA) plays a crucial role in mucosal immune responses. We demonstrate in this study that RA-producing retinaldehyde dehydrogenase (RALDH) enzymes are postnatally induced in mesenteric lymph node (MLN) dendritic cells (DCs) and MLN stromal cells. RALDH enzyme activity in lamina propria-derived CD103(+) MLN-DCs did not depend on TLR signaling. Remarkably, RA itself could directly induce RALDH2 in both DCs and stromal cells in vitro. Furthermore, upon provision of a vitamin A-deficient diet, it was found that RA-mediated signaling was strongly reduced within the small intestines, while RALDH2 mRNA and RALDH enzyme activity in lamina propria DCs and MLN-DCs, as well as RALDH2 mRNA expression in MLN stromal cells, were strongly diminished. Moreover, supply of vitamin A to vitamin A-deficient mice restored RA-mediated signaling in the intestine and RALDH activity in lamina propria-derived CD103(+) MLN-DCs. Our results show that RA-dependent signaling within the intestine is indispensable for RALDH activity in the draining MLN.

Topics: Aldehyde Oxidoreductases; Animal Feed; Animals; Dendritic Cells; Gene Expression Regulation; Intestinal Mucosa; Intestine, Small; Lymph Nodes; Mesentery; Mice; Retinal Dehydrogenase; Stromal Cells; Tretinoin; Vitamin A; Vitamin A Deficiency

RPE65 gene knockout (Rpe65⁻/⁻) mice showed abolished isomerohydrolase activity in the visual cycle and were considered a model for vitamin A deficiency in the retina. The purpose of this study was to compare the retinal phenotypes between vitamin A-deficient (VAD) mice and Rpe65⁻/⁻ mice under normal diet.. The VAD mice were fed with a vitamin A-deprived diet after birth. The age-matched control mice and Rpe65⁻/⁻ mice were maintained under normal diet. The structure of photoreceptor outer segment was compared using electron microscopy. Photoreceptor-specific gene expression was determined using real-time RT-PCR. The isomerohydrolase and lecithin-retinol acyltransferase (LRAT) activities were measured using an in vitro enzymatic activity assay. Endogenous retinoid profiles were analyzed by HPLC in mouse eyecup homogenates.. Compared to wild-type mice under normal diet, scanning and transmission electron microscopy showed that the outer segments of photoreceptors were disorganized in VAD mice and were not disorganized in Rpe65⁻/⁻ mice, although they were shortened in the latter. VAD mice showed more prominent downregulation of middle wavelength cone opsin, whereas Rpe65⁻/⁻ mice displayed more suppressed expression of short wavelength cone opsin. In vitro enzymatic activity assay and Western blot analysis showed that vitamin A deprivation downregulated LRAT expression and activity in the eyecup, but Rpe65⁻/⁻ mice showed unchanged LRAT expression and activity. The depressed LRAT activity in VAD mice was partially rescued by the intraperitoneal injection of retinoic acid.. VAD and Rpe65⁻/⁻ mice are different in cone photoreceptor degeneration, photoreceptor-specific gene regulation, isomerohydrolase activity, endogenous retinoid profile, and LRAT activity.

Topics: Acyltransferases; Animals; Blotting, Western; Carrier Proteins; Cell Count; Chromatography, High Pressure Liquid; cis-trans-Isomerases; Eye Proteins; Gene Expression; Injections, Intraperitoneal; Mice; Mice, Knockout; Microscopy, Electron, Scanning; Opsins; Phenotype; Photoreceptor Cells, Vertebrate; Retinal Degeneration; Retinal Photoreceptor Cell Outer Segment; Retinal Pigment Epithelium; Reverse Transcriptase Polymerase Chain Reaction; Tretinoin; Vitamin A; Vitamin A Deficiency

Numerous wrinkles are observed in the skin of streptozotocin (STZ)-induced type 1 diabetic rats, which are similar to those seen in vitamin A-deficient (VAD) rats. Retinoic acid (RA), the active form of vitamin A, promotes the production of collagen in dermis and induces cell growth and inhibition of epidermal differentiation in skin tissues. Normal skin function is maintained by the extracellular matrix (ECM)-degrading enzymes, matrix metalloproteinase (MMP) and hyaluronidase (HAase). This study is the first comparison of MMP and HAase activities in skin tissues of type 1 diabetic, VAD and RA-treated animal models. In skin tissues of type 1 diabetic and VAD rats or VAD mice, both MMP-2 and HAase activities increased as compared with controls. In contrast, MMP and HAase activities were reduced in the skin tissues of RA-treated mice. Blood retinol levels in type 1 diabetic rats were lower than controls. These results indicate a close relationship between type 1 diabetes and vitamin A-deficiency on MMP and HAase in skin tissues, suggesting that type 1 diabetic rats could be vitamin A-deficient. Vitamin A-derived RA might be a significant regulator of ECM-degrading enzyme expression and diabetic symptoms.

Topics: Aging; Animals; Diabetes Mellitus, Type 1; Extracellular Matrix; Hyaluronoglucosaminidase; Male; Matrix Metalloproteinases; Mice; Mice, Hairless; Rats; Rats, Wistar; Skin; Skin Aging; Tretinoin; Vitamin A Deficiency

Vitamin A is essential for lung development and pulmonary cell differentiation and its deficiency results in alterations of lung structure and function. Basement membranes (BMs) are also involved in those processes, and retinoic acid, the main biologically active form of vitamin A, influences the expression of extracellular matrix macromolecules. Therefore, we have analyzed the ultrastructure and collagen content of lung alveolar BM in growing rats deficient in vitamin A and the recovering effect of all-trans retinoic acid. Male weanling pups were fed a retinol-adequate or -deficient diet until they were 60 days old. A group of vitamin A-deficient pups were recovered by daily intraperitoneal injections of all-trans retinoic acid for 10 days. Alveolar BM in vitamin A-deficient rats doubled its thickness and contained irregularly scattered collagen fibrils. Immunocytochemistry revealed that these fibrils were composed of collagen I. Total content of both collagen I protein and its mRNA was greater in vitamin-deficient lungs. In agreement with the greater size of the BM the amount of collagen IV was also increased. Proinflammatory cytokines, IL-1alpha, IL-1beta and TNF-alpha, did not change, but myeloperoxidase and TGF-beta1 were increased. Treatment of vitamin A-deficient rats with retinoic acid reversed all the alterations, but the BM thickness recovered only partially. Retinoic acid recovering activity occurred in the presence of increasing oxidative stress. In conclusion, vitamin A deficiency results in alterations of the structure and composition of the alveolar BM which are probably mediated by TGF-beta1 and reverted by retinoic acid. These alterations could contribute to the impairment of lung function and predispose to pulmonary disease.

Topics: Animals; Basement Membrane; Collagen Type I; Collagen Type IV; Gene Expression Regulation; Immunohistochemistry; Interleukins; Lung; Male; Malondialdehyde; Oxidative Stress; Peroxidase; Protein Subunits; Pulmonary Alveoli; Rats; Rats, Wistar; RNA, Messenger; Transforming Growth Factor beta1; Tretinoin; Tumor Necrosis Factor-alpha; Vitamin A Deficiency

Retinoic acid (RA) is a crucial factor for maintaining homeostasis in the gut, including lymphocyte homing, immunoglobulin (Ig) A production, and T regulatory cells (Treg) and T helper cell 17 (T(H)17) generation. Until now, most attention has focused on the function of dendritic cells (DCs) to initiate adaptive immunity including T and B lymphocytes through RA. To investigate the effects of RA on DCs of gut-associated lymphoid tissue (GALT), we analyzed the phenotype and function of DC subsets from GALT of vitamin A-deficient (VAD) mice.. VAD mice were prepared by feeding them a VAD diet over 12 weeks from gestational days 10-14.. Here, we report that tremendous increase of langerin(+) DCs occurred in the mesenteric lymph nodes (MLNs) and gut lamina propria of VAD mice dependent on CCR7 signaling. Langerin(+) DCs have phenotypes more similar to those of bone marrow-derived dermal langerin(+) DCs than epidermal Langerhans cells. Moreover, RA receptor antagonists enhance the differentiation of langerin(+) DCs from mouse and human precursors of bone marrow and peripheral blood. Langerin(+) DCs were highly differentiated but less inflammatory than langerin(-) DCs of MLNs of VAD mice. Moreover, tolerance to orally delivered antigen was completely abrogated by depletion of langerin(+) DCs in the VAD mice.. These results suggest that generation of langerin(+) DCs in the GALT is tightly regulated by RA and that the microenvironment of tissues determines the phenotype of DCs.

Topics: Aldehyde Oxidoreductases; Animals; Antigens, Surface; CD4-Positive T-Lymphocytes; Cell Differentiation; Dendritic Cells; Immune Tolerance; Intestines; Langerhans Cells; Lectins, C-Type; Lymphocyte Activation; Lymphoid Tissue; Mannose-Binding Lectins; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Receptors, CCR7; Tretinoin; Vitamin A Deficiency

Vitamin A deficiency (VAD) is associated with increased susceptibility to carcinogenesis in animal models and elevated risk for a number of human cancers. Here, we found that CYP26A1, the gene encoding a cytochrome P450 enzyme specifically involved in metabolic inactivation of retinoic acid (RA), the most active vitamin A derivative, is highly expressed in 42% (27/65) of primary breast cancers. We also showed that enhanced expression of CYP26A1 suppresses cellular responses to anoikis and consequently promotes anchorage-independent growth. This transformed phenotype was sufficient to markedly increase tumorigenic and metastatic potential. Suppression of CYP26A1 significantly reversed the CYP26A1-mediated oncogenic characteristics, suggesting a direct link between intracellular RA status and tumorigenicity. Our observations provide strong evidence for oncogenic and cell survival properties of CYP26A1 in carcinogenesis, and suggest mechanisms whereby VAD might promote cancer development.

Topics: Animals; Anoikis; Breast Neoplasms; Carcinogenicity Tests; Carcinogens; Cell Proliferation; Cell Survival; Cell Transformation, Neoplastic; Cytochrome P-450 Enzyme System; Disease Models, Animal; Drug Interactions; Gene Expression Profiling; Humans; Mice; Mice, Knockout; Neoplasms; Retinoic Acid 4-Hydroxylase; Reverse Transcriptase Polymerase Chain Reaction; Tretinoin; Tumor Cells, Cultured; Vitamin A; Vitamin A Deficiency

Hirschsprung disease is a serious disorder of enteric nervous system (ENS) development caused by the failure of ENS precursor migration into the distal bowel. We now demonstrate that retinoic acid (RA) is crucial for GDNF-induced ENS precursor migration, cell polarization and lamellipodia formation, and that vitamin A depletion causes distal bowel aganglionosis in serum retinol-binding-protein-deficient (Rbp4(-/-)) mice. Ret heterozygosity increases the incidence and severity of distal bowel aganglionosis induced by vitamin A deficiency in Rbp4(-/-) animals. Furthermore, RA reduces phosphatase and tensin homolog (Pten) accumulation in migrating cells, whereas Pten overexpression slows ENS precursor migration. Collectively, these data support the hypothesis that vitamin A deficiency is a non-genetic risk factor that increases Hirschsprung disease penetrance and expressivity, suggesting that some cases of Hirschsprung disease might be preventable by optimizing maternal nutrition.

Topics: Animals; Cell Movement; Disease Models, Animal; Embryonic Stem Cells; Enteric Nervous System; Female; Glial Cell Line-Derived Neurotrophic Factor; Heterozygote; Hirschsprung Disease; Humans; In Vitro Techniques; Maternal Nutritional Physiological Phenomena; Mice; Mice, Inbred C57BL; Mice, Knockout; Models, Biological; Pregnancy; PTEN Phosphohydrolase; Receptors, Retinoic Acid; Retinol-Binding Proteins, Plasma; Signal Transduction; Tretinoin; Vitamin A; Vitamin A Deficiency

Chronic vitamin A deficiency induces a substantial delay in the rates of weight and height gain in both humans and experimental animals. This effect has been associated with an impaired nutrient metabolism and loss of body protein. Therefore, we analyzed the effect of vitamin A deficiency on endogenous proteolysis and nitrogen metabolism and its reversibility with all-trans retinoic acid (RA). Male weanling rats, housed in pairs, were pair-fed a vitamin A-deficient (VAD) or control diet until they were 60 d old. A group of deficient rats were further treated with daily intraperitoneal injections of all-trans RA for 10 d. Final body and tissue (i.e. liver and heart) weights were significantly lower and tissue:body weight ratios were similar in VAD rats and in controls. Conversely, the epididymal white fat:body weight ratio and the plasma concentrations of alanine aminotransferase and adiponectin were significantly higher in VAD rats, which also had hepatic macrovesicular lipid accumulations. Plasma and gastrocnemius muscle 3-methylhistidine, urine nitrogen, and plasma and urine urea concentrations were all significantly higher in the VAD group. The expression of the genes encoding urea cycle enzymes and their activities increased in VAD livers. These changes were partially reverted by all-trans RA. We propose that fuel partitioning in vitamin A deficiency may shift from fatty acids to protein catabolism as an energy source. Our results emphasize the importance of vitamin A on the energy balance control system and they provide an explanation for the role of vitamin A in protein turnover, development, and growth.

Topics: Animals; Antioxidants; Enzyme Induction; Lipid Peroxidation; Liver; Male; Methylhistidines; Muscle, Skeletal; Nitrogen; Rats; Retinoids; Tretinoin; Triglycerides; Urea; Vitamin A Deficiency

Nephron number varies widely between 0.3 and 1.3 million per kidney in humans. During fetal life, the rate of nephrogenesis is influenced by local retinoic acid (RA) level such that even moderate maternal vitamin A deficiency limits the final nephron number in rodents. Inactivation of genes in the RA pathway causes renal agenesis in mice; however, the impact of retinoids on human kidney development is unknown. To resolve this, we tested for associations between variants of genes involved in RA metabolism (ALDH1A2, CYP26A1, and CYP26B1) and kidney size among normal newborns. Homozygosity for a common (1 in 5) variant, rs7169289(G), within an Sp1 transcription factor motif of the ALDH1A2 gene, showed a significant 22% increase in newborn kidney volume when adjusted for body surface area. Infants bearing this allele had higher umbilical cord blood RA levels compared to those with homozygous wild-type ALDH1A2 rs7169289(A) alleles. Furthermore, the effect of the rs7169289(G) variant was evident in subgroups with or without a previously reported hypomorphic RET 1476(A) proto-oncogene allele that is critical in determining final nephron number. As maternal vitamin A deficiency is widespread in developing countries and may compromise availability of retinol for fetal RA synthesis, our study suggests that the ALDH1A2 rs7169289(G) variant might be protective for such individuals.

Topics: Alleles; Cytochrome P-450 Enzyme System; Developing Countries; Genotype; Humans; Infant; Infant, Newborn; Kidney; Kidney Diseases; Nephrons; Organogenesis; Oxidoreductases; Proto-Oncogene Mas; Proto-Oncogenes; Retinoic Acid 4-Hydroxylase; Retinoids; Tretinoin; Vitamin A; Vitamin A Deficiency

The developmental abnormalities associated with disruption of signaling by retinoic acid (RA), the biologically active form of vitamin A, have been known for decades from studies in animal models and humans. These include defects in the respiratory system, such as lung hypoplasia and agenesis. However, the molecular events controlled by RA that lead to formation of the lung primordium from the primitive foregut remain unclear. Here, we present evidence that endogenous RA acts as a major regulatory signal integrating Wnt and Tgfbeta pathways in the control of Fgf10 expression during induction of the mouse primordial lung. We demonstrated that activation of Wnt signaling required for lung formation was dependent on local repression of its antagonist, Dickkopf homolog 1 (Dkk1), by endogenous RA. Moreover, we showed that simultaneously activating Wnt and repressing Tgfbeta allowed induction of both lung buds in RA-deficient foreguts. The data in this study suggest that disruption of Wnt/Tgfbeta/Fgf10 interactions represents the molecular basis for the classically reported failure to form lung buds in vitamin A deficiency.

Topics: Animals; Digestive System; Embryonic Development; Fibroblast Growth Factor 10; Lung; Mice; Mice, Knockout; Proteins; Signal Transduction; Transforming Growth Factor beta; Tretinoin; Vitamin A Deficiency

Retinoic acid (RA), a well-known vitamin A metabolite, mediates inhibition of the IL-6-driven induction of proinflammatory Th17 cells and promotes anti-inflammatory regulatory T cell generation in the presence of TGF-beta, which is mainly regulated by dendritic cells. To directly address the role of RA in Th17/regulatory T cell generation in vivo, we generated vitamin A-deficient (VAD) mice by continuous feeding of a VAD diet beginning in gestation. We found that a VAD diet resulted in significant inhibition of Th17 cell differentiation in the small intestine lamina propria by as early as age 5 wk. Furthermore, this diet resulted in low mRNA expression levels of IL-17, IFN regulatory factor 4, IL-21, IL-22, and IL-23 without alteration of other genes, such as RORgammat, TGF-beta, IL-6, IL-25, and IL-27 in the small intestine ileum. In vitro results of enhanced Th17 induction by VAD dendritic cells did not mirror in vivo results, suggesting the existence of other regulation factors. Interestingly, the VAD diet elicited high levels of mucin MUC2 by goblet cell hyperplasia and subsequently reduced gut microbiome, including segmented filamentous bacteria. Much like wild-type mice, the VAD diet-fed MyD88-/-TRIF-/- mice had significantly fewer IL-17-secreting CD4+ T cells than the control diet-fed MyD88-/-TRIF-/- mice. The results strongly suggest that RA deficiency altered gut microbiome, which in turn inhibited Th17 differentiation in the small intestine lamina propria.

Topics: Animals; CD4-Positive T-Lymphocytes; Cell Differentiation; Cell Separation; Down-Regulation; Flow Cytometry; Immunologic Factors; Interleukin-17; Intestine, Small; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Reverse Transcriptase Polymerase Chain Reaction; Tretinoin; Vitamin A Deficiency

Vitamin A-deficient (VAD) quail embryos lack the vitamin A-active form, retinoic acid (RA) and are characterized by a phenotype that includes a grossly abnormal cardiovascular system that can be rescued by RA. Here we report that the transforming growth factor, TGFbeta2 is involved in RA-regulated cardiovascular development. In VAD embryos TGFbeta2 mRNA and protein expression are greatly elevated. The expression of TGFbeta receptor II is also elevated in VAD embryos but is normalized by treatment with TGFbeta2-specific antisense oligonucleotides (AS). Administration of this AS or an antibody specific for TGFbeta2 to VAD embryos normalizes posterior heart development and vascularization, while the administration of exogenous active TGFbeta2 protein to normal quail embryos mimics the excessive TGFbeta2 status of VAD embryos and induces VAD cardiovascular phenotype. In VAD embryos pSmad2/3 and pErk1 are not activated, while pErk2 and pcRaf are elevated and pSmad1/5/8 is diminished. We conclude that in the early avian embryo TGFbeta2 has a major role in the retinoic acid-regulated posterior heart morphogenesis for which it does not use Smad2/3 pathways, but may use other signaling pathways. Importantly, we conclude that retinoic acid is a critical negative physiological regulator of the magnitude of TGFbeta2 signals during vertebrate heart formation.

Topics: Animals; Gene Expression Regulation, Developmental; Heart; Morphogenesis; Quail; Receptors, Transforming Growth Factor beta; Transforming Growth Factor beta2; Tretinoin; Vitamin A Deficiency

CYP26A1, which catalyzes the oxidation of all-trans (at)-retinoic acid (RA), is induced moderately by RA in numerous tissues, but is highly responsive in liver. To understand this difference, we have examined the CYP26A1 gene sequence, identified multiple RA response elements (RAREs) and tested them functionally in HepG2 cells as model hepatocytes, and in the liver of vitamin A (VA)-adequate and -deficient rats. Analysis of a 2.2 kbp 5'-flanking region upstream of the CYP26A1 transcription start site (TSS) identified 3 conserved hexameric direct repeat-5 elements, RARE1, -2 and -3, and a half site, RARE4. The full-length promoter containing all 4 elements was sufficient and necessary to increase promoter activity similar to levels of endogenous CYP26A1 mRNA produced in HepG2 cells treated with at-RA. In DNA binding and chromatin immunoprecipitation assays, the binding of RARs to the proximal RARE1 and distal RARE2, -3, and -4 regions of the CYP26A1 promoter was increased in RA-treated HepG2 cells, and greater in VA-sufficient than VA-deficient liver. Moreover, RA increased the binding of RNA polymerase-II in the distal as well as the proximal region, indicating that the distal region may be looped to become positioned close to the TSS, a process favored by retinoic acid receptors. The results support a cooperative model in which the functioning of multiple RAREs may account for the strong inducibility of CYP26A1 in liver, which, in turn, may be important physiologically for restoring retinoid homeostasis when the concentration of RA rises.

Topics: Animals; Base Sequence; Cell Line; Chromatin; Conserved Sequence; Cytochrome P-450 Enzyme System; Enzyme Induction; Hep G2 Cells; Humans; Kidney; Liver; Male; Protein Binding; Rats; Rats, Sprague-Dawley; Receptors, Retinoic Acid; Response Elements; Retinoic Acid 4-Hydroxylase; RNA Polymerase II; Transfection; Tretinoin; Vitamin A Deficiency

Interleukin-2 (IL-2) is an essential cytokine for T-lymphocyte homeostasis. We have previously reported that all-trans retinoic acid (atRA) enhances the secretion of IL-2 from human peripheral blood T cells in vitro, followed by increased proliferation and inhibition of spontaneous cell death. In this study we used a transgenic IL-2 gene luciferase reporter model to examine the effects of atRA in vivo. In contrast to the observations in human T cells, we found an overall reduction in luciferase-reported IL-2 gene expression in mice treated with atRA. Whole-body luminescence of anti-CD3-treated and non-treated mice was reduced in mice receiving atRA. Accordingly, after 7 hr, IL-2 gene expression was on average 55% lower in the atRA-treated mice compared with the control mice. Furthermore, mice fed a vitamin A-deficient diet had a significantly higher basal level of luciferase activity compared with control mice, demonstrating that vitamin A modulates IL-2 gene expression in vivo. Importantly, the atRA-mediated inhibition of IL-2 gene expression was accompanied by decreased DNA synthesis in murine T cells, suggesting a physiological relevance of the reduced IL-2 gene expression observed in transgenic reporter mice.

Topics: Animals; Benzoates; Cell Proliferation; Cells, Cultured; Chromans; Female; Gene Expression Regulation; Interleukin-2; Lymphocyte Activation; Mice; Mice, Transgenic; NF-kappa B; Receptors, Retinoic Acid; Spleen; T-Lymphocytes; Tretinoin; Vitamin A Deficiency

The farnesoid X receptor/retinoid X receptor-alpha (FXR/RXRalpha) complex regulates bile salt homeostasis, in part by modulating transcription of the bile salt export pump (BSEP/ABCB11) and small heterodimer partner (SHP/NR0B2). FXR is activated by bile salts, RXRalpha by the vitamin A derivative 9-cis retinoic acid (9cRA). Cholestasis is associated with vitamin A malabsorption. Therefore, we evaluated the role of vitamin A/9cRA in the expression of human and mouse bile salt export pump (hBSEP/mBsep), small heterodimer partner (hSHP/mShp), and mouse sodium-dependent taurocholate co-transporting polypeptide (mNtcp). HBSEP and hSHP transcription were analyzed in FXR/RXRalpha-transfected HepG2 cells exposed to chenodeoxycholic acid (CDCA) and/or 9cRA. BSEP promoter activity was determined by luciferase reporter assays, DNA-binding of FXR and RXRalpha by pull-down assays. Serum bile salt levels and hepatic expression of Bsep, Shp, and Ntcp were determined in vitamin A-deficient (VAD)/cholic acid (CA)-fed C57BL/6J mice. Results indicated that 9cRA strongly repressed the CDCA-induced BSEP transcription in HepG2 cells, whereas it super-induced SHP transcription; 9cRA reduced DNA-binding of FXR and RXRalpha. The 9cRA repressed the CDCA-induced BSEP promoter activity irrespective of the exact sequence of the FXR-binding site. In vivo, highest Bsep messenger RNA (mRNA), and protein expression was observed in CA-fed VAD mice. Shp transcription was highest in CA-fed vitamin A-sufficient mice. Ntcp protein expression was strongly reduced in CA-fed VAD mice, whereas mRNA levels were normal. CA-fed control and VAD mice had similarly increased serum bile salt levels.. We showed that 9cRA has opposite effects on bile salt-activated transcription of FXR/RXRalpha target genes. Vitamin A deficiency in CA-fed mice leads to high BSEP expression. Clearance of serum bile salts may, however, be limited because of post-transcriptional reduction of Ntcp. The molecular effects of vitamin A supplementation during cholestasis need further analysis to predict a therapeutic effect.

Topics: Alitretinoin; Animals; ATP Binding Cassette Transporter, Subfamily B, Member 11; ATP-Binding Cassette Transporters; Carcinoma, Hepatocellular; Cell Line, Tumor; Chenodeoxycholic Acid; Cholic Acid; DNA-Binding Proteins; Gene Expression Regulation; Humans; Male; Mice; Mice, Inbred C57BL; Organic Anion Transporters, Sodium-Dependent; Receptors, Cytoplasmic and Nuclear; Response Elements; Retinoid X Receptor alpha; Symporters; Transcription Factors; Tretinoin; Vitamin A; Vitamin A Deficiency

Vitamin A (VA), all-trans-retinol (at-ROL), and its derivative, all-trans-retinoic acid (at-RA), are required for neuron development. The effects of these retinoids are dependent upon the nutritional status of the rat and tissue-specific dynamics of retinoid access and utilization. The purpose of this study was to determine the status of at-ROL and at-RA in the peripheral olfactory organ of postnatal rats fed a normal diet and rats fed a VA-deficient (VAD) diet. Extracted retinoids were analyzed by HPLC. Resolved sample peaks were identified by comparing their elution times and spectra with those of authentic standards. Mean at-RA and at-ROL concentrations of 23 pmol/g olfactory tissue and 0.13 nmol/g, respectively, were recovered from olfactory tissue. The ratio of at-RA:at-ROL in olfactory was approximately 2 times that in testis and 200 times that in liver. at-ROL was depleted from the liver and olfactory organ of rats fed a VAD diet from birth to 70 d of age. Surprisingly, at-RA was still present in olfactory tissue from these rats. At 90 d of age, the VAD rats were frankly deficient and at-RA was no longer detectable in olfactory tissue. The comparatively high ratio of at-RA:at-ROL in the peripheral olfactory organ and the persistence of at-RA in at-ROL-depleted tissues strongly suggests that maintenance of local stores of at-RA is functionally relevant in this tissue.

Topics: Animals; Diet; Female; Male; Neurons; Olfactory Mucosa; Rats; Rats, Long-Evans; Tretinoin; Vitamin A; Vitamin A Deficiency

Vitamin A (retinol) is required for male and female reproduction as well as to support many developmental processes. In the male, meiotic entry of germ cells occurs after birth and throughout adulthood, whereas in the female, the entry into meiosis I occurs during embryonic development. Evidence from cultured embryonic ovaries suggests that the vitamin A metabolite, all-trans retinoic acid (atRA), initiates this process. However, in vivo evidence to support a normal role for atRA in meiotic entry is lacking. The present study demonstrates that although germ cell number is normal in ovaries from both vitamin A-sufficient (VAS) embryos and those that are deficient in atRA, the majority of germ cells in the most severely atRA-deficient group fail to enter meiosis and remain in an undifferentiated state. In contrast, in a group that is only moderately deficient in atRA, a small number of ovarian germ cells enter meiosis (30%) compared with 75% of cells in the VAS control group. The expression of the atRA-responsive gene, Stra8, is reduced by approximately 90% and 50% in the severely and moderately atRA-deficient ovaries, respectively, compared with the VAS controls. These results provide the first in vivo evidence that vitamin A regulates the entry of germ cells into meiosis in the developing ovary.

Topics: Animals; Cell Count; Cell Cycle Proteins; Chromosomes; Female; Fluorescent Antibody Technique; Male; Meiosis; Ovary; Ovum; Proteins; Rats; Rats, Sprague-Dawley; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Testis; Tretinoin; Vitamin A Deficiency

Retinoic acid plays a positive role in induction of FoxP3(+) regulatory T cells. Because retinoic acid is produced as a metabolite of vitamin A in the intestine and FoxP3(+) T cells regulate intestinal inflammation, we investigated the impact of vitamin A status on the regulatory T cells and inflammation in the intestine.. The SAMP1/YP model is a mouse model of Crohn's disease. We made vitamin A-deficient, vitamin A-excessive, and normal SAMP1/YP mice and assessed the intestinal inflammation. We also investigated the phenotype and function of FoxP3(+) T cells induced in different levels of vitamin A availability in regulation of intestinal inflammation in a T-cell-induced inflammation model in SCID mice.. The limited and excessive vitamin A conditions induced distinct FoxP3(+) T-cell subsets in vivo, and both ameliorated the intestinal inflammation in SAMP1/YP mice. The limited vitamin A condition greatly induced unusual CD103(+)CCR7(+) FoxP3(+) cells, while the high vitamin A condition induced CCR9(+)alpha4beta7(+) FoxP3(+) T cells in the intestine. Both FoxP3(+) T-cell populations, when transferred into mice with ongoing intestinal inflammation, were highly effective in reversing the inflammation. Blockade or lack of occupancy of RARalpha is a mechanism to induce highly suppressive CD103(+)CCR7(+) FoxP3(+) cells in both the thymus and periphery in limited vitamin A availability.. Our results identify novel pathways of inducing highly suppressive FoxP3(+) regulatory T cells that can effectively control intestinal inflammation. The results have significant ramifications in treating inflammatory bowel diseases.

Topics: Adoptive Transfer; Animals; Anti-Inflammatory Agents; Antigens, CD; Cell Movement; Cells, Cultured; Coculture Techniques; Crohn Disease; Disease Models, Animal; Forkhead Transcription Factors; Gastrointestinal Agents; Immunophenotyping; Integrin alpha Chains; Intestines; Mice; Mice, Inbred BALB C; Mice, SCID; Receptors, CCR; Receptors, CCR7; Receptors, Retinoic Acid; Retinoic Acid Receptor alpha; Spleen; T-Lymphocyte Subsets; Th1 Cells; Th2 Cells; Thymus Gland; Tretinoin; Vitamin A; Vitamin A Deficiency

To study the influence of atRA on iron metabolism in cultured primary rat hepatocyte.. Rat primary hepatocytes were isolated by two-step in situ collagenase perfusion method by Seglen, and after that Cell viability was observed by 0.4% trypan blue. And then primary hepatocyte were treated into 6 wells plate with 0, 0.5, 1 and 50 micromol/L atRA and DMEM contained 10% fetal bovine serum. After 72h, IRP2 mRNA, TFR mRNA, Fnm RNA levels were measured by RT-PCR.. VA deficiency can decrease the viability and function. Moreover hepatic IRP2 mRNA and TFRmRNA levels were increased by VA deficiency, which diminishing expression of Fn mRNA.. vitamin A deficiency can change cellular iron metabolism by inducing IRP2-Fn-TFR pathway. AtRA supplementation inhibited the increase in IRP2 mRNA expression. Taken together, these results indicate that vitamin A deficiency can regulate iron metabolism by IRP2-TFR-Fn pathway.

Topics: Animals; Cells, Cultured; Female; Hepatocytes; Iron; Iron Regulatory Protein 2; Male; Rats; Rats, Sprague-Dawley; Receptors, Transferrin; RNA, Messenger; Tretinoin; Vitamin A Deficiency

Vitamin A (retinol) and its analogs (retinoids) are important regulators of cell proliferation, differentiation, immune function, and apoptosis. The kidneys are target organs for vitamin A action. Retinoic acid (RA), a vitamin A metabolite, is involved in embryonic kidney patterning through the control of receptor tyrosine kinase expression, which modulates ureteric bud branching morphogenesis. Vitamin A status of the mother profoundly affects kidney organogenesis of the newborn. In rodents, mild vitamin A deficiency results in a 20% reduction of nephron number. In adult humans, nephron number varies between 0.3 and 1.3 million per kidney, which is accepted as normal. However, recent studies indicate that humans at the low end of nephron number are predisposed to primary hypertension. Because RA regulates nephron mass, its optimal availability during nephrogenesis is critical. RA levels in the embryo are affected by several factors, such as maternal vitamin A nutrition and disturbances in retinol metabolism. Maternal vitamin A deficiency during pregnancy is widespread in developing countries and segments of these populations may be exposed to low vitamin A during fetal life when nephron number is determined. Infants are likely to be born with suboptimal nephrons and may develop primary hypertension later in life. Although maternal vitamin A deficiency is not common in developed countries, congenital nephron number nevertheless varies widely, indicating low fetal RA levels due to common variants of the enzymes that convert retinol to RA. These infants might require heightened surveillance for hypertension later in life.

Topics: Animals; Female; Fetus; Gene Expression Regulation, Developmental; Humans; Hypertension; Infant, Newborn; Maternal-Fetal Exchange; Nephrons; Pregnancy; Pregnancy Complications; Tretinoin; Vitamin A; Vitamin A Deficiency

A dysfunction of retinoid hippocampal signaling pathway has been involved in the appearance of affective and cognitive disorders. However, the underlying neurobiological mechanisms remain unknown. Hippocampal granule neurons are generated throughout life and are involved in emotion and memory. Here, we investigated the effects of vitamin A deficiency (VAD) on neurogenesis and memory and the ability of retinoic acid (RA) treatment to prevent VAD-induced impairments. Adult retinoid-deficient rats were generated by a vitamin A-free diet from weaning in order to allow a normal development. The effects of VAD and/or RA administration were examined on hippocampal neurogenesis, retinoid target genes such as neurotrophin receptors and spatial reference memory measured in the water maze. Long-term VAD decreased neurogenesis and led to memory deficits. More importantly, these effects were reversed by 4 weeks of RA treatment. These beneficial effects may be in part related to an up-regulation of retinoid-mediated molecular events, such as the expression of the neurotrophin receptor TrkA. We have demonstrated for the first time that the effect of vitamin A deficient diet on the level of hippoccampal neurogenesis is reversible and that RA treatment is important for the maintenance of the hippocampal plasticity and function.

Topics: Animals; Hippocampus; Memory Disorders; Neurogenesis; Rats; Receptor, trkA; Regeneration; Treatment Outcome; Tretinoin; Up-Regulation; Vitamin A Deficiency

Vitamin A plays an essential role in vertebrate embryogenesis. In the present study, pregnant vitamin A-deficient (VAD) rats were maintained during early pregnancy on the short half-life vitamin A metabolite, all-trans retinoic acid (atRA), in an amount sufficient to support normal development to E10.5, with a higher level of atRA (250 microg atRA/g diet) provided from embryonic day (E) 8.5-10.5 to prevent mid-gestational resorption. When limiting amounts of atRA (1.5 or 12 microg/g diet) were provided after E10.5, a highly reproducible and penetrant state of late fetal vitamin A deficiency (late VAD) was induced in the organs of developing fetuses. In addition, late VAD fetuses displayed both anteriorization of cervical regions and novel posteriorization events at the thoracic and sacral levels of the skeleton, and showed sternal and pelvic malformations not previously observed in early VAD or genetic models. The expression of several Hox genes (Hoxd3 and Hoxb4) was altered in late VAD embryos, with a reduction in Hoxd3 noted as early as 1 day after instituting deficiency. All late VAD-induced malformations were prevented by the addition of retinol starting at E10.5, whereas provision of a high level of atRA throughout pregnancy improved but could not completely rescue the development of all organ systems. This work defines a nutritional model in which vitamin A deficiency can be induced during fetal development, and reveals new functions for the vitamin in the development of the axial and appendicular skeleton.

Topics: Animal Nutritional Physiological Phenomena; Animals; Bone Development; Disease Models, Animal; Fetal Development; Gastrula; In Situ Hybridization; Organogenesis; Rats; Rats, Sprague-Dawley; Tretinoin; Vitamin A Deficiency

Congenital diaphragmatic hernia (CDH) is a congenital malformation that occurs with a frequency of 0.08 to 0.45 per 1,000 births. Children with CDH are born with the abdominal contents herniated through the diaphragm and exhibit an associated pulmonary hypoplasia which is frequently accompanied by severe morbidity and mortality. Although the etiology of CDH is largely unknown, considerable progress has been made in understanding its molecular mechanisms through the usage of genetic, teratogenic, and surgical models. The following review focuses on the teratogenic and surgical models of CDH and the possible molecular mechanisms of nitrofen (a diphenyl ether, formerly used as an herbicide) in both induction of CDH and pulmonary hypoplasia. In addition, the mechanisms of other compounds including several anti-inflammatory agents that have been linked to CDH will be discussed. Furthermore, this review will also explore the importance of vitamin A in lung and diaphragm development and the possible mechanisms of teratogen interference in vitamin A homeostasis. Continued exploration of these models will bring forth a clearer understanding of CDH and its molecular underpinnings, which will ultimately facilitate development of therapeutic strategies.

Topics: Animals; Diaphragm; Disease Models, Animal; Dogs; Hernia, Diaphragmatic; Lung; Models, Animal; Phenyl Ethers; Rabbits; Rats; Sheep; Signal Transduction; Species Specificity; Teratogens; Tretinoin; Vitamin A; Vitamin A Deficiency

Vitamin A supplementation for infants and young children is recommended by WHO/UNICEF for countries with a high prevalence of vitamin A deficiency, and vitamin A is often administered at immunization contacts. Using a rat model, we tested whether supplementation with vitamin A or other retinoids at the time of neonatal immunization has prospective benefit in terms of preventing postweaning vitamin A deficiency and promoting antibody responses to T-cell dependent (TD) antigens administered at the neonatal stage and at the young adult stage. Rats were treated orally on postnatal d 6-8 with oil (placebo control), vitamin A, retinoic acid, or a combination of both (VARA) (n > or = 12/group), and immunized with tetanus toxoid (TT) on d 7. The primary anti-TT response was measured on d 21, after which weanling rats were fed the vitamin A-deficient diet until approximately 10 wk. At 8 wk, rats were immunized again with TT to determine the recall response, and with a novel TD antigen, keyhole limpet hemocyanin (KLH), to assess the adult primary response. None of the supplements affected the plasma titer of anti-TT immunoglobulin G (IgG) on d 21 (P = 0.25). However, neonatal-age supplementation with vitamin A or VARA at the young adult stage resulted in: >5 times higher anti-TT IgG recall response (P < 0.01); 5- and 9-times higher anti-KLH primary IgM and IgG responses, respectively (P < 0.05), and plasma retinol in the normal range (approximately 1.0 micromol/L vs. approximately 0.35 micromol/L in retinoic acid-treated and control groups, P < 0.0001). We conclude that early-life supplementation with vitamin A or VARA can prospectively benefit the primary and recall antibody responses to TD antigens administered at the young adult stage, which may involve the maintenance of normal plasma retinol levels.

Topics: Adjuvants, Immunologic; Animals; Animals, Newborn; Antibodies; Antigens; Diet; Drug Administration Schedule; Drug Combinations; Female; Hemocyanins; Immunization; Male; Rats; Rats, Sprague-Dawley; T-Lymphocytes; Tetanus Toxoid; Tretinoin; Vitamin A; Vitamin A Deficiency; Weaning

We have investigated the role of retinoic acid (RA) in eye development using the vitamin A deficient quail model system, which overcomes problems of retinoic acid synthesising enzyme redundancy in the embryo. In the absence of retinoic acid, the ventral optic stalk and ventral retina are missing, whereas the dorsal optic stalk and dorsal retina develop appropriately. Other ocular abnormalities observed were a thinner retina and the lack of differentiation of the lens. In an attempt to explain this, we studied the expression of various dorsally and ventrally expressed genes such as Pax2, Pax6, Tbx6, Vax2, Raldh1 and Raldh3 and noted that they were unchanged in their expression patterns. In contrast, the RA catabolising enzymes Cyp26A1 and Cyp26B1 which are known to be RA-responsive were not expressed at all in the developing eye. At much earlier stages, the expression domain of Shh in the prechordal plate was reduced, as was Nkx2.1 and we suggest a model whereby the eye field is specified according to the concentration of SHH protein that is present. We also describe another organ, Rathke's pouch which fails to develop in the absence of retinoic acid. We attribute this to the down-regulation of Bmp2, Shh and Fgf8 which are known to be involved in the induction of this structure.

Topics: Animals; Coturnix; Embryo, Nonmammalian; Embryonic Structures; Eye; Gene Expression Regulation, Developmental; Models, Biological; Tretinoin; Vitamin A Deficiency

The lungs require an adequate supply of vitamin A for normal embryonic development, postnatal maturation, and maintenance and repair during adult life. We have previously shown that a nutrient-metabolite combination of vitamin A admixed with a small proportion (10%) of retinoic acid (RA), referred to as VARA, acts synergistically to increase lung retinyl ester (RE) concentration in neonatal rats. A series of studies was designed to test whether VARA increases RE in adult lungs, and whether VARA is more effective than vitamin A when given by the i.m. route. Orally administered VARA increased RE in the lungs of vitamin A-marginal adult rats more than either vitamin A or RA alone (P < 0.05). In vitamin A-deficient young adult rats, lung RE was increased by VARA when administered by the i.m. route. When a tracer of (3)H-retinol was added to the placebo (oil), vitamin A, and VARA doses, total (3)H and (3)H-RE increased in the lungs more with VARA than vitamin A alone, for oral and i.m. dosing. Nevertheless, when VARA and vitamin A were given by the oral route, they were more effective in increasing RE in the liver. Plasma retinol was increased similarly in vitamin A-deficient rats after administration of VARA and vitamin A, by either the oral or the i.m. route. Overall, VARA can increase retinol uptake and esterification in adult lungs when delivered intramuscularly as well as orally.

Topics: Administration, Oral; Animals; Biological Transport; Female; Injections, Intramuscular; Lipids; Lung; Rats; Rats, Sprague-Dawley; Tretinoin; Vitamin A; Vitamin A Deficiency

Although retinoic acid (RA) has been implicated as one of the diffusible signals regulating forebrain development, patterning of the forebrain has not been analyzed in detail in knockout mouse mutants deficient in embryonic RA synthesis. We show that the retinaldehyde dehydrogenase 2 (RALDH2) enzyme is responsible for RA synthesis in the mouse craniofacial region and forebrain between the 8- and 15-somite stages. Raldh2-/- knockout embryos exhibit defective morphogenesis of various forebrain derivatives, including the ventral diencephalon, the optic and telencephalic vesicles. These defects are preceded by regionally decreased cell proliferation in the neuroepithelium, correlating with abnormally low D-cyclin gene expression. Increases in cell death also contribute to the morphological deficiencies at later stages. Molecular analyses reveal abnormally low levels of FGF signaling in the craniofacial region, and impaired sonic hedgehog signaling in the ventral diencephalon. Expression levels of several regulators of diencephalic, telencephalic and optic development therefore cannot be maintained. These results unveil crucial roles of RA during early mouse forebrain development, which may involve the regulation of the expansion of neural progenitor cells through a crosstalk with FGF and sonic hedgehog signaling pathways.

Topics: Aldehyde Oxidoreductases; Animals; Body Patterning; Cell Death; Cell Proliferation; Fibroblast Growth Factors; Gestational Age; Hedgehog Proteins; Mice; Mice, Knockout; Models, Neurological; Neural Crest; Prosencephalon; Signal Transduction; Telencephalon; Trans-Activators; Tretinoin; Visual Pathways; Vitamin A Deficiency

In mammals, meiosis is initiated at different time points in males and females, but the mechanism underlying this difference is unknown. Female germ cells begin meiosis during embryogenesis. In males, embryonic germ cells undergo G0/G1 mitotic cell cycle arrest, and meiosis begins after birth. In mice, the Stimulated by Retinoic Acid Gene 8 (Stra8) has been found to be required for the transition into meiosis in both female and male germ cells. Stra8 is expressed in embryonic ovaries just before meiotic initiation, whereas its expression in testes is first detected after birth. Here we examine the mechanism underlying the sex-specific timing of Stra8 expression and meiotic initiation in mice. Our work shows that signaling by retinoic acid (RA), an active derivative of vitamin A, is required for Stra8 expression and thereby meiotic initiation in embryonic ovaries. We also discovered that RA is sufficient to induce Stra8 expression in embryonic testes and in vitamin A-deficient adult testes in vivo. Finally, our results show that cytochrome p450 (CYP)-mediated RA metabolism prevents premature Stra8 expression in embryonic testes. Treatment with an inhibitor specific to RA-metabolizing enzymes indicates that a cytochrome p450 from the 26 family (CYP26) is responsible for delaying Stra8 expression in embryonic testes. Sex-specific regulation of RA signaling thus plays an essential role in meiotic initiation in embryonic ovaries and precludes its occurrence in embryonic testes. Because RA signaling regulates Stra8 expression in both embryonic ovaries and adult testes, this portion of the meiotic initiation pathway may be identical in both sexes.

Topics: Adaptor Proteins, Signal Transducing; Animals; Cytochrome P-450 Enzyme System; Female; Male; Meiosis; Mice; Ovary; Proteins; Retinoic Acid 4-Hydroxylase; Sex Determination Processes; Testis; Time Factors; Tretinoin; Vitamin A Deficiency

Retinoic acid (RA) is commonly used in vitro to differentiate stem cell populations including adult neural stem cells into neurons; however, the in vivo function of RA during adult neurogenesis remains largely unexplored. We found that depletion of RA in adult mice leads to significantly decreased neuronal differentiation within the granular cell layer of the dentate gyrus. RA contribution to neurogenesis occurs early, for RA deficiency also results in a decrease in newborn cells expressing an immature neuronal marker. Furthermore, although proliferation is unaffected during RA absence, cell survival is significantly reduced. Finally, a screen for retinoid-induced genes identifies metabolic targets including the lipid transporters, CD-36 and ABCA-1, the lipogenic master regulator SREBP1c as well as components of the Wnt signaling pathway. Our results reveal RA as a crucial contributor to early stages of adult neurogenesis and survival in vivo.

Topics: Animals; Cell Differentiation; Cell Proliferation; Cell Survival; Dentate Gyrus; Gene Expression; In Vitro Techniques; Mice; Mice, Inbred SENCAR; Neurons; Stem Cells; Tretinoin; Vitamin A Deficiency

Recent data have revealed that disruption of vitamin A signaling observed in Alzheimer's disease (AD) leads to a deposition of beta-amyloid (Abeta). The aim of this study was to precise the role of vitamin A and its nuclear receptors (RAR) in the processes leading to the Abeta deposits. Thus, the effect of vitamin A depletion and subsequent administration of retinoic acid (RA, the active metabolite of vitamin A) on the expression of RARbeta, and of proteins involved in amyloidogenic pathway, e.g., amyloid precursor protein (APP), beta-secretase enzyme (BACE), and APP carboxy-terminal fragment (APP-CTF) was examined in the whole brain, hippocampus, striatum, and cerebral cortex of rats. Rats fed a vitamin A-deprived diet for 13 weeks exhibited decreased amount of RARbeta, APP695, BACE, and of APP-CTF in the whole brain and in the cerebral cortex. Administration of RA is able to restore all expression. The results suggest that fine regulation of vitamin A mediated gene expression seems fundamental for the regulation of APP processing.

Topics: Amyloid beta-Peptides; Amyloid beta-Protein Precursor; Amyloid Precursor Protein Secretases; Animals; Aspartic Acid Endopeptidases; Blotting, Western; Cerebral Cortex; Endopeptidases; Male; Polymerase Chain Reaction; Rats; Rats, Wistar; Receptors, Retinoic Acid; RNA, Messenger; Tretinoin; Vitamin A; Vitamin A Deficiency

beta-Carotene exhibits biological activity as provitamin A. Key step in vitamin A formation is the cleavage of beta-carotene to retinal by an enzyme designated as beta-carotene 15,15'-monooxygenase (BCM). Recently, it is reported that expression of BCM gene in the intestine is under feedback regulation by retinoic acid (RA). However, the regulation of BCM gene expression in various other tissues is still unknown.. In the present study, we identified the full-length cDNA encoding the rat BCM gene and investigated the regulation of its expression in several tissues by RA in the presence of vitamin A deficiency.. We cloned the full-length cDNA encoding BCM gene from a rat intestinal cDNA library by hybridization screening. The BCM gene expression was examined using Northern blotting and reverse transcription-PCR analysis. We also investigated whether BCM gene expression was regulated by retinoids in several tissues of vitamin A-deficient rats.. Sequence analysis of this clone revealed an open reading frame of 1,701 bases encoding a protein of 566 amino acids. The predicted polypeptide showed 94%, 81%, and 66% identity with mouse, human, and chicken BCM, respectively. Rat BCM mRNA was highly expressed in the intestine and liver, while there was weak expression in the testes, kidneys, and lungs. Immunoblotting revealed that rat BCM is a 64-kDa protein. BCM gene expression was increased in the small intestine by vitamin A deficiency compared with that in rats on a control diet, while this upregulation was suppressed by all-trans RA (ATRA) or 9-cis RA (9-cis RA). BCM gene expression in the lungs and testes was also suppressed by ATRA or 9-cis RA in rats with vitamin A deficiency. However, hepatic BCM gene expression was only decreased by ATRA and renal expression was not affected by either retinoid. As the small intestine is the major site of beta-carotene conversion, intestinal BCM gene expression may be more tightly regulated.. These data suggest that BCM gene expression in several tissues may be down-regulated by RA at the level of conversion of beta-carotene to retinal. To prevent an excess of retinol, homeostasis may occur at the level of conversion of beta-carotene to retinal in several tissues.

Topics: Amino Acid Sequence; Animals; beta-Carotene 15,15'-Monooxygenase; Cloning, Molecular; DNA, Complementary; Gene Expression Regulation, Enzymologic; Gene Library; Humans; Immunoblotting; Molecular Sequence Data; Open Reading Frames; Organ Specificity; Rats; Reverse Transcriptase Polymerase Chain Reaction; Sequence Homology, Amino Acid; Tretinoin; Vitamin A Deficiency

Retinoids, including all-trans-retinoic acid (RA), are considered to have anti-inflammatory properties and are used therapeutically for diseases of the skin and certain cancers. However, few studies have addressed the effects of disease states on RA metabolism. The present study was conducted to better understand the effects of exogenous RA, both in the absence and presence of inflammation, on the distribution and metabolism of a dose of [3H]RA. Female Sprague-Dawley rats fed a low vitamin A diet were pretreated with RA (po), a low dose of lipopolysaccharide (LPS, ip), or their combination. Twelve hours later, albumin-bound [3H]RA was injected intravenously, and tissue organic- and aqueous-phase 3H was determined after 10 and 30 min. In liver and plasma, 3H-labeled organic metabolites (e.g., 4-oxo- and 4-hydroxy-RA) were isolated by solid-phase extraction. LPS-induced inflammation significantly reduced plasma retinol by 47%, increased total 3H in plasma at 10 min, and reduced total 3H in liver at both times. In contrast, RA pretreatment did not affect plasma retinol, significantly increased total 3H in plasma at both times, and did not affect liver total 3H. However, by 30 min, RA significantly increased [3H]RA metabolism in plasma, liver, lung, and small intestine, as indicated by greater 3H-labeled aqueous-phase and 3H-labeled organic-phase metabolites. The results presented here demonstrate that, although LPS-induced inflammation affects the organ distribution of RA, the ability of RA to induce its own catabolism is maintained during inflammation. Thus we conclude that RA and LPS act independently to alter RA metabolism in vitamin A-marginal rats.

Topics: Animals; Drug Combinations; Female; Homeostasis; Inflammation; Lipopolysaccharides; Metabolic Clearance Rate; Organ Specificity; Rats; Rats, Sprague-Dawley; Tissue Distribution; Tretinoin; Vitamin A; Vitamin A Deficiency

Normal intestinal mucosa contains abundant immunoglobulin A (IgA)-secreting cells, which are generated from B cells in gut-associated lymphoid tissues (GALT). We show that dendritic cells (DC) from GALT induce T cell-independent expression of IgA and gut-homing receptors on B cells. GALT-DC-derived retinoic acid (RA) alone conferred gut tropism but could not promote IgA secretion. However, RA potently synergized with GALT-DC-derived interleukin-6 (IL-6) or IL-5 to induce IgA secretion. Consequently, mice deficient in the RA precursor vitamin A lacked IgA-secreting cells in the small intestine. Thus, GALT-DC shape mucosal immunity by modulating B cell migration and effector activity through synergistically acting mediators.

Topics: Animals; B-Lymphocytes; Cell Movement; Cells, Cultured; Chemotaxis, Leukocyte; Dendritic Cells; Immunity, Mucosal; Immunoglobulin A; Interleukin-5; Interleukin-6; Intestinal Mucosa; Intestines; Lymphoid Tissue; Mice; Mice, Inbred C57BL; Receptors, Antigen, B-Cell; Tretinoin; Vitamin A; Vitamin A Deficiency; Vitamins

Vitamin A is essential for the visual system. It is metabolized in the retina and the resulting product, retinoic acid (RA), greatly affects the structure and functions of retinal pigment epithelial (RPE) cells. RPE cells produce a variety of extracellular matrix (ECM) proteins and angiogenic factors, both of which are expressed at varying levels in the normal RPE layer. In this study, we investigated the effect of all-trans-retinoic acid on the production of an ECM protein, thrombospondin-1 (TSP-1), and two angiogenic factors, pigment epithelium-derived factor (PEDF) and vascular endothelial growth factor (VEGF) by RPE cells. RA increased the release of TSP-1 and PEDF, but not that of VEGF, from human RPE cells in vitro. In vitamin A-deficient mice, the expression of TSP-1 and PEDF in the RPE layer considerably decreased compared with that of normal control mice. The vitamin A deficiency hardly affected the accumulation of VEGF in the RPE layer. These findings suggest that vitamin A modulates the structure and anti-angiogenic functions of the RPE layer partly by up-regulating the expression of the angiogenesis-related ECM protein, TSP-1, and the anti-angiogenic factor, PEDF.

Topics: Animals; Cells, Cultured; Choroid; Culture Media; Dose-Response Relationship, Drug; Epithelial Cells; Eye Proteins; Humans; Keratolytic Agents; Mice; Nerve Growth Factors; Pigment Epithelium of Eye; Reverse Transcriptase Polymerase Chain Reaction; Serpins; Thrombospondin 1; Tretinoin; Up-Regulation; Vascular Endothelial Growth Factor A; Vitamin A; Vitamin A Deficiency

Studies using genetics and vitamin A deficiency (VAD) have shown that vitamin A and retinoids play essential roles in spermatogenesis at the pre-meiotic stage. To understand the mechanisms of control in spermatogenesis by retinoic acid, we investigated whether retinoic acids regulate the expression of downstream transcription factors that are essential for spermatogenesis. In this study, we found that administration of all-trans retinoic acid (ATRA) or retinol to VAD rats down-regulates the testicular mRNA levels of the cAMP responsive element modulator (CREM), an essential transcription factor for spermatogenesis. Conversely, depletion of retinoids from the diet leds to an up-regulation of CREM expression in adult testes. In addition, RT-PCR analysis indicated that ATRA specifically represses the expression of the activator spliced variant of CREM (CREMtau). These results suggest that retinoids function as a negative regulator of CREM expression in testes.

Topics: Animals; Cyclic AMP Response Element Modulator; DNA-Binding Proteins; Down-Regulation; Male; Rats; Rats, Wistar; RNA, Messenger; Testis; Transcription Factors; Tretinoin; Vitamin A; Vitamin A Deficiency

To study the effects of exogenous retinoic acid on vitamin A (VA) metabolism, we analyzed previously collected tracer kinetic data on VA dynamics in rats with low vitamin A (LA) status either with (LA+RA) or without (LA) retinoic acid supplementation. In spite of low VA intake ( approximately 7 nmol/d), the LA+RA rats were in a slight positive VA balance (0.325 nmol/d vs. -0.168 for LA) for 35 d after administration of [(3)H]retinol-labeled plasma. Using the Windows version of the Simulation, Analysis and Modeling software, we determined that the VA disposal rate was lower in LA+RA than in LA rats (3.98 vs. 5.00 nmol/d) as was the system fractional catabolic rate (0.0548 vs. 0.110 d(-1)). Model-predicted traced mass and residence times (the average time that a molecule of retinol spends in an organ before irreversible loss) were higher for liver (19.4 vs. 1.8 nmol; 5.0 vs. 0.36 d), kidneys (7.0 vs. 2.1 nmol; 1.4 vs. 0.42 d), small intestine (2.1 vs. 0.42 nmol; 0.43 vs. 0.084 d), and lungs (3.2 vs. 0.10 nmol; 1.6 vs. 0.021 d) in the LA+RA compared with the LA rats; there were no major differences for eyes, testes, adrenal glands, or remaining carcass. We conclude that RA supplementation of rats with low VA status affects VA metabolism at both the whole-body level and in specific organs. These organs (liver, kidneys, small intestine, and lungs) have the enzymatic capability and an appropriate cell type to store retinyl esters.

Topics: Animals; Carbon Radioisotopes; Diet; Kinetics; Male; Organ Specificity; Rats; Rats, Sprague-Dawley; Tretinoin; Vitamin A; Vitamin A Deficiency

All-trans retinoic acid (atRA) is an endogenous morphogen that regulates gene transcription. Maternal exposure to atRA results in severe developmental abnormalities by disrupting normal patterns of atRA distribution. Previously, we have shown that the pontine nucleus, which originates from the rhombic lip, is severely atrophied in the mouse on exposure to atRA at gestational days 9 and 10. In this study, we show that this same period of atRA exposure has the contrary effect on the inferior olive and this rhombic lip derivative is expanded in volume and probably contains an increased number of cells. The posterior region of the inferior olive maintains a relatively normal shape but is significantly expanded in size. In contrast, the organization of the anterior inferior olive is severely disrupted. Because endogenous atRA levels are known to be higher in the region of the posterior inferior olive at the time of birth of inferior olivary neurons, these results suggest that endogenous atRA may promote the generation, or select the fate, of posterior neurons of the inferior olive. In support of this concept, a reduction in atRA resulting from vitamin A deficiency results in loss of cells of the posterior inferior olive.

Topics: Animals; Brain; Embryo, Mammalian; Gene Expression Regulation, Developmental; In Situ Hybridization; Mice; Mice, Inbred C57BL; Models, Anatomic; Olivary Nucleus; Rats; RNA; Time Factors; Transcription, Genetic; Tretinoin; Vitamin A; Vitamin A Deficiency

Bronchial hyperreactivity is influenced by properties of the conducting airways and the surrounding pulmonary parenchyma, which is tethered to the conducting airways. Vitamin A deficiency (VAD) is associated with an increase in airway hyperreactivity in rats and a decrease in the volume density of alveoli and alveolar ducts. To better define the effects of VAD on the mechanical properties of the pulmonary parenchyma, we have studied the elastic modulus, elastic fibers and elastin gene-expression in rats with VAD, which were supplemented with retinoic acid (RA) or remained unsupplemented.. Parenchymal mechanics were assessed before and after the administration of carbamylcholine (CCh) by determining the bulk and shear moduli of lungs that that had been removed from rats which were vitamin A deficient or received a control diet. Elastin mRNA and insoluble elastin were quantified and elastic fibers were enumerated using morphometric methods. Additional morphometric studies were performed to assess airway contraction and alveolar distortion.. VAD produced an approximately 2-fold augmentation in the CCh-mediated increase of the bulk modulus and a significant dampening of the increase in shear modulus after CCh, compared to vitamin A sufficient (VAS) rats. RA-supplementation for up to 21 days did not reverse the effects of VAD on the elastic modulus. VAD was also associated with a decrease in the concentration of parenchymal elastic fibers, which was restored and was accompanied by an increase in tropoelastin mRNA after 12 days of RA-treatment. Lung elastin, which was resistant to 0.1 N NaOH at 98 degrees, decreased in VAD and was not restored after 21 days of RA-treatment.. Alterations in parenchymal mechanics and structure contribute to bronchial hyperreactivity in VAD but they are not reversed by RA-treatment, in contrast to the VAD-related alterations in the airways.

Topics: Adaptation, Physiological; Animals; Elasticity; Elastin; Female; Gene Expression Regulation; Lung; Rats; Rats, Inbred Lew; Tretinoin; Vitamin A Deficiency

We consider here how morphogenetic signals involving retinoic acid (RA) are switched on and off in the light of positive and negative feedback controls which operate in other embryonic signalling systems. Switching on the RA signal involves the synthetic retinaldehyde dehydrogenase (RALDH) enzymes and it is currently thought that switching off the RA signal involves the CYP26 enzymes which catabolise RA. We have tested whether these enzymes are regulated by the presence or absence of all-trans-RA using the vitamin A-deficient quail model system and the application of excess retinoids on beads to various locations within the embryo. The Raldhs are unaffected either by the absence or presence of excess RA, whereas the Cyps are strongly affected. In the absence of RA some, but not all domains of Cyp26A1, Cyp26B1 and Cyp26C1 are down-regulated, in particular the spinal cord (Cyp26A1), the heart and developing vasculature (Cyp26B1) and the rhombomeres (Cyp26C1). In the presence of excess RA, the Cyps show a differential regulation-Cyp26A1 and Cyp26B1 are up-regulated whereas Cyp26C1 is down-regulated. We tested whether the Cyp products have a similar influence on these genes and indeed 4-oxo-RA, 4-OH-RA and 5,6-epoxy-RA do. Furthermore, these 3 metabolites are biologically active in that they fully rescue the vitamin A-deficient quail embryo. Finally, by using retinoic acid receptor selective agonists we show that these compounds regulate the Cyps through the RARalpha receptor. These results are discussed with regard to positive and negative feedback controls in developing systems.

Topics: Animals; Base Sequence; Coturnix; Cytochrome P-450 Enzyme System; DNA, Complementary; Feedback; Gene Expression Regulation, Developmental; Gene Expression Regulation, Enzymologic; Morphogenesis; Receptors, Retinoic Acid; Retinoic Acid 4-Hydroxylase; Signal Transduction; Tretinoin; Vitamin A Deficiency

Lecithin:retinol acyltransferase (LRAT) catalyzes the esterification of retinol (vitamin A) in the liver and in some extrahepatic tissues, including the lung. We produced an LRAT gene knock-out mouse strain and assessed whether LRAT-/- mice were more susceptible to vitamin A deficiency than wild type (WT) mice. After maintenance on a vitamin A-deficient diet for 6 weeks, the serum retinol level was 1.34 +/- 0.32 microM in WT mice versus 0.13 +/- 0.06 microM in LRAT-/- mice (p < 0.05). In liver, lung, eye, kidney, brain, tongue, adipose tissue, skeletal muscle, and pancreas, the retinol levels ranged from 0.05 pmol/mg (muscle and tongue) to 17.35 +/- 2.66 pmol/mg (liver) in WT mice. In contrast, retinol was not detectable (<0.007 pmol/mg) in most tissues from LRAT-/- mice after maintenance on a vitamin A-deficient diet for 6 weeks. Cyp26A1 mRNA was not detected in hepatic tissue samples from LRAT-/- mice but was detected in WT mice fed the vitamin A-deficient diet. These data indicate that LRAT-/- mice are much more susceptible to vitamin A deficiency and should be an excellent animal model of vitamin A deficiency. In addition, the retinol levels in serum rapidly increased in the LRAT-/- mice upon re-addition of vitamin A to the diet, indicating that serum retinol levels in LRAT-/- mice can be conveniently modulated by the quantitative manipulation of dietary retinol.

Topics: Acyltransferases; Animals; Carboxylic Ester Hydrolases; Chromatography, High Pressure Liquid; Cytochrome P-450 Enzyme System; Exons; Gene Expression Regulation; Genetic Vectors; Genotype; Liver; Lung; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Models, Genetic; Mutation; Retinoic Acid 4-Hydroxylase; Reverse Transcriptase Polymerase Chain Reaction; RNA; RNA, Messenger; Time Factors; Tissue Distribution; Tretinoin; Vitamin A; Vitamin A Deficiency

Airway hyperresponsiveness (AHR) is influenced by structural components of the bronchial wall, including the smooth muscle and connective tissue elements and the neuromuscular function. AHR is also influenced by parenchymally derived tethering forces on the bronchial wall, which maintain airway caliber by producing outward radial traction. Our previous work has shown that vitamin A-deficient (VAD) rats exhibit cholinergic hyperresponsiveness and a decrease in the expression and function of the muscarinic-2 receptors (M2R). We hypothesized that if decreases in radial traction from airway or parenchymal structures contributed to the VAD-related increase in AHR, then the radial traction would normalize more slowly than VAD-related alterations in neurotransmitter signaling. Rats remained vitamin A sufficient (VAS) or were rendered VAD and then maintained on the VAD diet in the presence or absence of supplementation with all-trans retinoic acid (RA). VAD was associated with an approximately twofold increase in respiratory resistance and elastance compared with VAS rats. Exposure to RA for 12 days but not 4 days restored resistance and elastance to control (VAS) levels. In VAD rats, AHR was accompanied by decreases in bronchial M2R gene expression and function, which were restored after 12 days of RA supplementation. Subepithelial bronchial elastic fibers were decreased by approximately 50% in VAD rats and were significantly restored by RA. The increase in AHR that is associated with VAD is accompanied by decreases in M2R expression and function that can be restored by RA and a reduction in airway elastic fibers that can be partially restored by RA.

Topics: Animals; Antineoplastic Agents; Bronchial Hyperreactivity; Bronchoconstrictor Agents; Elasticity; Female; Gene Expression; Methacholine Chloride; Muscarinic Agonists; Pilocarpine; Pulmonary Alveoli; Rats; Rats, Inbred Lew; Receptor, Muscarinic M2; Specific Pathogen-Free Organisms; Tretinoin; Vitamin A Deficiency

Retinoic acid (RA) was recently shown to modify testosterone secretion of the fetal testis in vitro. We characterized this effect by culturing rat testes explanted at various ages, from Fetal Day 14.5 to Postnatal Day 3. In basal medium, RA inhibited, in a dose-dependent manner, both basal and acute LH-stimulated testosterone secretion by testes explanted on Fetal Days 14.5, 15.5, and 16.5. It had no effect on testes from older animals. The negative effect of RA did not result from a diminution in the number of Leydig cells but from a decrease in P450c17 mRNA levels and in LH-stimulated cAMP production. However, the RA-induced decrease in P450C17 mRNA levels was also observed with neonatal testes, suggesting that this enzymatic step is no longer rate limiting at this developmental stage. To study the physiological relevance of RA effects, we used fetuses and neonates issued from mothers fed a vitamin A-deficient (VAD) diet, resulting in a threefold decrease of plasma retinol concentration. On Fetal Day 18.5 and on Posnatal Day 3, testosterone secretion by the testis ex vivo was significantly increased in VAD animals. This shows that the endogenous retinol inhibits differentiation and/or function of fetal Leydig cells before Fetal Day 18.5 and is required for the normal regression of fetal Leydig cell function that occurs after Fetal Day 18.5. In conclusion, our results show that retinoids play a negative role on the steroidogenic activity during the differentiation of rat fetal Leydig cells.

Topics: Animals; Animals, Newborn; Base Sequence; Cell Differentiation; DNA; Female; Fetus; Gestational Age; Leydig Cells; Luteinizing Hormone; Male; Organ Culture Techniques; Pregnancy; Rats; Rats, Sprague-Dawley; RNA, Messenger; Steroid 17-alpha-Hydroxylase; Testis; Testosterone; Tretinoin; Vitamin A Deficiency

Our previous data showed that vitamin A deficiency (VAD) induces, in whole brain, a reduced amount of mRNA for brain retinoic acid (RA) and triiodothyronine (T3) nuclear receptors (i.e., RAR, RXR, and TR, respectively), which is accompanied by reduced amounts of mRNA and protein of neurogranin (RC3, a neuronal protein involved in synaptic plasticity) as well as selective behavioral impairment. Given the important role of retinoids for optimal brain functioning, the effects of vitamin A depletion and subsequent administration of RA or T3 on the mRNA levels of RA and T3 nuclear receptors and on two target genes' (RC3 and neuromodulin or GAP43) mRNA and protein levels were examined in the hippocampus, striatum, and cerebral cortex. A quantitative real-time polymerase chain reaction (PCR), in situ hybridization, and Western blot analysis demonstrated that the striatal region is the brain site where both RA and T3 signaling pathways are most affected by VAD. Indeed, rats fed a vitamin A-free diet for 10 weeks exhibited decreased expression of RAR, RXR, TR, RC3, and GAP43 in the striatum. The administration of T3 to these vitamin A-deprived rats reversed the reduction in mRNA levels of RA and T3 nuclear receptors and in mRNA and protein levels of target genes in this region. These data suggest that modifications that appear preferentially in the striatum, a region highly sensitive to vitamin A bioavailability, may contribute to neurobiological alterations and the spatial learning impairment that occurs in vitamin A-deprived animals.

Topics: Animals; Calmodulin-Binding Proteins; Cerebral Cortex; Corpus Striatum; Down-Regulation; GAP-43 Protein; Gene Expression Regulation; Hippocampus; Male; Nerve Tissue Proteins; Neurogranin; Rats; Rats, Wistar; Receptors, Retinoic Acid; Receptors, Thyroid Hormone; RNA, Messenger; Tretinoin; Triiodothyronine; Vitamin A Deficiency

Our study aimed to investigate, in vivo, the relationship between vitamin A status and NF-kappaB activity, a transcription factor central in regulating inflammatory and immune responses. We used a novel transgenic murine NF-kappaB-luciferase reporter model that enabled molecular imaging of NF-kappaB activity in live mice via an intensified image-capture apparatus. Whole-body luminescence, which reflects overall NF-kappaB activity, was elevated 2.2-fold in vitamin A-deficient (VAD) mice compared with control mice. Specifically, NF-kappaB activity in VAD mice was increased 1.8-fold in the lymph nodes and 1.4-fold in the thymus and, NF-kappaB induction in UVB radiation-exposed skin was also enhanced in VAD mice compared with control mice. The administration of all-trans retinoic acid to VAD mice resulted in a transient reduction in NF-kappaB activity and, conversely, a single dose of the RAR-pan-antagonist, AGN 194310, administered to control mice, led to a marked, transient induction of whole-body luminescence. Our results suggest that vitamin A status, and vitamin A itself, affects NF-kappaB activity in vivo and that the elevated NF-kappaB activity in VAD may be a mechanism underlying some of the features of VAD syndrome.

Topics: Animals; Benzoates; Genes, Reporter; Immunoglobulin kappa-Chains; Luciferases; Lymphoid Tissue; Mice; Mice, Transgenic; NF-kappa B; Skin; T-Lymphocytes; Thiophenes; Transcriptional Activation; Tretinoin; Ultraviolet Rays; Vitamin A Deficiency

We have disrupted the retinoid signalling pathway in adult rats by a dietary deficiency of vitamin A. After 1 year of this dietary deficiency, there was a deposition of amyloid beta in the cerebral blood vessels. There is a downregulation of retinoic acid receptor alpha in the forebrain neurons of the retinoid-deficient rats and a loss of choline acetyl transferase expression, which precedes amyloid beta deposition. In neocortex of pathology samples of patients with Alzheimer's disease, the same retinoic acid receptor alpha deficit in the surviving neurons was observed. We have identified the retinoid-synthesizing enzymes involved in this process, retinaldehyde dehydrogenase-2 and class IV alcohol dehydrogenase, only the former is downregulated in patients with Alzheimer's disease. This suggests that retinoids are important for the maintenance of the adult nervous system and their loss may in part play a role in Alzheimer's disease.

Topics: Aldehyde Oxidoreductases; Alzheimer Disease; Amyloid beta-Peptides; Animals; Brain; Humans; Rats; Rats, Wistar; Receptors, Retinoic Acid; Retinal Dehydrogenase; Retinoic Acid Receptor alpha; Signal Transduction; Tretinoin; Vitamin A Deficiency

Although respiratory tract defects that result from disruption of retinoic acid (RA) signaling have been widely reported, the mechanism by which endogenous RA regulates early lung morphogenesis is unknown. Here, we provide novel evidence that a major role for RA is to selectively maintain mesodermal proliferation and induce fibroblast growth factor 10 (Fgf10) expression in the foregut region where the lung forms. By using a pan-RAR antagonist (BMS493) in foregut explant cultures, we show that bud initiation is selectively blocked in the prospective respiratory region by failure to induce Fgf10 in the corresponding mesoderm. The RA regulation of Fgf10 expression occurs only in this region, within a defined developmental window, and is not seen in other foregut derivatives such as thyroid and pancreas where Fgf10 is also required for normal development. Furthermore, we show that RA activity is essential in the lung field to maintain lung cell identity in the endoderm; RAR antagonism disrupts expression of thyroid transcription factor 1 (Ttf1), an early marker of the respiratory region in the endoderm, and surfactant protein C (Sp-C) mRNAs. Our observations in mouse foregut cultures are corroborated by data from an in vivo model of vitamin A deficiency in rats. Our study supports RA as an essential regulator of gene expression and cellular activities during primary bud formation.

Topics: Animals; Digestive System; Fibroblast Growth Factor 10; Fibroblast Growth Factors; Gene Expression Regulation, Developmental; In Situ Hybridization; Lung; Mice; Mice, Transgenic; Nuclear Proteins; Pancreas; Rats; Rats, Sprague-Dawley; Receptors, Retinoic Acid; RNA, Messenger; Signal Transduction; Thyroid Gland; Thyroid Nuclear Factor 1; Transcription Factors; Tretinoin; Vitamin A Deficiency

Retinoids modulate many physiological processes such as the differentiation and growth of different cell types, including cells from the immune system. We have previously shown that retinoids modulate IgE production in vitro and in vivo. In the present study we investigated the effects of retinoids in non-sensitized and ovalbumin-sensitized mice that were fed for 11 weeks with three different vitamin A (VA) diets: a) VA-deficiency diet, b) base diet, and c) base diet supplemented with 0.5% all-trans-retinoic acid (ATRA). Phorbol-myristate-acetate (PMA)/ionomycin-stimulated SMC (splenic mononuclear cells) from mice fed with ATRA and the vitamin A-deficient diet group showed increased interleukin-4 (IL-4) responses in non-sensitized mice. After ovalbumin sensitization in the VA-deficient and the ATRA supplementation diet groups, no significant effects on IL-4 production were observed. By contrast, gamma interferon (IFN-gamma production from PMA/ionomycin-stimulated SMC was enhanced in the VA-deficient diet group in ovalbumin-sensitized mice, and also in non-sensitized mice compared to the base and the ATRA-supplemented diet group. The data indicate that VA and retinoid content in a diet influences the cytokine response in non-sensitized and also ovalbumin-sensitized mice. Therefore these molecules may serve as active modulators of cytokine production in vivo that are responsible for the induction and persistence of atopic diseases.

Topics: Animals; Cells, Cultured; Cytokines; Diet; Female; Flow Cytometry; Immunization; Interferon-gamma; Interleukin-4; Ionomycin; Lymphocyte Count; Lymphocyte Subsets; Lymphocytes; Mice; Ovalbumin; Retinoids; Spleen; Tetradecanoylphorbol Acetate; Tretinoin; Vitamin A Deficiency

In a prior study, vitamin A-deficient rats subjected to submassive small bowel resections did not mount a normal intestinal adaptive response by 10 days postoperatively, although adaptive increases in crypt cell proliferation were not attenuated and there were no differences in apoptotic indexes. The present study was designed to address the mechanisms by which vitamin A status effects adaptation by analyzing proliferation, apoptosis, and enterocyte migration in the early postoperative period (16 and 48 h) in vitamin A-sufficient, -deficient, and partially replenished sham-resected and resected rats. At 16 h postresection, apoptosis was significantly greater in the remnant ileum of resected vitamin A-deficient rats compared with the sufficient controls. Crypt cell proliferation was increased by resection in all dietary groups at both timepoints. However, at 48 h postresection, proliferation was significantly decreased in the vitamin A-deficient and partially replenished rats. By 48 h after resection, vitamin A deficiency also reduced enterocyte migration rates by 44%. This occurred in conjunction with decreased immunoreactive collagen IV at 48 h and 10 days postoperation. Laminin expression was also reduced by deficiency at 10 days postresection, whereas fibronectin and pancadherin were unchanged at 48 h and 10 days. These studies indicate that vitamin A deficiency inhibits intestinal adaptation following partial small bowel resection by reducing crypt cell proliferation, by enhancing early crypt cell apoptosis, and by markedly reducing enterocyte migration rates, which may be related to changes in the expression of collagen IV and other extracellular matrix components.

Topics: Adaptation, Physiological; Animals; Apoptosis; Cadherins; Cell Division; Cell Movement; Collagen Type IV; Diet; Enterocytes; Female; Fibronectins; Intestines; Laminin; Male; Rats; Rats, Sprague-Dawley; Short Bowel Syndrome; Tretinoin; Vitamin A Deficiency

Recent studies have revealed that retinoids play an important role in the adult central nervous system and cognitive functions. Previous investigations in mice have shown that vitamin A deficiency (VAD) generates a hypo-expression of retinoic acid (RA, the active metabolite of vitamin A) receptors and of neurogranin (RC3, a neuronal protein involved in synaptic plasticity) and a concomitant selective behavioural impairment. Knowing that RC3 is both a triiodothyronine (T3) and a RA target gene, and in consideration of the relationships between the signalling pathways of retinoids and thyroid hormones, the involvement of T3 on RA signalling functionality in VAD was investigated. Thus, the effects of vitamin A depletion and subsequent administration with RA and/or T3 on the expression of RA nuclear receptors (RAR, RXR), T3 nuclear receptor (TR) and on RC3 in the brain were examined. Rats fed a vitamin A-deficient diet for 10 weeks exhibited a decreased expression of RAR, RXR and TR mRNA and of RC3 mRNA and proteins. RA administration to these vitamin A-deficient rats reversed only the RA hypo-signalling in the brain. Interestingly, T3 is able to restore its own brain signalling simultaneously with that of vitamin A and the hypo-expression of RC3. These results obtained in vivo revealed that one of the consequences of VAD is a dysfunction in the thyroid signalling pathway in the brain. This seems of crucial importance since the down regulation of RC3 observed in the depleted rats was corrected only by T3.

Topics: Animals; Blotting, Western; Brain Chemistry; Calmodulin-Binding Proteins; Diterpenes; GTP-Binding Proteins; Liver; Male; Nerve Tissue Proteins; Neurogranin; Protein Glutamine gamma Glutamyltransferase 2; Rats; Rats, Wistar; Receptors, Retinoic Acid; Receptors, Thyroid Hormone; Retinoid X Receptors; Retinyl Esters; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Transcription Factors; Transglutaminases; Tretinoin; Triiodothyronine; Vitamin A; Vitamin A Deficiency

The functional links of specific retinoid receptors to early developmental events in the avian embryo are not known. Before such studies are undertaken, knowledge is required of the spatiotemporal expression patterns of the receptor genes and their regulation by endogenous retinoic acid levels during the early stages of development. Here, we report the expression patterns of mRNAs for RARalpha, RARalpha2, RARbeta2, RARgamma, RARgamma2, RXRalpha, and RARgamma from neurulation to HH10 in the normal and vitamin A-deficient (VAD) quail embryo. The transcripts for all retinoid receptors are detectable at HH5, except for RXRgamma, which is detected at the beginning of HH6. At the 4/5 somite stage of HH8, when retinoid signaling is initiated in the avian embryo, mRNAs of all receptors are present, with very strong and ubiquitous expression patterns for RARalpha, RARalpha2, RARgamma, RARgamma2, and RXRalpha, a persistent expression of RARgamma in the neural tissues, a strong expression of RARbeta2 in lateral plate mesoderm and somites, and an anterior expression of RXRgamma. All retinoid receptors are expressed in the heart primordia. In the VAD quail embryo, the general pattern of retinoid receptor transcript localization is similar to that of the normal, except that the expression of RARalpha2 and RARbeta2 is severely diminished. Administration of retinol or retinoic acid to VAD embryos at or before the 4/5 somite stage rescues the expression of RARalpha2 and RARbeta2 within approximately 45 min and restores normal development. RARbeta2 expression requires the expression of RARalpha2. After neurulation, the expression of all retinoid receptors in the VAD quail embryo becomes independent of vitamin A status and is similar to that of the normal. The mRNA levels and sites of expression of the key enzyme for retinoic acid biosynthesis, Raldh-2, are not affected by vitamin A status; the expression pattern is restricted and does not correspond to that of retinoid receptors at all sites. The general patterns and intensity of retinoid receptor gene expression during early quail development are comparable to those of the mammalian and thus validate the application of results from retinoid-regulated avian development studies to those of the mammalian.

Topics: Aldehyde Oxidoreductases; Animals; Coturnix; Embryo, Nonmammalian; Fetal Death; Gene Expression Regulation, Developmental; Mice; Receptors, Retinoic Acid; Retinal Dehydrogenase; Retinoic Acid Receptor alpha; Retinoic Acid Receptor gamma; Retinoid X Receptors; Transcription Factors; Transcription, Genetic; Tretinoin; Vitamin A Deficiency

Vitamin A and its derivatives, the retinoids, have recently been reported to be implicated in the synaptic plasticity of the hippocampus and in cognitive functions. Acting via transcription factors, retinoids can regulate gene expression via their nuclear receptors [retinoic acid receptors (RARs) and retinoid X receptors (RXRs)]. We recently showed that a moderate (about 30%) hypoexpression of brain (and hippocampal) retinoid signalling, like that naturally occurring in the aged brain of mice, might be related to a selective relational memory deficit. To further assess this hypothesis, the present study investigated the effects of Vitamin A deprivation of varying duration both on the brain expression of retinoid receptors (RARbeta and RXRbeta/gamma) and two associated target genes [tissue-type transglutaminase (tTG) and neurogranin, (RC3)], and on radial maze discrimination learning using young adult mice as subjects. We observed that irrespective of its duration (i.e. 31 or 39 weeks), Vitamin A deprivation resulted in a significant reduction (25-30%) in the expression of brain RARbeta, RXRbeta/gamma and tTG mRNAs. Conversely, only the 39-week condition was found to induce a significant decrease in brain RC3 mRNAs contents and a selective relational memory impairment. Finally, daily administration of retinoic acid (RA) failed to reverse the 39-week Vitamin A deficiency (VAD)-related cognitive deficit and to fully normalise the associated brain retinoid hyposignalling. In particular, there was no evidence for an up-regulating effect of RA on whole brain (and hippocampal) RC3 mRNAs of the 39-week-depleted mice. The results show that post-natal VAD may induce a selective memory impairment and give further support to the hypothesis that the fine regulation of retinoid-mediated gene expression is important for optimal brain functioning and higher cognition.

Topics: Animals; Behavior, Animal; Brain; Brain Chemistry; Calmodulin-Binding Proteins; Choice Behavior; Discrimination Learning; Male; Memory Disorders; Mice; Mice, Inbred C57BL; Nerve Tissue Proteins; Neurogranin; Reaction Time; Receptors, Retinoic Acid; Retinoids; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Time Factors; Transglutaminases; Tretinoin; Vitamin A Deficiency

The retinamide, N-(4-hydroxyphenyl)retinamide (4-HPR), has shown promising anti-tumor activity, but it is unclear whether this compound is hydrolyzed to all-trans retinoic acid (atRA) and if so, whether this plays any role in its chemotherapeutic activity. To address this issue, the ability of 4-hydroxybenzylretinone (4-HBR), a carbon-linked analog of 4-HPR, to support growth in vitamin A-deficient (VAD) animals and to activate an atRA-responsive gene in vivo was compared to 4-HPR and atRA. Further, the non-hydrolyzable 4-HBR analog was used to determine whether the presence of the labile amide linkage in 4-HPR is essential for the induction of apoptosis in cultured MCF-7 breast cancer cells. Studies in VAD rats showed that 4-HPR, like atRA, supports animal growth and induces CYP26B1 mRNA expression in lung whereas 4-HBR does not. Analysis of plasma from 4-HPR- and atRA-treated VAD animals revealed the presence of atRA whereas it was not detected in plasma from animals given 4-HBR. To determine whether hydrolysis to atRA is necessary for apoptosis induced by 4-HPR in MCF-7 breast cancer cells, morphological and biochemical assays for apoptosis were performed. 4-HBR, like 4-HPR, induced apoptosis in MCF-7 cells. Apoptosis was not induced even at high concentrations of atRA, showing that 4-HPR and 4-HBR act in cells via a distinct signaling pathway. These results show that although limited hydrolysis of 4-HPR occurs in vivo, the ability to liberate atRA is not required for these 4-hydroxyphenyl retinoids to induce apoptosis in MCF-7 breast cancer cells. Thus the non-hydrolyzable analog, 4-HBR, may have significant therapeutic advantage over 4-HPR because it does not liberate atRA that can contribute to the adverse side effects of drug administration in vivo.

Topics: Administration, Oral; Animals; Apoptosis; Body Weight; Breast Neoplasms; Cell Line, Tumor; Cell Survival; Dose-Response Relationship, Drug; Fenretinide; Humans; Hydrolysis; Male; Rats; Rats, Sprague-Dawley; Tretinoin; Vitamin A; Vitamin A Deficiency

We have examined the role of the signalling molecule, retinoic acid, in the process of neurulation and the subsequent growth and differentiation of the central nervous system using quail embryos that have developed in the absence of retinoic acid. Such retinoic acid-free embryos undergo abnormal neural tube formation in terms of its shape and structure, but the embryos do not display spina bifida or exencephaly. The neural tubes have a wider floor plate, a thicker roof plate and a different dorsoventral shape. Phalloidin staining and electron microscopy revealed alterations in the actin filaments and the junctional complexes of the cell layer lining the lumen. Initially the neural tubes proliferated at the same rate as normal, but later the proliferation rate declined drastically and neuronal differentiation was highly deficient. There were very few motoneurons extending neurites into the periphery, and within the neural tube axon trajectories were chaotic. These results reveal several functions for retinoic acid in the morphogenesis and growth of the neural tube, many of which can be explained by defective notochord signalling, but they do not suggest that this molecule plays a role in neural tube closure.

Topics: Actin Cytoskeleton; Animals; Apoptosis; Cell Division; Central Nervous System; In Situ Hybridization; Microscopy, Electron; Models, Animal; Morphogenesis; Notochord; Quail; Signal Transduction; Staining and Labeling; Tretinoin; Vitamin A Deficiency

Phosphoenolpyruvate carboxykinase (PEPCK) gene expression is decreased in vitamin A-deficient (VAD) mice. However, the underlying molecular mechanism at the PEPCK promoter that contributes to this alteration in gene expression remains unexplained and thus serves as the basis for our investigation in this report. Using liver from vitamin A-sufficient (VAS) and VAD mice in the chromatin immunoprecipitation (ChIP) assay, we determined that histones H3 and H4 were in the acetylated or active state in VAS mice at each of the three retinoic acid response elements (RARE1, RARE2 and RARE3) of the PEPCK promoter. The same acetylation pattern was seen in VAD mice, but with relatively lower levels of acetylated H3 and H4 bound at the region encompassing PEPCK RARE1/RARE2. In ChIP assays conducted with an antibody to RNA polymerase II (RNA Pol II), the association of RNA Pol II with PEPCK RARE1/RARE2 was significantly decreased in vitamin A deficiency. The reduction in RNA Pol II association is indicative of an interruption in the direct interactions of RNA Pol II with the PEPCK promoter, with general transcription factors and/or with coregulator molecules that contribute to the activation of the PEPCK gene. These results increase our understanding of the molecular basis for decreased PEPCK gene expression in VAD mice in vivo and offer additional insight into the regulation of other retinoid-responsive genes.

Topics: Acetylation; Alitretinoin; Animals; Chromatin; Drug Combinations; Food Deprivation; Histones; Mice; Mice, Inbred C57BL; Phosphoenolpyruvate Carboxykinase (GTP); Precipitin Tests; Promoter Regions, Genetic; Response Elements; RNA Polymerase II; Tretinoin; Vitamin A Deficiency

Retinyl ester, the most abundant form of vitamin A (retinol), is synthesized by the enzyme lecithin:retinol acyltransferase (LRAT). Previously, we cloned a 2.5 kb LRAT cDNA from rodent liver which codes for functional LRAT activity. However, Northern blots of tissues probed with the 2.5 kb cDNA revealed the presence of a larger transcript of approximately 5 kb as well as several smaller transcripts. To elucidate the nature of the large LRAT transcript, a high-molecular-mass adrenal gland cDNA library was screened. Two similar clones of 3962 and 3187 nt were identified which appeared to be part of the 3'-untranslated region (UTR) of a 5358 nt LRAT mRNA. The 5.3 kb cDNA was then amplified from liver by reverse transcriptase PCR (RT-PCR) and demonstrated to encode functional LRAT activity. The 3'-UTR of the 5.3 kb cDNA contains several AAUAAA polyadenylation signals. Analysis of the 3' ends of LRAT mRNA transcripts from liver, intestine and testis showed the usage of both canonical and non-canonical polyadenylation signals. To further analyse the LRAT mRNAs expressed in vivo, Northern blot analysis was performed using four probes spanning sections from the 5' end to the distal 3' end of the 5.3 kb LRAT cDNA. The results show that the major 5.3 kb transcript uses the canonical signal AAUAAA located at nt 5308, and the major short transcript of approximately 1.5 kb uses the non-canonical signal AUUAAA located at nt 1330. The 5.3 kb LRAT transcript was predominant in the liver of retinoic acid-repleted vitamin A-deficient rats, coincident with increased quantitative expression of LRAT mRNA and enzyme activity. The differential usage of these polyadenylation signals can explain the presence of multiple LRAT mRNA transcripts which are expressed in different tissue-specific patterns.

Topics: 3' Untranslated Regions; Acyltransferases; Age Factors; Amino Acid Sequence; Animals; Base Sequence; Cloning, Molecular; Female; Gene Expression Regulation; Humans; Liver; Male; Molecular Sequence Data; Polyadenylation; Protein Biosynthesis; Rats; Rats, Inbred Lew; RNA, Messenger; Tretinoin; Vitamin A Deficiency

Vitamin A (VA) and insulin-like growth factors (IGF) are important regulators of a wide range of physiological processes. To investigate the IGF system's involvement in the physiological actions of VA, we examined the effects of VA status on components of the IGF system in rats. Male rats (3-wk-old) fed a VA-deficient diet for 11 wk developed VA deficiency, as confirmed by the depletion of serum retinol and hepatic retinyl palmitate. Rats fed the VA-deficient diet had significantly lower body weight (p < 0.05) and lower serum IGF-I concentrations than the rats fed the control diet. The decreases in serum IGF-I levels were accompanied by approximately 40% lower levels of the IGF-I mRNA in the liver and lungs. With respect to the gene expression of other IGF system components, VA deficiency caused a twofold induction of IGF-I receptor (IGF-IR) mRNA in the heart and a twofold reduction in IGFBP-6 mRNA in the lungs, but did not alter the expression of IGF-II, IGFBP-1, IGFBP-3, IGFBP-4 or IGFBP-5 in all tissues examined. When VA-deficient rats received a single injection of retinoic acid (2 mg/rat), tissue IGF-I and IGF-IR gene expression did not change after 4 or 8 h, while the expression of IGF-II, IGFBP-4, and IGFBP-6 mRNAs in some tissues increased rapidly. These results suggest a possible involvement of the IGF system in mediating the physiological actions of VA, including VA-supported growth, in the rat.

Topics: Animals; Blotting, Northern; Diterpenes; Gene Expression Regulation; Insulin-Like Growth Factor Binding Proteins; Male; Random Allocation; Rats; Rats, Wistar; Retinyl Esters; RNA, Messenger; Somatomedins; Tissue Distribution; Tretinoin; Vitamin A; Vitamin A Deficiency

Retinoic acid (RA) is essential for cellular growth and differentiation in developing and adult animals. The central nervous system (CNS) suffers developmental defects if embryonic levels of RA are too high or too low. The production and function of RA in adult brain are unclear. We report that RA is present throughout the brain and spinal cord of adult, vitamin A-deficient (VAD) rats treated with a physiological amount of all-trans-retinol. The hippocampus/cortex contained the highest proportion of RA in the brain (27.2 +/- 2.9% of the organic phase radioactivity, and 23.5 +/- 0.8% of the organic phase radioactivity extracted from spinal cord was RA). RA comprises a higher proportion of the retinoid pool in the CNS compared with amounts reported in other target tissues (E Werner and HF DeLuca. Arch Biochem Biophys 393: 262-270, 2001). However, RA is not preferentially transported from the blood to the brain. There were 2.90 +/- 0.20 fmol RA/g tissue transported to the brain of VAD rats treated with 2.00 nmol [20-(3)H]all-trans-retinoic acid, but higher amounts of RA were delivered to the liver, testis, and spleen. Because RA is not transported preferentially to brain, this tissue likely synthesizes RA more efficiently than other target tissues.

Topics: Animals; Biological Transport; Blood-Brain Barrier; Brain; Brain Chemistry; Brain Stem; Carbon Radioisotopes; Cerebellum; Cerebral Cortex; Chromatography, High Pressure Liquid; Hippocampus; Inulin; Liver; Male; Rats; Rats, Sprague-Dawley; Spinal Cord; Spleen; Testis; Tretinoin; Tritium; Vitamin A; Vitamin A Deficiency; Weaning

The ability of class I alcohol dehydrogenase (ADH1) and class IV alcohol dehydrogenase (ADH4) to metabolize retinol to retinoic acid is supported by genetic studies in mice carrying Adh1 or Adh4 gene disruptions. To differentiate the physiological roles of ADH1 and ADH4 in retinoid metabolism we report here the generation of an Adh1/4 double null mutant mouse and its comparison to single null mutants. We demonstrate that loss of both ADH1 and ADH4 does not have additive effects, either for production of retinoic acid needed for development or for retinol turnover to minimize toxicity. During gestational vitamin A deficiency Adh4 and Adh1/4 mutants exhibit completely penetrant postnatal lethality by day 15 and day 24, respectively, while 60% of Adh1 mutants survive to adulthood similar to wild-type. Following administration of a 50-mg/kg dose of retinol to examine retinol turnover, Adh1 and Adh1/4 mutants exhibit similar 10-fold decreases in retinoic acid production, whereas Adh4 mutants have only a slight decrease. LD(50) studies indicate a large increase in acute retinol toxicity for Adh1 mutants, a small increase for Adh4 mutants, and an intermediate increase for Adh1/4 mutants. Chronic retinol supplementation during gestation resulted in 65% postnatal lethality in Adh1 mutants, whereas only approximately 5% for Adh1/4 and Adh4 mutants. These studies indicate that ADH1 provides considerable protection against vitamin A toxicity, whereas ADH4 promotes survival during vitamin A deficiency, thus demonstrating largely non-overlapping functions for these enzymes in retinoid metabolism.

Topics: Alcohol Dehydrogenase; Animals; Chromatography, High Pressure Liquid; Female; Genetic Vectors; Genotype; Heterozygote; Male; Mice; Mice, Transgenic; Models, Genetic; Mutation; Time Factors; Tretinoin; Vitamin A; Vitamin A Deficiency

Influence of vitamin A (retinol) on growth depends on its sequential oxidation to retinal and then to retinoic acid (RA), producing a ligand for RA receptors essential in development of specific tissues. Genetic studies have revealed that aldehyde dehydrogenases function as tissue-specific catalysts for oxidation of retinal to RA. However, enzymes catalyzing the first step of RA synthesis, oxidation of retinol to retinal, remain unclear because none of the present candidate enzymes have expression patterns that fully overlap with those of aldehyde dehydrogenases during development. Here, we provide genetic evidence that alcohol dehydrogenase (ADH) performs this function by demonstrating a role for Adh3, a ubiquitously expressed form. Adh3 null mutant mice exhibit reduced RA generation in vivo, growth deficiency that can be rescued by retinol supplementation, and completely penetrant postnatal lethality during vitamin A deficiency. ADH3 was also shown to have in vitro retinol oxidation activity. Unlike the second step, the first step of RA synthesis is not tissue-restricted because it is catalyzed by ADH3, a ubiquitous enzyme having an ancient origin.

Topics: Alcohol Dehydrogenase; Alcohol Oxidoreductases; Animals; Cytosol; Genotype; Mice; Mice, Transgenic; Mutation; Oxygen; Retinaldehyde; Time Factors; Tretinoin; Vitamin A; Vitamin A Deficiency

Lecithin:retinol acyltransferase (LRAT), a retinol esterifying enzyme, plays a major role in the metabolism and storage of vitamin A in several animal tissues. Groups of vitamin A (VA)-adequate (control) and VA-deficient rats were treated with vehicle or 5 mg of all-trans-retinoic acid (RA); an additional group of VA-deficient rats were fed 100 microg of RA. In control rats, lung LRAT mRNA and LRAT specific activity were approximately 50% of the levels expressed in the liver. In the lung of VA-deficient rats, LRAT mRNA and specific activity levels were <10% of those in the control group. Treatment of VA-deficient rats with 100 microg RA increased lung LRAT mRNA (P < 0.005) and specific activity (P < 0.0001), and treatment with 5 mg of RA increased LRAT mRNA level and specific activity more than approximately 15- and 6-fold above those in control lung, respectively (both P < or = 0.001). The lung tissue of VA-adequate rats contained retinyl ester (approximately 3 nmol/g tissue), whereas none was detected in the lung tissue of VA-deficient rats. These results show that LRAT expression and vitamin A storage are regulated by vitamin A status and by treatment with all-trans-RA in the adult lung. These results suggest that the regulated storage of vitamin A may be important for maintaining the integrity and physiologic functions of the lung.

Topics: Acyltransferases; Animals; Enzyme Induction; Gene Expression Regulation, Enzymologic; Liver; Lung; Rats; Rats, Inbred Lew; RNA, Messenger; Tretinoin; Vitamin A; Vitamin A Deficiency

A novel retinoic acid (RA)-inducible cytochrome P450 (P450 RAI or CYP26), previously cloned from human, zebra fish, and mouse, functions in the metabolism of all-trans-RA to polar metabolites including 4-hydroxy-RA and 4-oxo-RA. To further study CYP26 in the rat model, we first cloned rat CYP26 cDNA. The nucleotide sequence predicts a 497-amino-acid protein whose sequence is 95% identical to mouse and 91% homologous to human CYP26. Animal studies showed that CYP26 mRNA expression is very low (0.01+/-0.008;P<0.05) in vitamin-A-deficient rats compared to pair-fed vitamin-A-sufficient rats (defined as 1.0). In a kinetic study, vitamin-A-deficient rats were treated with approximately 100 microg of all-trans-RA and liver was collected after 3-72 h for analysis of CYP26 mRNA by quantitative real-time PCR. Liver CYP26 mRNA increased to nearly 10-fold above control after 3 h (P<0.01), reaching a peak of about 2000-fold greater around 10 h (P<0.001) and then decreased rapidly. The CYP26 dose response to RA was nearly linear (R(2)=0.9638). Additionally, significant regulation of CYP26 gene expression was observed in the vitamin-A-deficient, control, and RA-treated condition in lung, testis, and small intestine. We conclude that CYP26 mRNA expression is dynamically regulated in vivo by diet and RA in hepatic and extrahepatic tissues. The long-term down-regulation of CYP26 in retinoid deficiency may be critical for conserving RA, while the acute up-regulation of CYP26 may be important for preventing a deleterious overshoot of RA derived from either dietary or exogenous sources.

Topics: Amino Acid Sequence; Animals; Base Sequence; Cloning, Molecular; Cytochrome P-450 Enzyme System; DNA, Complementary; Gene Expression Regulation, Enzymologic; Humans; Liver; Male; Mice; Mixed Function Oxygenases; Molecular Sequence Data; Rats; Rats, Inbred Lew; Retinoic Acid 4-Hydroxylase; RNA, Messenger; Sequence Homology, Amino Acid; Tissue Distribution; Tretinoin; Vitamin A Deficiency

Retinoic acid-mediated gene activation is important for normal vertebrate development. The size and nature of retinoic acid make it difficult to identify the precise cellular location of this signaling molecule throughout an embryo. Additionally, retinoic acid (RA) signaling is regulated by a complex combination of receptors, coactivators, and antagonizing proteins. Thus, in order to integrate these signals and identify regions within a whole developing embryo where cells can respond transcriptionally to retinoic acid, we have used a reporter transgenic approach. We have generated several stable lines of transgenic zebrafish which use retinoic acid response elements to drive fluorescent protein expression. In these zebrafish lines, transgene expression is localized to regions of the neural tube, retina, notochord, somites, heart, pronephric ducts, branchial arches, and jaw muscles in embryos and larvae. Transgene expression can be induced in additional regions of the neural tube and retina as well as the immature notochord, hatching gland, enveloping cell layer, and fin by exposing embryos to retinoic acid. Treatment with retinoic acid synthase inhibitors, citral and diethylaminobenzaldehyde (DEAB), during neurulation, greatly reduces transgene expression. DEAB treatment of embryos at gastrulation phenocopies the embryonic effects of vitamin A deprivation or targeted disruption of the RA synthase retinaldehyde dehydrogenase-2 in other vertebrates. Together these data suggest that the reporter expression we see in zebrafish is dependent upon conserved vertebrate pathways of RA synthesis.

Topics: Acyclic Monoterpenes; Aldehyde Oxidoreductases; Animals; Benzaldehydes; Cell Communication; Cytochrome P-450 Enzyme Inhibitors; Gastrula; Gene Expression Regulation, Developmental; Genes, Reporter; Genetic Engineering; Monoterpenes; Oxygenases; Promoter Regions, Genetic; Retinal Dehydrogenase; Teratogens; Terpenes; Transcriptional Activation; Transgenes; Tretinoin; Vitamin A Deficiency; Zebrafish; Zebrafish Proteins

Acute promyelocytic leukemia (APL) is always associated with chromosomal translocations that disrupt the retinoic acid receptor alpha (RARalpha) gene. Whether these translocations relate to a role for endogenous RARalpha in normal granulopoiesis remains uncertain because most studies addressing this question have used non-physiological overexpression systems. Granulocyte differentiation in cells derived from RARalpha-deficient (RARalpha(-/-)) mice was studied and evaluated in the context of agonist-bound and ligand-free RARalpha. Our results demonstrate that RARalpha is dispensable for granulopoiesis, as RARalpha(-/-) mice have a normal granulocyte population despite an impaired ability to respond to retinoids. However, although it is not absolutely required, RARalpha can bidirectionally modulate granulopoiesis. RARalpha stimulates differentiation in response to exogenous retinoic acid. Furthermore, endogenous retinoids control granulopoiesis in vivo, as either vitamin A-deficient mice or animals treated with an RAR antagonist accumulate more immature granulocytes in their bone marrow. Conversely, RARalpha acts to limit differentiation in the absence of ligand because granulocyte precursors from RARalpha(-/-) mice differentiate earlier in culture. Thus, the block in granulopoiesis exerted by RARalpha fusion proteins expressed in APL cells may correspond to an amplification of a normal function of unliganded RARalpha.

Topics: Animals; Bone Marrow Cells; Cell Culture Techniques; Cell Differentiation; Granulocytes; Mice; Mice, Mutant Strains; Receptors, Retinoic Acid; Retinoic Acid Receptor alpha; Retinoids; Retinol-Binding Proteins; Retinol-Binding Proteins, Cellular; Tretinoin; Vitamin A Deficiency

The effect of retinoids on the expression of kappa opioid receptor (KOR) gene was examined in normal and transgenic animals. KOR-lacZ transgene expression was specifically elevated in KOR-positive areas of the developing CNS by depleting vitamin A from animal diets. The endogenous KOR mRNA species, including all three isoforms, were also upregulated by depleting vitamin A in developing animals. Change in the expression of isoforms a and b is similar in prenatal stages but differs during postnatal development. Interestingly, upregulation of isoform c is most significant postnatally. The regulation of KOR gene by vitamin A was substantiated in a mouse embryonal carcinoma P19 culture system in which retinoic acid (RA), the most potent ingredient of vitamin A, was able to suppress the expression of all the three KOR isoforms and KOR protein. The RA-mediated suppression was blocked by an RA receptor antagonist and a histone deacetylase (HDAC) inhibitor. By using a reporter transfection assay in P19 cells, the potential genetic element responsible for RA-mediated suppression of KOR gene expression was located to intron 1 of the mouse KOR gene, which could also be blocked by HDAC inhibitor. Furthermore, suppression of KOR gene expression by RA in P19 cells appeared to be an indirect event and required protein synthesis. A role of RA in KOR gene regulation during developmental stages was discussed.

Topics: Animals; Diet; Embryonal Carcinoma Stem Cells; Enzyme Inhibitors; Female; Gene Expression Regulation; Genes, Reporter; Histone Deacetylase Inhibitors; Introns; Male; Mice; Mice, Transgenic; Neoplastic Stem Cells; Organ Specificity; Protein Isoforms; Receptors, Opioid, kappa; Receptors, Retinoic Acid; Recombinant Fusion Proteins; Regulatory Sequences, Nucleic Acid; RNA, Messenger; Transfection; Transgenes; Tretinoin; Tumor Cells, Cultured; Vitamin A Deficiency

To gain insight into the in vivo modulation of the expression of the adipogenic transcription factors PPAR gamma 2, C/EBP alpha, and ADD1/SREBP1c by retinoids and its relationship with whole-body adiposity.. Three-week-old mice were fed with standard chow or a vitamin A-deficient diet for 10 weeks. During the 4 days immediately before they were killed, the animals were treated either with all-trans retinoic acid (tRA; 100 mg/kg per day, subcutaneously) or vehicle. The specific levels of the mRNAs for the three transcription factors were analyzed in epididymal white adipose tissue (eWAT) and inguinal white adipose tissue and in brown adipose tissue (BAT). Other parameters determined were leptin and UCP2 levels in white adipose tissue depots, total cholesterol and triglyceride serum levels, energy intake, body weight, and adiposity.. Vitamin A-deficient diet feeding led to a marked increase of adiposity and to a small increase of body weight. Hypertrophy of white adipose tissue depots correlated with enhanced PPAR gamma 2 expression. Hypertrophy of BAT, in contrast, correlated with a decrease of PPAR gamma 2 expression that may contribute to the known reduced thermogenic potential of BAT under conditions of vitamin A restriction. Treatment with tRA triggered a reduction of adiposity and body weight that correlated with a down-regulation of PPAR gamma 2 expression in all adipose tissues. The effects of tRA were more pronounced in eWAT, where C/EBP alpha and ADD1/SREBP1c levels were also reduced. The response to tRA was impaired in the eWAT and BAT of animals fed the vitamin A-deficient diet.. The results emphasize the importance of retinoids as physiological regulators of adipose tissue development and function in intact animals.

Topics: Adipose Tissue; Adipose Tissue, Brown; Animals; Body Weight; Cholesterol; Energy Intake; Gene Expression; Gene Expression Regulation; Ion Channels; Male; Membrane Transport Proteins; Mice; Mitochondrial Proteins; Nutritional Status; Proteins; Receptors, Cytoplasmic and Nuclear; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Transcription Factors; Tretinoin; Triglycerides; Uncoupling Agents; Uncoupling Protein 2; Vitamin A; Vitamin A Deficiency

Because only retinol and not all-trans-retinoic acid (atRA) can satisfy all of the functions of vitamin A, we have investigated the retinol metabolites in tissues of vitamin A-deficient (VAD) rats responding to a radioactive dose of [20-(3)H]all-trans-retinol. As expected, atRA is the major vitamin A metabolite present in the target tissues of VAD rats given a physiological dose (1 microg) of [20-(3)H]all-trans-retinol (atROL). Both atROL and atRA were detected by high-performance liquid chromatographic (HPLC) analysis of the radioactivity extracted from the liver, kidney, small intestine, lung, spleen, bone, skin, or testis of these animals. Novel retinol metabolites were observed in the aqueous extracts from the testis, lung, and skin. However, these metabolites were detected in very small amounts and were not characterized further. Importantly, neither 9-cis-retinoic acid (9cRA), 9-cis-retinol (9cROL), nor 13-cis-retinoic acid (13cRA) was present in detectable amounts. The amounts of atRA varied in each tissue, ranging from 0.29 +/- 0.05 fmol of RA/g of tissue in the femurs to 12.9 +/- 4.3 fmol of RA/g of tissue in the kidneys. The absence of 9cRA in vivo was not due to degradation of this retinoid during the extraction procedure or HPLC analysis of the extracted radioactivity. As atROL completely fulfills all of the physiological roles of vitamin A, and 9cRA is not detected in any of the tissues analyzed, these results suggest that 9cRA may have no physiological relevance in the rat.

Topics: Alitretinoin; Animals; Chromatography, High Pressure Liquid; Lung; Male; Protein Isoforms; Rats; Rats, Sprague-Dawley; Retinoids; Testis; Tissue Distribution; Tretinoin; Vitamin A; Vitamin A Deficiency

The vitamin A deficient (VAD) quail embryo lacks active retinoids, fails to express normally GATA-4, and develops a nonlooping heart tube morphogenetic defect that is a model for congenital cardiomyopathy. VAD quail embryos, or chick embryos depleted specifically for GATA factors, show in addition abnormal foregut development, characterized by apoptosis of the endoderm cells associated with presumptive myocardium during the process of heart tube formation. Exogenous retinoic acid or transplantation of normal chick embryo anterior endoderm is sufficient to rescue apoptosis as well as GATA-4 expression and results in normal development and heart tube morphogenesis. Normal posterior endoderm also contains retinoids but is unable to rescue the VAD defect. Our results indicate that a retinoid-dependent transcriptional program, mediated at least in part by GATA factors, is critical in presumptive foregut endoderm for normal heart tube morphogenesis.

Topics: Animals; Apoptosis; Chick Embryo; Chimera; Digestive System; DNA-Binding Proteins; Endoderm; GATA4 Transcription Factor; GATA5 Transcription Factor; GATA6 Transcription Factor; Gene Expression; Heart; Heart Defects, Congenital; In Situ Hybridization; Phenotype; Quail; RNA, Messenger; Transcription Factors; Transplantation, Heterologous; Tretinoin; Vitamin A Deficiency

The developing nervous system is particularly vulnerable to vitamin A deficiency. Retinoid has been proposed to be a posteriorizing factor during hindbrain development, although direct evidence in the mammalian embryo is lacking. In the present study, pregnant vitamin A-deficient (VAD) rats were fed purified diets containing varying levels of all-trans-retinoic acid (atRA; 0, 0.5, 1.5, 6, 12, 25, 50, 125, or 250 microg/g diet) or were supplemented with retinol. Hindbrain development was studied from embryonic day 10 to 12.5 ( approximately 6 to 40 somites). Normal morphogenesis was observed in all embryos from groups fed 250 microg atRA/g diet or retinol. The most caudal region of the hindbrain was the most sensitive to retinoid insufficiency, as evidenced by a loss of the hypoglossal nerve (cranial nerve XII) in embryos from the 125 microg atRA/g diet group. Further reduction of atRA to 50 microg/g diet led to the loss of cranial nerves IX, X, XI, and XII and associated sensory ganglia IX and X in all embryos as well as the loss of hindbrain segmentation caudal to the rhombomere (r) 3/4 border in a subset of embryos. Dysmorphic orthotopic otic vesicles or immature otic-like vesicles in both orthotopic and caudally ectopic locations were also observed. As the level of atRA was reduced, a loss of caudal hindbrain segmentation was observed in all embryos and the incidence of otic vesicle abnormalities increased. Perturbations in hindbrain segmentation, cranial nerve formation, and otic vesicle development were associated with abnormal patterning of the posterior hindbrain. Embryos from VAD dams fed between 0.5 and 50 microg atRA/g diet exhibited Hoxb-1 protein expression along the entire neural tube caudal to the r3/r4 border at a time when it should be restricted to r4. Krox-20 protein expression was expanded in r3 but absent or reduced in presumptive r5. Hoxd-4 mRNA expression was absent in the posterior hindbrain, and the rostral limit of Hoxb-5 protein expression in the neural tube was anteriorized, suggesting that the most posterior hindbrain region (r7/r8) had been deleted and/or improperly patterned. Thus, when limiting amounts of atRA are provided to VAD dams, the caudal portion of the hindbrain is shortened and possesses r4/r5-like characteristics, with this region finally exhibiting r4-like gene expression when retinoid is restricted even more severely. Thus, regions of the anterior hindbrain (i.e., r3 and r4) appear to be greatly expanded, wherea

Topics: Animals; Biomarkers; Cranial Nerves; DNA-Binding Proteins; Dose-Response Relationship, Drug; Ear; Early Growth Response Protein 2; Homeodomain Proteins; Immunohistochemistry; In Situ Hybridization; Models, Biological; Rats; Rats, Sprague-Dawley; Rhombencephalon; Time Factors; Transcription Factors; Tretinoin; Vitamin A; Vitamin A Deficiency

Retinoic acid (RA) is necessary for normal differentiation of the tail bud into the secondary neural tube. Excess RA, however, is teratogenic and causes neural tube defects (NTDs). The way in which RA modulates secondary neurulation is unclear but probably involves RA-regulated downstream genes such midkine (MK), which encodes a growth factor implicated in tail bud mesenchymal-neuroepithelial conversion. Our objective was to determine whether RA-deficiency would produce similar defects and if MK is involved.. Citral, a drug that blocks endogenous RA formation, as well as a neutralizing antibody, were used to block RA activity in chick embryos. Immunohistochemistry and in situ hybridization were used to localize RA and MK in the tail bud. Competitive RT-PCR was used to examine the effects of excess RA and RA deficiency due to citral on the expression of MK mRNA.. Citral-induced NTDs displayed a morphological resemblance to those caused by excess RA. However, citral treatment did not significantly increase embryonic mortality, and RA rescue of citral-treated embryos proved unsuccessful. MK mRNA was detected in the differentiating tail bud by in situ hybridization. Competitive RT-PCR showed that excess RA decreased MK expression by 60%. Doses of citral that caused a comparable incidence of defects, however, caused only a 25% decrease.. The results show that excess RA and RA deficiency both cause defects of secondary neurulation. While excess RA decreased MK expression, RA deficiency had minimal effects. However, whether or not MK is an intermediary in the developmental phenomena regulated physiologically or pathologically by RA remains to be elucidated.

Topics: Abnormalities, Drug-Induced; Acyclic Monoterpenes; Animals; Carrier Proteins; Chick Embryo; Cytokines; Dose-Response Relationship, Drug; Immunohistochemistry; In Situ Hybridization; Keratolytic Agents; Midkine; Monoterpenes; Nerve Growth Factors; Neural Tube Defects; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Tail; Teratogens; Terpenes; Time Factors; Tretinoin; Vitamin A Deficiency

Antibody responses to T cell-dependent antigens are reduced during vitamin A (VA) deficiency and restored by retinoids. To test whether retinoic acid (RA) and polyinosinic:polycytidylic acid (PIC), an inducer of interferons, can increase specific antibody production, VA-deficient rats were treated with all-trans-RA, PIC, or both at the time of primary immunization with tetanus toxoid. VA-deficient rats produced low primary and secondary anti-tetanus IgG responses (P<.001 vs. VA-sufficient controls). Both responses were increased synergistically by RA plus PIC (P<.0001). In VA-deficient spleens, mRNAs were low for interleukin (IL)-2 receptor-beta, interferon regulatory factor-1, and signal transducer and activator of transcription 1. Each, however, was induced by RA plus PIC (P<.0001 vs. controls). Conversely, IL-12 and IL-10 mRNAs were elevated in VA deficiency and were induced by PIC and suppressed by RA. Thus, RA plus PIC appears to be a promising combination for stimulating antigen-specific immunity. Several molecular factors identified here may partially account for the observed enhancement.

Topics: Animals; Antibody Formation; DNA-Binding Proteins; Drug Synergism; Female; Gene Expression Regulation; Immunoglobulin G; Interferon Regulatory Factor-1; Lactation; Male; Phosphoproteins; Poly I; Rats; Rats, Inbred Lew; Receptors, Chemokine; Receptors, Interleukin; Receptors, Interleukin-8B; RNA, Messenger; Signal Transduction; Spleen; STAT1 Transcription Factor; Tetanus Toxoid; Trans-Activators; Transcription Factors; Transcription, Genetic; Tretinoin; Vitamin A Deficiency

The relationship between interscapular brown adipose tissue (IBAT) thermogenic potential and vitamin A status was investigated by studying the effects of feeding a vitamin A-deficient diet and all-trans retinoic acid (tRA) treatment on body weight and IBAT parameters in mice. Feeding a vitamin A-deficient diet tended to trigger opposite effects to those of tRA treatment, namely increased body weight, IBAT weight, adiposity and leptin mRNA expression, and reduced IBAT thermogenic potential in terms of uncoupling protein 1 (UCP1) mRNA and UCP2 mRNA expression. The results emphasize the importance of retinoids as physiological regulators of brown adipose tissue.

Topics: Adipose Tissue, Brown; Animals; Blotting, Northern; Body Temperature Regulation; Body Weight; Carrier Proteins; Hypertrophy; Ion Channels; Leptin; Male; Membrane Proteins; Membrane Transport Proteins; Mice; Mice, Inbred Strains; Mitochondrial Proteins; Proteins; RNA, Messenger; Tretinoin; Uncoupling Protein 1; Uncoupling Protein 2; Vitamin A Deficiency

Normal embryonic development and survival in utero is dependent on an adequate supply of vitamin A. Embryos from vitamin A-deficient (VAD) pregnant rats fed an inadequate amount of all-trans retinoic acid (atRA; 12 microg per g of diet or approximately 230 microg per rat per day) exhibit severe developmental abnormalities of the anterior cardinal vein and hindbrain by embryonic day (E) 12.5 and die shortly thereafter.. In the present study, we sought to determine whether supplementation of VAD-RA supported (12 microg per g of diet) pregnant rats with retinol (ROL) at the late-gastrula (presomite or rat E9.5) or early somite stages (E10.5), or provision of higher levels of atRA throughout this period could prevent abnormalities in the developing cardiovascular and nervous systems.. A newly described defect in the sinuatrial venus valve along with enlarged anterior cardinal veins and nervous system abnormalities and the later death of embryos are prevented by supplementing pregnant animals with ROL on the morning of E9.5. If ROL supplementation is delayed by 1 day (E10.5), most embryos are abnormal and die by E18.5. Supplementation of VAD rats with atRA (250 microg per g of diet) between E8.5 and E10.5 also prevents the cardiovascular and nervous system abnormalities and a significant number of these embryos survive to parturition. Thus, high levels of atRA can obviate the need for ROL between E9.5 and E10.5.. These results support an essential role for retinoid signaling between the late gastrula and early somite stages in the rat embryo for normal morphogenesis of the primitive heart tube and the posterior hindbrain. Further, these results suggest that embryonic death occurring at midgestation in the VAD rat may be linked to the abnormal development of one or both of these embryonic structures.

Topics: Abnormalities, Multiple; Animal Feed; Animals; Cranial Nerves; Diterpenes; Dose-Response Relationship, Drug; Embryonic and Fetal Development; Female; Fetal Death; Fetal Heart; Fetal Resorption; Gastrula; Genes, Homeobox; Gestational Age; Morphogenesis; Pregnancy; Pregnancy Complications; Rats; Retinyl Esters; Rhombencephalon; Transcription Factors; Tretinoin; Veins; Vitamin A; Vitamin A Deficiency

Vitamin A (VA) deficiency compromises antibody responses to T-cell-dependent antigens such as tetanus toxoid, but this effect can be reversed through administration of retinol or retinoic acid (RA). To test whether RA and polyriboinosinioc : polyribocytidylic acid (PIC), a known inducer of several forms of interferon (IFN), can cooperate to increase specific immunoglobulin (Ig)G and IgM production during VA deficiency, rats and mice were made VA-deficient, immunized with TT and treated with all-trans-RA, PIC or their combination. VA-deficient rats produced low primary and secondary anti-tetanus IgG responses (VA-deficient controls v. VA-sufficient controls P < 0.001), although total IgG was slightly elevated when compared with VA-sufficient control rats. Although RA administered alone elevated antibody production during VA deficiency to control levels, RA combined with PIC synergistically enhanced these responses (RA and PIC group v. all other groups P < 0.0001). In contrast, Balb/c mice maintained on a VA-deficient diet and immunized in a similar fashion showed no impairment in antigen-specific IgG levels, but treatment with a combination of RA and PIC still evoked an additive enhancement in antigen-specific antibody production. Additionally, RA and PIC administration to VA-sufficient mice resulted in elevated antibody responses, suggesting that this combination should be evaluated further for its immuno-stimulatory effects.

Topics: Animals; Drug Synergism; Female; Gene Expression Regulation; Immunoglobulin G; Immunoglobulin M; Lactation; Male; Mice; Mice, Inbred BALB C; Poly I-C; Rats; Rats, Inbred Lew; Tetanus Toxoid; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Humans; Receptors, Retinoic Acid; Skin; Sunlight; Tretinoin; Ultraviolet Rays; Vitamin A Deficiency

We report here that ultraviolet irradiation substantially reduced the mRNA and protein of the two major nuclear retinoid receptors, RAR-gamma and RXR-alpha, in human skin in vivo. Pre-treatment with retinoic acid mitigated this loss of nuclear retinoid receptors. Ultraviolet irradiation caused a near-total loss of retinoic acid induction of two RAR/RXR target genes, cellular retinoic acid binding protein-II and RA 4-hydroxylase, but did not affect 1,25-dihydroxyvitamin D3 induction of the vitamin D receptor/RXR-regulated gene vitamin D 24-hydroxylase. In effect, ultraviolet irradiation causes a functional vitamin A deficiency that may have deleterious effects on skin function, contributing to skin photo-aging and carcinogenesis.

Topics: Administration, Topical; Adult; Biopsy; Cell Nucleus; Cytochrome P-450 Enzyme System; Humans; Male; Middle Aged; Receptors, Calcitriol; Receptors, Retinoic Acid; Retinoic Acid 4-Hydroxylase; Retinoid X Receptors; RNA, Messenger; Skin; Steroid Hydroxylases; Transcription Factors; Tretinoin; Ultraviolet Rays; Vitamin A Deficiency; Vitamin D3 24-Hydroxylase

Topics: Adolescent; Esters; Female; Humans; Liver; Mutation; Retina; Retinol-Binding Proteins; Tretinoin; Vitamin A; Vitamin A Deficiency

Adh4, a member of the mouse alcohol dehydrogenase (ADH) gene family, encodes an enzyme that functions in vitro as a retinol dehydrogenase in the conversion of retinol to retinoic acid, an important developmental signaling molecule. To explore the role of Adh4 in retinoid signaling in vivo, gene targeting was used to create a null mutation at the Adh4 locus. Homozygous Adh4 mutant mice were viable and fertile and demonstrated no obvious defects when maintained on a standard mouse diet. However, when subjected to vitamin A deficiency during gestation, Adh4 mutant mice demonstrated a higher number of stillbirths than did wild-type mice. The proportion of liveborn second generation vitamin A-deficient newborn mice was only 15% for Adh4 mutant mice but 49% for wild-type mice. After retinol administration to vitamin A-deficient dams in order to rescue embryonic development, Adh4 mutant mice demonstrated a higher resorption rate at stage E12.5 (69%), compared with wild-type mice (30%). The relative ability of Adh4 mutant and wild-type mice to metabolize retinol to retinoic acid was measured after administration of a 100-mg/kg dose of retinol. Whereas kidney retinoic acid levels were below the level of detection in all vehicle-treated mice (< 1 pmol/g), retinol treatment resulted in very high kidney retinoic acid levels in wild-type mice (273 pmol/g) but 8-fold lower levels in Adh4 mutant mice (32 pmol/g), indicating a defect in metabolism of retinol to retinoic acid. These findings demonstrate that another retinol dehydrogenase can compensate for a lack of Adh4 when vitamin A is sufficient, but that Adh4 helps optimize retinol utilization under conditions of both retinol deficiency and excess.

Topics: Alcohol Dehydrogenase; Animals; Animals, Newborn; Female; Homozygote; Isoenzymes; Mice; Mice, Knockout; Pregnancy; Signal Transduction; Tretinoin; Vitamin A; Vitamin A Deficiency

Retinoids long have been implicated in limb development and their endogenous contributions to this process are finally being elucidated. Here we use an established model of retinoid depletion during specific gestational windows to investigate the role of endogenous retinoic acid (RA) in supporting limb outgrowth. Rat embryos were deprived of RA starting at days-postcoitum (dpc) 3.0, 5.5, or 7.0 and harvested at the 35-somite stage (dpc 12-12.5). Although embryos from all these windows possessed many characteristics of gestational retinoid deficiency (frontonasal hypoplasia, straight tail, reduced CRBPI and RAR beta), their limb buds emerged with only modest size reductions. Molecular analysis of RA-deficient limb buds revealed enhanced gli-3 and reduced hoxd-12, hoxd-13, shh, and fgf-4, while fgf-8, en-1, and wnt-7a expression remained unaltered. Occasional posterior truncations were observed at low incidence in the longest deficiency window; otherwise, the deficiency window length had no discernable impact on the severity of these changes. At the 45-somite stage, RA-deficient limbs had additional losses of hoxd-13 and fgf-8, accompanied by a flattened AER, suggestive of an ultimate failure in limb bud outgrowth. Results could not confirm a function for endogenous retinoids in limb initiation, but show they are required to maintain the signaling loops between the developing mesenchyme and AER that govern limb outgrowth after the initial emergence of limb bud.

Topics: Abnormalities, Multiple; Animals; Embryonic and Fetal Development; Embryonic Induction; Female; Fetal Resorption; Gene Expression Regulation, Developmental; Hedgehog Proteins; Homeodomain Proteins; Limb Buds; Pregnancy; Proteins; Rats; Rats, Sprague-Dawley; Signal Transduction; Trans-Activators; Transcription Factors; Tretinoin; Vitamin A Deficiency

Topics: Epithelium; Eye Diseases; Humans; Lung Diseases; Mouth Diseases; Tracheal Diseases; Tretinoin; Vitamin A; Vitamin A Deficiency

By way of differential screening of testis cDNA libraries from vitamin A-deficient (VAD) rats before and after administration of all-trans retinoic acid (ATRA), genes, the transcription of which was influenced by ATRA, were isolated. Most clones with an increased transcription encoded different subunits of the same mitochondrial protein complex, cytochrome c oxidase (COX). The mRNA expression of COX increased by a factor 3.9 +/- 1.5 (mean +/- SD, n = 4). This increased expression seems to reflect an increased energy demand in the ATRA-supplemented VAD testis. Also, one gene was isolated, the transcription of which was reduced to about 70% by ATRA. This gene, sulfated glycoprotein 2 (Sgp-2), is a major secretion product of Sertoli cells, the function of which is still unknown. The effect of ATRA on Sgp-2 expression may be direct, since the promoter of Sgp-2 contains a putative ATRA-responsive element (RARE).

Topics: Animals; Clusterin; DNA, Complementary; Electron Transport Complex IV; Female; Gene Expression Regulation; Genomic Library; Glycoproteins; Male; Molecular Chaperones; Pregnancy; Rats; Rats, Wistar; Spermatogenesis; Testis; Tretinoin; Vitamin A Deficiency

In vitro incubation of all-trans-retinol (atROL) with kidney homogenate from vitamin A-deficient and retinoic acid-supplemented (VAD-RAS) female rats produces a new retinol metabolite. Reverse-phase (RP) and normal-phase (NP) high-performance liquid chromatography (HPLC) analysis showed that this metabolite coelutes with the unknown all-trans-retinol (atROL) metabolite previously found in the day 10 conceptus and kidneys of vitamin A-deficient rats maintained on all-trans-retinoic acid (VAD-RA) and given 2 microg of [3H]atROL. Normal-phase (NP) HPLC purification of the metabolite collected from a RP HPLC column further separated the radiolabeled material into two components. The two isolated compounds have identical or very similar spectroscopic properties. Their nuclear magnetic resonance (1H NMR) and mass spectra (MS) indicated that they are isomers. Spectroscopic studies of the metabolites and their derivatives showed that they are nine-carbon fragments resulting from an oxidative cleavage of the side chain of atROL. The cleavage occurs at C-9, and the product is then oxidized to a keto group. The primary hydroxy group from atROL is preserved in the metabolite. A sulfide bridge is formed between C-11 and C-14, which interrupts the conjugation. The formation of the new metabolites, possessing a 2,5-dihydrothiophene ring, is catalyzed by an enzyme(s) located in the cytosolic fraction of kidneys. The process represents a new retinol metabolic pathway; however, its biological significance is unknown.

Topics: Administration, Oral; Animals; Chromatography, High Pressure Liquid; Female; Kidney; Magnetic Resonance Spectroscopy; Mass Spectrometry; Rats; Spectrophotometry, Ultraviolet; Spectroscopy, Fourier Transform Infrared; Subcellular Fractions; Tretinoin; Vitamin A; Vitamin A Deficiency

Retinal dehydrogenase (RALDH) catalyzes the oxidation of retinal to all-trans and 9-cis retinoic acid, which function as ligands controlling RAR and RXR nuclear receptor-signaling pathways. We have recently shown the expression of RALDH transcript in the stomach and small intestine by reverse transcription polymerase chain reaction [Bhat, P.V., Labrecque J., Dumas, F., Lacroix, A. and Yoshida, A. (1995) Gene 166, 303-306]. We have examined RALDH expression in the stomach and small intestine before and during postnatal development and in vitamin A deficiency by assaying for mRNA levels and protein as well as for enzyme activity. In -2 day fetuses, RALDH expression was high in the small intestine, whereas RALDH protein was not detectable in the stomach. However, expression of RALDH was seen in the stomach after birth, and gradually increased with age and reached the highest level at postnatal day 42. In the intestine, RALDH expression decreased postnatally. Vitamin A deficiency up-regulated RALDH expression in the stomach and small intestine, and administration of retinoids down-regulated the RALDH expression in these tissues. These results show the differential expression of RALDH in the stomach and small intestine during postnatal development, and that vitamin A status regulates the expression of RALDH gene in these tissues.

Topics: Age Factors; Aldehyde Oxidoreductases; Animals; Gene Expression Regulation, Enzymologic; Intestine, Small; Rats; Rats, Sprague-Dawley; Retinal Dehydrogenase; RNA, Messenger; Stomach; Tretinoin; Vitamin A; Vitamin A Deficiency

In keeping with the in vivo observation that the conversion of retinoyl beta-glucuronide (RBG) to retinoic acid (RA) is enhanced in vitamin A-deficient (A-) rats, the relative rates of hydrolysis of RBG to RA by various organelles of liver, kidney and intestine were found to be higher in A- rats than in vitamin A-sufficient (A+) rats (mean ratio 1.28; range 1.05-1.63). The lysosomal fraction of kidney and the microsomal fraction of liver showed the highest ratios for RBG hydrolysis; namely, 1.63 and 1.57, respectively (p < 0.005). The rates of hydrolysis of an ether glucuronide, p-nitrophenyl-beta-D-glucosiduronate (pNPG), were also enhanced in A- rats. The ratios of activities were again highest in kidney lysosomes and in liver microsomes; namely, 1.51 (p < 0.005) and 1.42 (p < 0.05), respectively. The non-ionic detergent, Triton X-100, increased RBG hydrolysis in organelles of A+ (19-27%) more than in those of A- rats (8-14%). The ratios of activities +/- SEM with 0.02% Triton X-100 in organelles of kidney, liver and intestine were 1.25 +/- 0.03, 1.22 +/- 0.03 and 1.24 +/- 0.03 in A+ rats and were 1.11 +/- 0.02, 1.07 +/- 0.02 and 1.13 +/- 0.03, respectively, in A- rats. Thus, Triton X-100 had a significantly greater effect (p < 0.005) on the membranes of A+ rats than on those of A- rats. In conclusion, the increased appearance of RA in the plasma after RBG administration in vivo seems to be caused by enhanced activity of beta-glucuronidases in tissue organelles, augmented in part by better access of the substrate to the membrane-bound enzymes.

Topics: Animals; Hydrolysis; Intestines; Kidney; Lysosomes; Male; Microsomes, Liver; Octoxynol; Organelles; Rats; Rats, Wistar; Tretinoin; Vitamin A Deficiency; Weaning

Retinoid X receptors (RXRs) are key regulators in retinoid signaling. Knowledge about the effects of 9-cis-retinoic acid (9-cis-RA), the natural ligand for the RXRs, may also provide insight in the functions of RXRs. In this study, the effect of 9-cis-RA on spermatogenesis in vitamin A-deficient (VAD) mice was examined. Administration of 9-cis-RA stimulated the differentiation and subsequent proliferation of the growth-arrested A spermatogonia in the testis of VAD mice. However, compared with all-trans-retinoic acid (ATRA), relatively higher doses of 9-cis-RA were necessary. This could not simply be due to a lower or delayed activity of 9-cis-RA, as simultaneous administration of ATRA and 9-cis-RA did not cause a synergistic effect. Instead, the presence of 9-cis-RA diminished the effect of ATRA by approximately one third. Studies of in vivo transport and metabolism showed that ATRA and 9-cis-RA, after administration to VAD mice, penetrated the testis equally well. However, 9-cis-RA was metabolized much faster than ATRA, and other metabolites were formed. This may account for the above-described differential effects of ATRA and 9-cis-RA on spermatogenesis. Similar to ATRA, 9-cis-RA transiently induced the messenger RNA expression of the nuclear RA receptor RAR beta, suggesting a role for this receptor in the effects of retinoids on the differentiation and proliferation of A spermatogonia. In contrast, the messenger RNA expression of the nuclear retinoid receptors RXR alpha, -beta, and -gamma was not changed significantly by administration of their ligand, 9-cis-RA. Hence, 9-cis-RA does not seem to exert its effect on spermatogenesis through altered expression of the RXRs.

Topics: Alitretinoin; Animals; Cell Differentiation; Cell Division; Male; Mice; Receptors, Retinoic Acid; Retinoid X Receptors; RNA, Messenger; Spermatogenesis; Spermatogonia; Testis; Transcription Factors; Tretinoin; Vitamin A Deficiency

Vitamin A is required for reproduction and normal embryonic development. We have determined that all-trans-retinoic acid (atRA) can support development of the mammalian embryo to parturition in vitamin A-deficient (VAD) rats. At embryonic day (E) 0.5, VAD dams were fed purified diets containing either 12 micrograms of atRA per g of diet (230 micrograms per rat per day) or 250 micrograms of atRA per g of diet (4.5 mg per rat per day) or were fed the purified diet supplemented with a source of retinol (100 units of retinyl palmitate per day). An additional group was fed both 250 micrograms of atRA per g of diet in combination with retinyl palmitate. Embryonic survival to E12.5 was similar for all groups. However, embryonic development in the group fed 12 micrograms of atRA per g of diet was grossly abnormal. The most notable defects were in the region of the hindbrain, which included a loss of posterior cranial nerves (IX, X, XI, and XII) and postotic pharyngeal arches as well as the presence of ectopic otic vesicles and a swollen anterior cardinal vein. All embryonic abnormalities at E12.5 were prevented by feeding pharmacological amounts of atRA (250 micrograms/g diet) or by supplementation with retinyl palmitate. Embryos from VAD dams receiving 12 micrograms of atRA per g of diet were resorbed by E18.5, whereas those in the group fed 250 micrograms of atRA per g of diet survived to parturition but died shortly thereafter. Equivalent results were obtained by using commercial grade atRA or atRA that had been purified to eliminate any potential contamination by neutral retinoids, such as retinol. Thus, 250 micrograms of atRA per g of diet fed to VAD dams (approximately 4.5 mg per rat per day) can prevent the death of embryos at midgestation and prevents the early embryonic abnormalities that arise when VAD dams are fed insufficient amounts of atRA.

Topics: Animals; Diet; Female; Fetal Resorption; Keratolytic Agents; Maternal-Fetal Exchange; Pregnancy; Rats; Rats, Sprague-Dawley; Rhombencephalon; Tretinoin; Vitamin A Deficiency

The cellular distribution of enzymes that esterify retinol and hydrolyze retinyl esters (RE) was studied in liver of vitamin A-sufficient, -deficient, and deficient rats treated with retinoic acid or N-(4-hydroxyphenyl)-retinamide. Livers were perfused and cell fractions enriched in hepatocytes, and nonparenchymal cells were obtained for assays of RE and enzyme activity. The specific activity of lecithin:retinol acyltransferase (LRAT) was approximately 10-fold greater in the nonparenchymal cell than the hepatocyte fraction from both vitamin A-sufficient and retinoid-treated rats. Total RE mass, newly synthesized [3H]RE and LRAT activity were positively correlated in liver and isolated cells of both normal (P < 0.0001) and retinoid-treated rats (P < 0.0002). In nonparenchymal cells, these three constituents were nearly equally enriched as evaluated by their relative specific activity values (RSA, defined as the percentage of recovered activity divided by the percentage of recovered protein), which were each significantly greater than 1.0, with values of 4.3 for total RE mass (P < 0.05), 3.6 for newly synthesized [3H]RE (P < 0.01) and 3.8 for LRAT activity (P < 0.01). In contrast, the specific activities of neutral and acid bile salt-independent retinyl ester hydrolases (REH) did not vary with vitamin A status, and their RSA values were close to 1.0 in both hepatocytes and nonparenchymal cells. These data show that LRAT and REH are differentially regulated by retinoids and that these enzymes also differ in their spacial distribution between liver parenchymal and nonparenchymal cells.

Topics: Acyltransferases; Animals; Carboxylic Ester Hydrolases; Female; Fenretinide; Liver; Male; Rats; Rats, Inbred Lew; Tretinoin; Vitamin A; Vitamin A Deficiency

The tissue concentration of retinol and its metabolites was determined in a group of vitamin A deficient rats after a single dose of 53 micrograms of [11,12-3H]retinol. The sensitive technique of high performance liquid chromatography was used to analyse the metabolites of retinol. The analysis of the metabolites in tissues at different days after the administration of radioactive retinol showed a rapid decrease in the amount of retinol and retinyl esters in the liver tissue, accompanied by an increase in the retinol and retinyl ester values in the kidney. In addition, the content of retinoic acid was higher in liver and kidney compared with intestine, testis, and blood. It reached maximum at 4 and 11 days, respectively. After 17 days the retinoid(s) concentrations decreased markedly in all tissues studied; yet the kidney showed higher concentrations of retinoic acid and retinyl esters. These studies indicate that the kidney retains more vitamin A as vitamin A becomes depleted in the body, probably as a reserve for the production of the active metabolite retinoic acid, needed for the growth and differentiation.

Topics: Animals; Body Weight; Esters; Male; Rats; Rats, Sprague-Dawley; Tissue Distribution; Tretinoin; Tritium; Vitamin A; Vitamin A Deficiency

Changes in ultrastructural characteristics and mucin gene expression were examined in rat tracheal explants cultured in a synthetic medium +/- retinoic acid (RA), benzo[a]pyrene (B[a]P) and N-methyl-N-nitrosourea (NMNU). In the RA(+) cultures, no changes in either ultrastructural features or mucin gene expression were detected after 48 h incubation. After 96 h incubation, however, the ultrastructural features associated with the squamous phenotype were characteristics of cultures containing the two carcinogens and the mucin gene expression was slightly reduced. Thus, in the presence of retinoic acid, the carcinogen induced changes in cytology to the squamous phenotypes were not matched by a marked loss of mucin gene expression. Explants cultured for 48 h without RA and +/- carcinogens showed none of the cytological changes associated with onset of the squamous phenotype. While mucin mRNA was still detected, it was clearly reduced compared to 48 h cultures in RA(+) medium. However, 48 h later, all explants exhibited pronounced squamous metaplasia and the mucin message decreased to trace levels. Thus, the results of these experiments with B[a]P and NMNU in RA(+) and RA(-) media indicates that at least the early carcinogen induced changes may be distinct from those associated with the retinoid pathway controlling expression of the mucin component of the mucociliary epithelium.

Topics: Animals; Benzo(a)pyrene; Carcinogens; Carcinoma, Squamous Cell; Cell Differentiation; Culture Media; Culture Techniques; Epithelium; Gene Expression; Lung Neoplasms; Metaplasia; Methylnitrosourea; Mucins; Phenotype; Rats; RNA, Messenger; Trachea; Tretinoin; Vitamin A Deficiency

We examined the effects of vitamin A deficiency and all-trans retinoic acid (RA) supplementation on regulation of three important genes in hepatic gluconeogenesis: the genes for phosphoenolpyruvate carboxykinase (PEPCK), fructose-1,6-bisphosphatase (Fru-1,6-P2ase) and 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (6-PF-2-K/Fru-2,6-P2ase). Mice were made vitamin A deficient in the second generation by initiating a vitamin A-deficient diet on d 10 of gestation. At 7 wk of age, vitamin A-deficient mice were treated with all-trans RA or vehicle alone and killed for RNA analysis. In liver, vitamin A deficiency resulted in PEPCK mRNA levels that were 74% lower and 6-PF-2-K/Fru-2,6-P2ase mRNA levels that were 42% lower than the respective mRNA measured in control mice. The Fru-1,6-P2ase mRNA abundance was not affected by vitamin A deficiency. The decrease in hepatic PEPCK and 6-PF-2-K/Fru-2,6-P2ase mRNA levels was reversed by treatment with all-trans RA within 3 h of administration. In mice fed the control diet, food deprivation for 15 h resulted in PEPCK mRNA levels that were 3.5-fold higher, Fru-1,6-P2ase mRNA levels that were 2-fold higher, and 6-PF-2-K/Fru-2,6-P2ase mRNA levels that were 3.4-fold higher than in fed mice. Vitamin A-deficient mice did not respond to food deprivation with induced PEPCK mRNA levels, whereas 6-PF-2-K/Fru-2,6-P2ase and Fru-1,6-P2ase mRNA levels were induced. The pattern of 6-PF-2-K/Fru-2,6-P2ase mRNA abundance with vitamin A deficiency and food deprivation was complex and different from that for either PEPCK or Fru-1,6-P2ase transcripts. The cAMP-responsiveness of the PEPCK gene in vitamin A-deficient mice was tested. Vitamin A deficiency caused a significant reduction in cAMP stimulation of PEPCK mRNA levels in liver. These results in the whole animal indicate that vitamin A regulation of the hepatic PEPCK gene is physiologically important; without adequate vitamin A nutriture, stimulation of the PEPCK gene by food deprivation or cAMP treatment is inhibited in the liver.

Topics: Animals; Bucladesine; Food Deprivation; Fructose-Bisphosphatase; Gene Expression Regulation, Enzymologic; Gluconeogenesis; Liver; Mice; Mice, Inbred C57BL; Phosphoenolpyruvate Carboxykinase (GTP); Phosphofructokinase-1; Promoter Regions, Genetic; RNA, Messenger; Tretinoin; Vitamin A; Vitamin A Deficiency

Rats deficient in vitamin A express low levels of P4502C7 mRNA in the liver. Administration of all-trans retinoic acid (at-RA) or growth hormone (GH) to deficient animals only partially restored the expression whereas the combined treatment returned the P4502C7 mRNA levels to that observed in normal rats. That a retinoid is the predominant inducer of P4502C7 at the cellular level is evident from studies performed with primary hepatocytes, but it became clear that GH is a prerequisite for the vitamin A effect in vivo. The at-RA induction of P4502C7 mRNA in primary rat hepatocytes was inhibited by ketoconazole, an inhibitor of P450 activity, and by cycloheximide, blocking ongoing protein synthesis. In contrast, the at-RA induction of RAR-beta2 mRNA was not affected by any of these compounds. This could indicate previously not recognized mechanisms of at-RA action. Interestingly, at-4-oxo-RA, an at-RA metabolite formed by a P450 catalyzed reaction, also induced P4502C7 mRNA. Induction of P4502C7 mRNA by the retinoic acid receptor (RAR) selective agonist TTNPB indicated that this pathway is preferred over the retinoid X receptor (RXR) pathway. In addition, analysis of RA metabolites in liver cell extracts revealed the formation of several as yet unidentified metabolites. The formation of some of these metabolites was inhibited by ketoconazole and they could therefore constitute potential inducers of CYP2C7. We suggest that metabolism of at-RA, possibly by a P450 enzyme, is an important step in the at-RA induction of P4502C7.

Topics: Animals; Cytochrome P-450 Enzyme System; Female; Gene Expression Regulation; Growth Hormone; Liver; Rats; Rats, Sprague-Dawley; RNA, Messenger; Tretinoin; Vitamin A Deficiency

The microsomal enzyme LRAT esterifies retinol and has been implicated in the hepatic storage of vitamin A. Previously, we showed that hepatic LRAT activity is negligible during vitamin A deficiency and that all-trans-retinoic acid (all-trans-RA) rapidly induces the activity of liver LRAT in retinoid-deficient rats. In the present studies, we have examined the ability of natural and synthetic retinoids to induce liver LRAT activity in retinoid-deficient rats. The natural retinoids retinol, all-trans-RA (100 microg), 9-cis-RA, or equal molar amounts of other retinoids were injected ip and LRAT specific activity was measured in liver homogenates 17-18 h later. In retinoid-deficient rats, liver LRAT activity was extremely low [0.13 +/- 0.03 pmol retinyl ester (RE)/min/mg liver protein, mean +/- SE]. The natural retinoids retinol and all-trans-RA strongly induced LRAT activity (12.71 +/- 1.09 and 13.10 +/- 1.55 pmol RE/min/mg, respectively), whereas 9-cis-RA induced a lower level of LRAT activity (3.96 +/- 1.88 pmol RE/min/mg, P < 0.001 vs all-trans-RA). The retinoic acid receptor (RAR)-selective analog (RAR pan-agonist) all-trans-UAB8 and the RAR-alpha-selective retinoid Am580 also strongly induced LRAT activity. In contrast, neither RXR-selective agonists nor retinoids having a retro structure were active. For retinoids with significant RAR-alpha binding activity there was a strong direct correlation between receptor binding in vitro and the ability to induce hepatic LRAT activity in vivo (r2 = 0.920). These data implicate the RARs in the induction of hepatic LRAT and suggest a predominant role for RAR-alpha-active ligands.

Topics: Acyltransferases; Animals; Female; Microsomes, Liver; Molecular Structure; Protein Binding; Rats; Rats, Inbred Strains; Receptors, Retinoic Acid; Retinoid X Receptors; Retinoids; Transcription Factors; Tretinoin; Vitamin A; Vitamin A Deficiency

Both retinoid receptor null mutants and classic nutritional deficiency studies have demonstrated that retinoids are essential for the normal development of diverse embryonic structures (e.g. eye, heart, nervous system, urogenital tract). Detailed analysis of retinoid-modulated events is hampered by several limitations of these models, including that deficiency or null mutation is present throughout gestation, making it difficult to isolate primary effects, and preventing analysis beyond embryolethality. We developed a mammalian model in which retinoid-dependent events are documented during distinct targeted windows of embryogenesis. This was accomplished through the production of vitamin A-depleted (VAD) female rats maintained on sufficient oral retinoic acid (RA) for growth and fertility. After mating to normal males, these RA-sufficient/VAD females were given oral RA doses which allowed for gestation in an RA-sufficient state; embryogenesis proceeded normally until retinoids were withdrawn dietarily to produce a sudden, acute retinoid deficiency during a selected gestational window. In this trial, final RA doses were administered on E11.5, vehicle at E12.5, and embryos analyzed on E13.5; during this 48 hour window, the last RA dose was metabolized and embryos progressed in a retinoid-deficient state. RA-sufficient embryos were normal. Retinoid-depleted embryos exhibited specific malformations of the face, neural crest, eyes, heart, and nervous system. Some defects were phenocopies of those seen in null mutant mice for RXR alpha(-/-), RXR alpha(-/-)/RAR alpha(-/-), and RAR alpha(-/-)/RAR gamma(-/-), confirming that RA transactivation of its nuclear receptors is essential for normal embryogenesis. Other defects were unique to this deficiency model, showing that complete ligand 'knock-out' is required to see those retinoid-dependent events previously concealed by receptor functional redundancy, and reinforcing that retinoid receptors have separate yet overlapping contributions in the embryo. This model allows for precise targeting of retinoid form and deficiency to specific developmental windows, and will facilitate studies of distinct temporal events.

Topics: Abnormalities, Multiple; Animals; Diet; Embryo Implantation; Embryonic and Fetal Development; Eye Abnormalities; Female; Liver; Nervous System; Nervous System Malformations; Neural Crest; Pregnancy; Rats; Rats, Mutant Strains; Rats, Sprague-Dawley; Receptors, Retinoic Acid; Retinoids; Tretinoin; Vitamin A Deficiency

The effect of ethanol on early avian cardiovascular development was investigated in stage 8 quail embryos grown in culture for 24 hr. When the culture medium contained 1% ethanol, 50% of the embryos developed abnormalities of the cardiovascular system, some of which resembled vitamin A deficiency. Only 15% of the embryos grown in control media developed abnormalities attributed to the manipulation of the embryo. When all-trans-retinoic acid, the active form of vitamin A, was added at 10(-8) M to the ethanol-containing medium, the cardiovascular development was similar to that of untreated controls. Inclusion of 4-methylpyrazole and citral, enzyme inhibitors for the conversion of retinol to retinoic acid, produced cardiovascular abnormalities in embryos similar to those observed in vitamin A deficiency. These abnormalities were partially prevented by the presence of 10(-8) M all-trans-retinoic acid in the medium. Immunohistochemical studies using antibodies specific for the heart muscle myosin heavy chain (MF-20) and quail endothelial cells (QH-1) revealed that looping of the heart of ethanol-treated embryos was prevented, and the embryonal circulation had no or minimal vascular connections to the extraembryonic circulation. Our studies provide indirect evidence that ethanol is producing vitamin A deficiency during embryonic cardiovascular development and that these effects are specifically prevented by the presence of retinoic acid. These findings may explain some of the symptoms of fetal alcohol syndrome.

Topics: Animals; Dose-Response Relationship, Drug; Embryo, Nonmammalian; Ethanol; Fetal Alcohol Spectrum Disorders; Heart Defects, Congenital; Quail; Tretinoin; Vitamin A Deficiency

Topics: Alitretinoin; Antineoplastic Agents; Carcinoma, Small Cell; Gene Deletion; Humans; Loss of Heterozygosity; Lung Neoplasms; Mutation; Neoplasms, Second Primary; Smoking; Survival Analysis; Tretinoin; Vitamin A Deficiency

We have studied the effects of retinoic acid (RA) and thyroid hormone (3,3',5-triiodothyronine; T3) on platelet-activating factor receptor (PAFR) gene expression in intact rats and the ability of two human PAFR gene promoters (PAFR promoters 1 and 2) to generate two transcripts (PAFR transcripts 1 and 2). Northern blotting showed that RA and T3 regulated PAFR gene expression only in rat tissues that express PAFR transcript 2. Functional analysis of the human PAFR promoter 2 revealed that responsiveness to RA and T3 was conferred through a 24-bp element [PAFR-hormone response element (HRE) located from -67 to -44 bp of the transcription start site, whereas PAFR promoter 1 did not respond to these hormones. The PAFR-HRE is composed of three direct repeated TGACCT-like hexamer motifs with 2-and 4-bp spaces, and the two upstream and two downstream motifs were identified as response elements for RA and T3. Thus, the PAF-PAFR pathway is regulated by the PAFR level altered by a tissue-specific response to RA and T3 through the PAFR-HRE of the PAFR promoter 2.

Topics: Animals; Base Sequence; DNA-Binding Proteins; Gene Expression Regulation; Humans; Molecular Sequence Data; Myocardium; Organ Specificity; Platelet Membrane Glycoproteins; Promoter Regions, Genetic; Protein Binding; Rats; Rats, Wistar; Receptors, Cell Surface; Receptors, G-Protein-Coupled; Receptors, Retinoic Acid; Receptors, Thyroid Hormone; Repetitive Sequences, Nucleic Acid; Skin; Tretinoin; Triiodothyronine; Vitamin A Deficiency

Both Hensen's node, the organizer center in chick embryo, and exogenous retinoic acid are known to induce limb duplication when grafted or applied to the host chick limb bud. Retinoic acid is known to be present in the node and has been proposed as the putative morphogen for chick limb development. Here, we report that Hensen's node from vitamin A-deficient quail embryo induces limb duplication in the host chick embryo similar to that induced by the node from vitamin A-sufficient control embryos. We also demonstrate that the expression of Sonic hedgehog (Shh), recently shown to be the mediator of polarizing activity in the chick limb bud, is not affected by the endogenous vitamin A status of the embryo. Furthermore, whole-mount in situ hybridization revealed asymmetry of Shh expression in the Hensen's node of both vitamin A-sufficient and -deficient quail embryos. Retinoids were not detectable in the eggs from which vitamin A-deficient embryos were obtained. Extracts from normal embryos induced a level of expression of reporter gene equivalent to the presence of 3.4 pg of active retinoids per embryo, while those from vitamin A-deficient embryos induced a baseline level of reporter gene expression similar to that of the controls. Our studies suggest that endogenous retinoic acid is not involved in Shh expression nor in regulating its asymmetry during normal early avian embryogenesis and support the current view that endogenous retinoic acid may not be a direct morphogen for limb bud duplication.

Topics: Animals; Biological Assay; Chick Embryo; Chromatography, High Pressure Liquid; Coturnix; Embryonic Induction; Extremities; Genes, Reporter; Hedgehog Proteins; In Situ Hybridization; Ovum; Proteins; Retinoids; Tissue Transplantation; Trans-Activators; Tretinoin; Vitamin A Deficiency

Vitamin A deficiency leads to an arrest of spermatogenesis and a loss of advanced germ cells in male mice. In the present study, the effects of several retinoids and carotenoids on these mouse testis were investigated. First, the proliferative activity of the growth-arrested A spermatogonia in vitamin A-deficient (VAD) mice testis was determined, 20, 24, or 28 h after administration of 0.5 mg all-trans-retinoic acid (RA). The bromodeoxy-uridine (BrdU) labeling index of A spermatogonia in control VAD testis was 5 +/- 1% (n = 4, mean +/- SD). When RA was injected (ip), the highest labeling index was found 24 h after RA administration; 49 +/- 5%. When various concentrations of RA, all-trans-4-oxo-retinoic acid (4-oxo-RA) or all-trans-retinol acetate (ROAc), ranging from 0.13-1 mg, were injected, the labeling index of A spermatogonia always increased in comparison with the VAD situation. A maximum index at 24 h was found when 0.5 mg 4-oxo-RA was injected: 56 +/- 3%. This labeling index was even higher than those after injection of RA or ROAc, 49 +/- 5% and 34 +/- 6% respectively. The increase of the BrdU labeling index was dose dependent. After an initial increase of the labeling indices with increasing retinoid doses, the labeling indices decreased at a higher concentration. This decrease is likely due to a concentration dependent timeshift of the optimum of BrdU labeling to shorter time intervals after retinoid administration because a labeling index of 66 +/- 1% was found 20 h after injection of 1 mg RA. At 24 h, this labeling index was halved: 33 +/- 2%. These indices show that the degree of synchronization of spermatogenesis is also dependent on the retinoid dose. When the dimers of RA and 4-oxo-RA, respectively beta-carotene (beta C) and canthaxanthin, were given, 24 h after administration BrdU-labeling indices comparable with the VAD value were found. Repeated injection of beta C twice a week did induce a reinitiation of spermatogenesis, but compared with RA, the activity of beta C was lower and delayed. It is concluded that 4-oxo-RA is active in adult mammals in vivo. It is at least as potent as RA in the induction of the differentiation and subsequent proliferation of growth-arrested A spermatogonia in VAD mice testis. Furthermore, the degree of synchronization of spermatogenesis is influenced by the retinoid dose. Finally, carotenoids were shown to act in the induction of spermatogonial cell proliferation too but with a lower and delayed activity.

Topics: Animals; beta Carotene; Bromodeoxyuridine; Carotenoids; Cell Differentiation; Cell Division; Male; Mice; Retinoids; Spermatogonia; Testis; Tretinoin; Vitamin A Deficiency

During fetal development, a specialized vessel the ductus arteriosus, shunts blood from the pulmonary artery to the aorta, thus bypassing the lungs. The ductus differs primarily from the great vessels in that it is a muscular rather than an elastic artery, and the etiology of this differential development remains controversial. We present evidence that retinoic acid (RA) may contribute to the unique muscle phenotype of the ductus arteriosus. Using a transgenic mouse carrying an RA response element-lacZ transgene that expresses beta-galactosidase (beta-gal) in response to endogenous RA signals during embryonic and fetal development, we observe a strong beta-gal signal in the ductus arteriosus. By immunofluorescence, this signal colocalizes with the expression of the adult-specific smooth muscle myosin heavy chain isoform, SM2. The beta-gal signal is present throughout fetal development and persists in the neonate until the ductus arteriosus is completely closed. beta-Gal-positive cells are first detected by immunofluorescence at 13.5 days postcoitum (dpc) in the mesenchyme surrounding the ductus. By 15.5 dpc, very intense beta-gal staining localizes to the ductus arteriosus but is absent or minimal in the pulmonary trunk and aortic arch; by 17.5 dpc, the smooth muscle layers of the tunica media in the ductus arteriosus exhibit positive beta-gal staining. Immunostaining with antibodies against smooth muscle myosins shows that, while SM1 is expressed in all embryonic vessels, SM2 is precociously expressed in the ductus arteriosus. Furthermore, SM2 expression can be detected in the ductus as early as 15.5 dpc. In the neonate, the beta-gal signal persists in the smooth muscle layer of the ductus and immunostaining colocalizes with SM2 expression. These data suggest that RA may play a role in inducing and maintaining smooth muscle differentiation in the developing ductus arteriosus and may promote precocious expression of the adult vascular phenotype.

Topics: Animals; Base Sequence; Ductus Arteriosus; Embryonic and Fetal Development; Female; Humans; Immunohistochemistry; Infant, Newborn; Isomerism; Mice; Mice, Transgenic; Molecular Probes; Molecular Sequence Data; Muscle, Smooth, Vascular; Myosin Heavy Chains; Phenotype; Pregnancy; Signal Transduction; Tretinoin; Vitamin A; Vitamin A Deficiency

All-trans-retinoyl beta-glucuronide (RAG) was chemically synthesized in high yields (up to 79%) by a new procedure involving the reaction of the tetrabutylammonium salt of glucuronic acid with all-trans-retinoic acid (RA) via the imidazole or triazole derivative. When RAG was fed orally to vitamin A-deficient rats, RA was identified as the major metabolite in the serum within hours of administration of RAG. Very little or no RAG was detected in the serum. Thus RAG, which was not appreciably hydrolysed to RA in vitamin A-sufficient rats [Barua and Olson (1987) Biochem. J. 263, 403-409], was rapidly converted into RA in vitamin A-deficient rats.

Topics: Acitretin; Animals; Chromatography, High Pressure Liquid; Glucuronates; Molecular Structure; Quaternary Ammonium Compounds; Rats; Rats, Sprague-Dawley; Tretinoin; Vitamin A Deficiency

It has been previously demonstrated that vitamin A-deficient rats supplemented with retinoic acid cannot complete gestation. Resorption of fetuses invariably occurs in these animals beginning Day 15 of gestation. Retinol must be administered on or before Day 10 of gestation in order to prevent this phenomenon. The administration of as little as 2 micrograms on Day 10 is sufficient to allow continuation of gestation through parturition. This study examines the metabolites of all-trans-retinol in the conceptuses of vitamin A-deficient, retinoic acid-supported pregnant animals at Day 10 of gestation. The retinoids recovered in this study could be identified as all-trans-retinol, retinyl palmitate, and other retinyl esters. Virtually no radiolabeled retinoic acid was found in the conceptuses of these animals over the 12-h period examined. This may suggest that retinoic acid that may be required by the fetus can arise directly from maternal sources. Additionally, the isolation of an early appearing, very polar metabolite of retinol is reported. This aqueous-soluble compound accounts for more than 20% of the radioactivity recovered at 1 h post-dose. The amount of this compound increases through 6 h post-dose. This metabolite is not found in urine from the same animals and is not likely to be an excretion product.

Topics: Animals; Female; Fetus; Gestational Age; Maternal-Fetal Exchange; Pregnancy; Pregnancy Complications; Rats; Rats, Sprague-Dawley; Stereoisomerism; Tretinoin; Vitamin A; Vitamin A Deficiency

Retinoic acid (RA) is a morphogenetically active signalling molecule thought to be involved in the development of severely embryonic systems (based on its effect when applied in excess and the fact that it can be detected endogenously in embryos). Here, we adopt a novel approach and use the vitamin A-deficient (A-) quail embryo to ask what defects these embryos show when they develop in the absence of RA, with particular reference to the nervous system.. We have examined the anatomy, the expression domains of a variety of genes and the immunoreactivity to several antibodies in these A- embryos. In addition to the previously documented cardiovascular abnormalities, we find that the somites are smaller in A- embryos, otic vesicle development is abnormal and the somites continue up to and underneath the otic vesicle. In the central nervous system, we find that neural crest cells need RA for normal development and survival, and the neural tube fails to extend any neurites into the periphery. Using general hindbrain morphology and the expression patterns of Hoxa-2, Hoxb-1, Hoxb-4, Krox-20 and FGF-3 as markers, we conclude that segmentation in the myelencephalon (rhombomeres 4-8) is disrupted. In contrast, the dorsoventral axis of the neural tube using Shh, islet-1 and Pax-3 as markers is normal.. These results demonstrate at least three roles for RA in central nervous system development: neural crest survival, neurite outgrowth and hindbrain patterning.

Topics: Animals; Coturnix; Medulla Oblongata; Neural Crest; Neural Tube Defects; Rhombencephalon; Tretinoin; Vitamin A Deficiency

Down-regulation of microsomal androgen-dependent CYP2C11 is produced in male rat liver by dietary vitamin A deficiency. Decreased circulating androgen concentrations also occur in vitamin A-deficient male rats. Both effects are prevented by addition of all-trans-retinoic acid to the diet. The present study evaluated directly whether androgen deficiency may be responsible for the down-regulation of 2C11 in vitamin A-deficient male rats. The major finding was that subcutaneous administration of the androgen methyltrienolone (MT) during the final week of the study restored CYP2C11 protein and its associated steroid 16alpha-hydroxylation activities to control levels; CYP2C11 mRNA was also restored. Despite the efficient restoration of CYP2C11 at a pretranslational level, no alteration in vitamin A status was apparent and animals remained vitamin A deficient after MT treatment. The possibility was assessed that vitamin A can maintain the microsomal content of CYP2C11 in normal liver. However, in contrast to MT, administration of ATRA to gonadectomized male rats did not restore 2C11 in liver. These findings establish that the major effect of vitamin A deficiency on CYP2C11 in male rat liver is mediated indirectly by androgen deficiency.

Topics: Androstenedione; Animals; Aryl Hydrocarbon Hydroxylases; Cytochrome P-450 Enzyme System; Cytochrome P450 Family 2; Diet; Down-Regulation; Hydroxylation; Liver; Male; Metribolone; Microsomes, Liver; Nutritional Status; Orchiectomy; Rats; Rats, Wistar; RNA, Messenger; Steroid 16-alpha-Hydroxylase; Steroid Hydroxylases; Testosterone Congeners; Tretinoin; Vitamin A Deficiency

Retinoic acid (RA), through its cognate receptors (retinoic acid receptors, RARs), plays an important role in the ontogenesis and maintenance of the normal function of murine Harderian and submandibular glands. In the present study, autoradiography was used to study RA binding to these glands. Both glands showed high radioactive labelling after [14C]-RA administration in normal and partially vitamin A-deficient (VAD) mice. The peak uptake was at 6 h after [14C]-RA administration in normal mice and at 0.5 h in VAD mice. At 24 h, RA binding remained high in normal mice, while it decreased significantly in VAD mice. In western blots with an antibody recognizing all forms of RARs, a band of molecular weight 51 kDa was seen in homogenates of both glands. Immunohistochemically, RAR staining was found in the nuclei of the glandular cells. The Harderian gland exhibited more intense staining than the submandibular gland. In the latter, the most intense staining was seen in the acinar cells, followed by the intercalated duct cells. The granular convoluted tubule showed weak immunostaining and the striated duct was negative. In the Harderian gland, RAR immunostaining was observed in both type I and II cells, but only part of them stained with RAR antibody. The expression of RAR alpha, beta, and gamma transcripts was studied by in situ hybridization using specific oligonucleotide probes. The cell-specific expression of RAR alpha mRNA in the submandibular gland corresponded to the RAR proteins detected by immunohistochemistry, while the RAR beta transcript was mainly seen in the striated duct. The transcripts of RAR alpha and beta were evenly distributed in type I and II glandular cells of the Harderian gland. RAR gamma labelling was below detectable levels in both glands. This result suggests that RA and RARs regulate the functions of Harderian and submandibular glands in a cell-specific manner.

Topics: Animals; Antibodies; Autoradiography; Carbon Radioisotopes; Harderian Gland; Immunoblotting; Immunohistochemistry; In Situ Hybridization; Male; Mice; Mice, Inbred Strains; Oligonucleotide Probes; Rats; Rats, Wistar; Receptors, Retinoic Acid; Reference Values; Retinoic Acid Receptor alpha; Retinoic Acid Receptor gamma; RNA, Messenger; Submandibular Gland; Transcription, Genetic; Tretinoin; Vitamin A Deficiency

In this study, we analyze the regulation of the squamous-specific gene, cornifin alpha, by estrogen and retinoic acid in vaginal and uterine epithelial cells. In ovariectomized animals, the vaginal epithelium consists of a stratified, nonkeratinizing epithelium which changes into a highly-stratified, keratinizing epithelium upon treatment with estradiol. This transition is accompanied by a dramatic induction of the crosslinked envelope precursor, cornifin alpha. An increase in cornifin mRNA can be detected as early as 3 h after treatment. A similar effect is observed for the synthetic estrogenic agent diethylstilbestrol while other steroid hormones, including testosterone, progesterone or dexamethasone have little effect on cornifin expression. In contrast to the vagina, estradiol induces neither squamous differentiation nor expression of cornifin alpha in the uterine epithelium. Similar to the action of estradiol, vitamin A-deficiency greatly enhances squamous differentiation and keratinization in the vaginal epithelium. But unlike estradiol, it induces squamous metaplasia in the normally columnar, uterine epithelium, which eventually is replaced by a keratinizing epithelium in severe deficiency. This transition is associated with an induction of cornifin alpha expression. Immunohistochemical and in situ hybridization analysis localizes cornifin protein and mRNA in the suprabasal layers of the squamous epithelium. Our results demonstrate that estrogen and retinoids play key roles in the regulation of differentiation and cornifin alpha expression in the uterine and vaginal epithelium.

Topics: Animals; Dexamethasone; Diethylstilbestrol; Dihydrotestosterone; Epithelial Cells; Epithelium; Estradiol; Female; Membrane Proteins; Ovariectomy; Progesterone; Rats; Rats, Sprague-Dawley; RNA, Messenger; Transcription, Genetic; Tretinoin; Uterus; Vagina; Vitamin A Deficiency

Topics: Antineoplastic Agents; Carcinoma, Hepatocellular; Humans; Liver Neoplasms; Neoplasms, Second Primary; Tretinoin; Vitamin A; Vitamin A Deficiency

In a previous study we investigated the effects of RA excess on TGF beta protein localization in early postimplantation stages of mouse development. Here we extend this investigation by comparing the effects of retinoid deficiency with those of excess, and by comparing the effects of altered retinoid status on TGF beta protein and RNA transcript distribution. In vitamin A-deficient embryos, TGF beta 1 RNA and protein distribution were both unaltered compared with controls; conversely, TGF beta 2 protein levels were reduced while RNA levels remained normal. In RA-treated embryos, the previous study showed that intracellular TGF beta 1 levels were decreased, while those of extracellular TFG beta 1 were initially decreased but subsequently increased; here we found that TGF beta 1 RNA transcript levels were reduced following exposure to RA excess. TGF beta 2 showed a clear disparity between the effects of RA excess on protein and RNA transcript levels: RNA transcript distribution was unchanged or showed a slight increase in RA-treated embryos, whereas the previous results showed greatly reduced protein levels. The new results provide further evidence for interaction between retinoids and TGF beta s during mouse development, and indicate that retinoids are capable of differentially regulating TGF beta isoforms through mechanisms involving different stages in the process of TGF beta synthesis and secretion. The long-term nature of the effects of transient exposure to RA excess suggests that the mechanisms of RA-TGF beta interaction may be indirect.

Topics: Animals; Cell Differentiation; Cell Division; Embryo, Mammalian; Embryonic and Fetal Development; Female; Gene Expression Regulation, Developmental; Immunohistochemistry; In Situ Hybridization; Mice; Mice, Inbred C57BL; Peptides; Pregnancy; Protein Biosynthesis; RNA, Messenger; Transcription, Genetic; Transforming Growth Factor beta; Tretinoin; Vitamin A Deficiency

Certain infections, like that with the human immunodeficiency virus-1, deplete vitamin A, and when vitamin A levels are low, immune dysfunctions establish susceptibility to further infection. Our research has focused on the immune dysfunctions that are a consequence of vitamin A deficiency and that predispose to further infection. We previously studied a helminth infection in mice, and showed that when vitamin A levels are low, the immune response develops a strong regulatory T cell imbalance with excessive T helper type-1 cell interferon (IFN)-gamma synthesis and insufficient T helper type-2 cell development and function. Here, we studied the T cell priming environment in vitamin A-deficient mice to learn how that priming environment might produce a regulatory T cell imbalance and consequently distort the ability of the immune system to respond to an infection. Our results show that during vitamin A deficiency, the priming environment included constitutive interleukin (IL)-12 and IFN-gamma transcripts, but it was devoid of constitutive IL-4 and IL-10 transcripts. Dietary all-trans-retinoic acid supplementation down-regulated the level of constitutive IL-12 and IFN-gamma transcripts. Furthermore, when T cells from naive vitamin A-deficient animals were stimulated through the T cell receptor, they produced excess IFN-gamma protein compared to T cells from control animals. In contrast, T cell stimulation failed to induce IL-4 or IL-10 secretion. The inducible IFN-gamma was largely from CD8+ T cells and all-trans-retinoic acid addition in vitro inhibited IFN-gamma production at the transcript level. Retinoic acid addition in vitro also decreased natural killer cell IFN-gamma synthesis at the transcript level. Taken together, the distorted constitutive and inducible cytokine gene expression patterns that occurred when vitamin A levels were low would be expected strongly to favor T helper type-1 development and limit T helper type-2 cell growth and differentiation, thereby limiting the animal's humoral immune response capability.

Topics: Animals; Cell Differentiation; Cells, Cultured; Diet; Down-Regulation; Interferon-gamma; Interleukin-12; Mice; T-Lymphocytes, Helper-Inducer; Transcription, Genetic; Tretinoin; Vitamin A Deficiency

Previously we reported that vitamin A-deficient rats have a low number of natural killer (NK) cells in their blood and spleen. The current studies were designed to address whether other cells of the immune system are also affected and whether dietary retinoic acid is able to reverse the changes caused by a deficiency of retinol and its metabolites. Total white blood cells, differential counts and spleen cell numbers were compared in vitamin A-sufficient rats (controls) and rats deficient in vitamin A, and lymphocyte and NK cell populations were identified and enumerated by flow cytometry. In comparison with control rats, the blood of deficient rats had three times the number of granulocytes, and fewer B lymphocytes (73% of control) and NK cells (38% of control). The numbers of splenic B cells (OX12+), CD5+ (OX19+) and CD4+ (W3/25+) T lymphocytes and NK cells (NKR-P1+) were also significantly reduced. When vitamin A-deficient rats were fed a retinoic acid supplement (4.2 mg all-trans retinoic acid/kg diet) for 28 d, the numbers of blood granulocytes and NK cells equaled those of control rats and NK cell cytotoxicity was significantly elevated. Blood lymphocyte number was increased 40% due to increases of B cells and T cells of the CD5+, CD4+ and CD8+ subsets. These data indicate that vitamin A deficiency affects a number of cells of the immune system and that repletion with retinoic acid effectively reestablishes the number of circulating lymphocytes. In addition, retinoic acid may stimulate NK cell function.

Topics: Animals; Body Weight; Diet; Flow Cytometry; Killer Cells, Natural; Leukocyte Count; Leukocytes; Phenotype; Rats; Rats, Inbred Lew; Spleen; Tretinoin; Vitamin A Deficiency

Vitamin A regulation of specific promoter domains of the phosphoenolpyruvate carboxykinase (PEPCK) gene was tested in a PEPCK/bovine growth hormone (bGH) transgenic mouse model. Vitamin A deficiency causes a significant decrease in hepatic bGH mRNA when expression is driven by either a 533-base-pair (bp) PEPCK promoter fragment (from position -460 to +73) or a 428-bp PEPCK promoter fragment (from position -355 to +73). Treatment of vitamin A deficient transgenic mice with all-trans retinoic acid (RA) increases bGH mRNA levels above those measured with the deficiency. Hepatic retinoic acid receptor (RAR)beta mRNA levels also change with vitamin A deficiency and supplementation, but not RAR alpha mRNA levels. These results indicate that all-trans RA plays a physiologic role in regulating expression of a gluconeogenic gene in liver.

Topics: Animals; Cattle; Gene Expression; Growth Hormone; Liver; Mice; Mice, Inbred C57BL; Mice, Transgenic; Phosphoenolpyruvate Carboxykinase (GTP); Promoter Regions, Genetic; Receptors, Retinoic Acid; Regulatory Sequences, Nucleic Acid; RNA, Messenger; Tretinoin; Vitamin A Deficiency

Homeobox-containing genes may play an important role in establishing embryonic patterns during development of vertebrates. Retinoic acid is able to induce expression of Hox genes in cells in culture and to alter expression patterns in the developing vertebrate embryos. Using wholemount in situ hybridization, we have examined and compared the expression patterns of a homeobox-containing gene, Msx-1, in early normal and vitamin A-deficient quail embryos. At gastrulation stage, Msx-1 is primarily expressed in the posterior half of both normal and vitamin A-deficient embryos. However, the gene is expressed wider and stronger in the vitamin A-deficient embryos. At neurulation stages, Msx-1 is continuously expressed in the posterior region up to Hensen's node and in the edge of the neural fold in both normal and vitamin A-deficient embryos. Notably, in the vitamin A-deficient embryos, Msx-1 is expressed more strongly and is also expressed ectopically in the anterior and precardiac regions. These results provide evidence that endogenous retinoids are involved in the normal expression of Msx-1 in avian embryo and that the expression of Msx-1 is downregulated by endogenous and physiological retinoids in vivo during early avian embryogenesis.

Topics: Animals; Coturnix; Embryo, Nonmammalian; Female; Gene Expression Regulation, Developmental; Homeodomain Proteins; In Situ Hybridization; MSX1 Transcription Factor; Transcription Factors; Tretinoin; Vitamin A Deficiency

Manipulation of vitamin A intake has been associated with altered rates of cytochrome P450 (P450)-mediated microsomal drug oxidation. Dietary vitamin A deficiency reportedly results in decreased rates of P450-dependent substrate oxidation, but the mechanisms underlying these changes remain unclear. In this study, the effects of dietary vitamin A modulation, as well as dietary inclusion of all-trans-retinoic acid (ATRA), on major constitutive P450s were defined. Total microsomal P450 in deficient male rats was decreased to 72% of control (0.63 +/- 0.07 vs. 0.88 +/- 0.08 nmol/mg of protein; P < .05); this was prevented by inclusion of ATRA (12 micrograms/g) in the deficient diet. Dietary vitamin A deficiency decreased rates of P450 2C11-mediated testosterone 2 alpha- and 16 alpha-hydroxylation in rat liver to 44 and 47% of respective adequate control, whereas rates of 6 beta- and 7 alpha-hydroxylation of the steroid were unaltered; inclusion of ATRA into the deficient diet prevented the loss of 2C11 activities. Immunoblot and RNA analysis revealed decreases in P450 2C11 apoprotein and its corresponding mRNA in liver from deficient rats that was prevented by inclusion of ATRA in the deficient diet. Serum testosterone concentrations were reduced in deficient rats and this also was prevented by dietary ATRA. To discern whether this was a direct effect of vitamin A on P450 2C11 regulation, further experiments evaluated the effect of ATRA administration to male rats maintained on standard rat chow (vitamin A-adequate). Dose- and time-dependent decreases in P450 2C11 activity were observed.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Aryl Hydrocarbon Hydroxylases; Base Sequence; Cytochrome P-450 Enzyme System; Cytochrome P450 Family 2; Diet; Down-Regulation; Liver; Male; Molecular Sequence Data; Protein Biosynthesis; Rats; Rats, Wistar; RNA, Messenger; Steroid 16-alpha-Hydroxylase; Steroid Hydroxylases; Testosterone; Tretinoin; Vitamin A Deficiency

The purposes of this study were to determine whether expression of the gene encoding ornithine aminotransferase (OAT) in the rat liver and kidney is regulated by retinoic acid (RA) and to characterize further the role of thyroid hormone in regulating the expression of this gene. The level of OAT messenger RNA (mRNA) was reduced 70% in the liver of animals fed a vitamin A-deficient diet relative to that in animals fed a vitamin A-sufficient diet. RA, administered at a dose of 20 micrograms/rat to A-deficient rats for 1 or 3 days, restored OAT mRNA to near the level observed in animals fed the A-sufficient diet. Retinol was also effective in this regard. T3, when injected alone at a dose of 10 micrograms/100 g BW, had no effect on the level of OAT mRNA in the liver. However, when injected concurrently with RA, T3 blocked the ability of RA to induce OAT mRNA in the liver of rats fed the vitamin A-deficient diet. Animals made both vitamin A deficient and hypothyroid responded to RA in a manner similar to vitamin A-deficient animals. The vitamin A-deficient, hypothyroid rats responded somewhat differently to T3, however. T3 was unable to block the induction of OAT mRNA in the liver of vitamin A-deficient, hypothyroid rats when injected concurrently with RA for 1 day, but did block the induction of OAT mRNA by RA when these two hormones were injected concurrently for 3 days. These data indicate that RA and T3 exert opposing effects on the level of OAT mRNA in the liver. The effects of RA and T3 on OAT mRNA were markedly different in the kidney. Neither vitamin A deficiency nor RA had any apparent affect on the level of OAT mRNA in the kidney. T3, however, increased the level of OAT mRNA in the kidney of vitamin A-deficient rats. In the kidney of vitamin A-deficient, hypothyroid rats, T3 was unable to increase OAT mRNA when injected for 1 day, but did increase this mRNA when injected for 3 days. Together, these data indicate cell-type specific effects of both RA and T3 on the OAT gene.

Topics: Animals; Animals, Newborn; Female; Gene Expression Regulation, Enzymologic; Hypothyroidism; Kidney; Lactation; Liver; Male; Organ Specificity; Ornithine-Oxo-Acid Transaminase; Propylthiouracil; Rats; Rats, Inbred Lew; RNA, Messenger; Thyroxine; Tretinoin; Triiodothyronine; Vitamin A; Vitamin A Deficiency

T3 inhibits transcription of the rat TSH beta gene, and two T3 response elements have been identified that bind T3 receptors and that share sequence homology with the consensus sequence that is also recognized by retinoic acid receptors (RARs). We, therefore, asked whether RA was a regulator of TSH beta gene expression in vivo. Using RNase protection analysis, we found that vitamin A deficiency led to a 2-fold increase in rat pituitary TSH beta messenger RNA (mRNA) levels, which returned to normal 18 h after treatment with RA (20 micrograms/rat). Vitamin A deficiency had no effect on TSH beta mRNA levels in hypothyroid rats. Coadministration of RA and T3 (10 micrograms/100g body wt) to either vitamin A-deficient or vitamin A-deficient, hypothyroid animals caused decreases in TSH beta mRNA content that were indistinguishable from those seen with T3 alone. Surprisingly, vitamin A deficiency had no significant effect on GH mRNA levels in euthyroid or hypothyroid rats. Furthermore, treatment of vitamin A-deficient, hypothyroid animals with RA for either 18 or 72 h had no effect on GH mRNA levels, whereas T3 caused 11-fold and 18-fold increases in GH mRNA, respectively, at these times. We also used transient transfection to test for direct, retinoid receptor-mediated regulation of TSH beta gene transcription by RA. A plasmid TSH beta luciferase, containing 0.8 kilobases of rat TSH beta gene 5'-flanking sequences, exon 1, and 150 base pairs of intron 1, was transfected into CV-1 cells. Cotransfection with RAR alpha and retinoid X receptor-beta induced TSH beta expression by 3.5-fold, and treatment with RA suppressed this induction by 46%. These results show that vitamin A levels play a significant role in regulating the expression of the TSH beta gene, but not the GH gene, in vivo and suggest that RA may suppress TSH beta gene transcription directly by an RAR-retinoid X receptor heterodimer-mediated mechanism.

Topics: Animals; Cells, Cultured; Gene Expression Regulation; Growth Hormone; Luciferases; Pituitary Gland; Rats; Receptors, Retinoic Acid; RNA, Messenger; Thyroid Hormones; Thyrotropin; Transcription, Genetic; Transfection; Tretinoin; Vitamin A Deficiency

Vitamin A-deficient, pregnant rats resorb their fetuses around Day 15 of gestation even when given retinoic acid (RA) daily. This resorption is prevented by the administration of retinol. Retinol was found to be required no later than Day 10 of gestation, and up to 500 micrograms of retinol administered on Day 11 or later was not able to change the course of fetal resorption in these animals. Vitamin A-deficient pregnant rats supported on 40 micrograms all-trans-RA daily that are given 2 micrograms of retinol on Day 10 and then placed on a retinol-sufficient diet at 24 h after administration of the retinol dose gave birth to normal pups. When a single 2-micrograms dose of retinol was administered on Day 10 and the vitamin A-deficient, RA-supported animals continued on the deficient diet supplemented with RA, the fetal resorption did not occur and the fetuses appeared normal when examined on Day 20 of gestation. Experiments using radiolabeled retinol tracer indicated that while there was some accumulation of radioactivity through Day 14 in the fetus, the administered retinol was mostly eliminated from the animal by Day 16. Thus, retinol is clearly required by Day 10 for normal gestation.

Topics: Animals; Female; Fetus; Pregnancy; Pregnancy Complications; Rats; Time Factors; Tretinoin; Vitamin A; Vitamin A Deficiency

The distribution of all-trans-retinoic acid (RA) was investigated using whole-body autoradiography of normal and partial vitamin A deficient (VAD) mice. Retinoic acid receptors (alpha, beta, and gamma) were also studied in normal mice using immunoblotting. Normal and VAD mice were injected with 5 muCi 14C RA. The distribution of RA was quantitatively studied using a computer-assisted image analysis system. 14C RA was incorporated 0.5 hr after RA administration in both normal and VAD mice, while the labelling peak was at 6 hr in most organs in normal and VAD mice. The most intense labeling was found in liver, kidney, intestine, lung, Harderian gland, and salivary gland at all time points. A band of M(r) 51K was found in all mouse tissues by immunoblotting using the polyclonal antibody RAR82 against total RARs or the RAR alpha-specific monoclonal antibody R alpha 13. In some tissue, an additional band of 55-58K was also found. Lung, large intestine, small intestine, testis, seminal vesicle, and spleen contained highest concentration of total RARs, while heart, lung, small intestine, spleen, salivary gland, and preputial gland had the highest concentration of RAR alpha. The uptake of labeled RA correlated well with RAR or RAR alpha concentration in the corresponding tissues.

Topics: Animals; Autoradiography; Blotting, Western; Image Processing, Computer-Assisted; Immunoblotting; Male; Mice; Receptors, Retinoic Acid; Tissue Distribution; Tretinoin; Vitamin A Deficiency

To determine the prevalence of subclinical vitamin A deficiency (vitamin A inadequacy) in Indonesian pregnant women as assessed by the modified relative dose response test.. Cross-sectional study of the vitamin A statuses of pregnant (second trimester) women randomly selected from ten different villages.. West Java, Indonesia.. A group of 144 pregnant women recruited from the local health posts.. Modified relative dose response (MRDR) test, serum retinol determination and gynecological examinations.. The mean (s.d.) MRDR ratio was 0.039 +/- 0.031. Of the women tested, the vitamin A status of 17% was provisionally classified as being marginal (subclinically deficient) (MRDR ratio > or = 0.060), of 35% as being uncertain (MRDR ratio between 0.030 and 0.060) and of 48% as being satisfactory (MRDR ratio < or = 0.030).. If the vitamin A statuses of the 'uncertain' group are also deemed to be unsatisfactory, approximately half of the pregnant Indonesian women tested could benefit from an increased intake of vitamin A.

Topics: Adolescent; Adult; Cross-Sectional Studies; Dose-Response Relationship, Drug; Female; Humans; Indonesia; Nutrition Surveys; Nutritional Status; Pregnancy; Pregnancy Complications; Prevalence; Tretinoin; Vitamin A; Vitamin A Deficiency

The effect of vitamin A on the expression of apoAI and CIII genes in intact rats was studied. In vitamin A-deficient rats, the hepatic level of apoAI mRNA was increased and enhanced by an oral administration of excess retinoic acid(RA). A similar administration to normal rats caused an increase in the apoAI mRNA level in the intestine without affecting that of hepatic mRNA. Though the hepatic level of apo CIII mRNA was not affected by the vitamin A status, the intestinal level was positively regulated by vitamin A. These findings show that vitamin A regulates the expression of apolipoprotein AI and CIII genes in a tissue-specific and complex fashion.

Topics: Animals; Apolipoprotein A-I; Apolipoprotein C-III; Apolipoproteins C; Gene Expression Regulation; Intestinal Mucosa; Intestines; Liver; Male; Rats; Rats, Wistar; Receptors, Retinoic Acid; RNA, Messenger; Tissue Distribution; Tretinoin; Vitamin A; Vitamin A Deficiency

Apolipoprotein A-I (apoA-I) gene expression is known to be regulated by nutritional and hormonal factors. Experiments were conducted to determine the effects of vitamin A deficiency and retinoic acid repletion on the in vivo expression of apoA-I in rat intestine and liver. The relative abundance of apoA-I mRNA (apoA-I/beta-actin ratio) in the intestine did not differ significantly between vitamin A-deficient and -sufficient rats. However, the relative abundance of hepatic apoA-I mRNA of vitamin A-deficient rats was 2.2- to 6-times that of sufficient rats. Even marginal vitamin A status resulted in a significant increase in hepatic apoA-I mRNA expression. Treatment of vitamin A-deficient rats with a single dose of retinoic acid (20 micrograms, 20 h before tissues were collected) reduced the hepatic apoA-I mRNA/beta-actin ratio by about 40%, while further reduction (about 60-65%) was observed after two treatments with retinoic acid. By nuclear run-on assay, the increase in hepatic apoA-I mRNA in vitamin A-deficient rats was attributable to increased transcription of the apoA-I gene. However, immunoblot analysis showed no apparent differences in apoA-I protein in either liver homogenates or plasma of vitamin A-deficient and -sufficient rats. These data indicate that apoA-I gene expression in vivo is sensitive to retinoid status and suggest that there is additional regulation of post-transcriptional events.

Topics: Animals; Apolipoprotein A-I; Blotting, Northern; Cell Nucleus; DNA Probes; Female; Intestinal Mucosa; Liver; Rats; Rats, Inbred Lew; RNA, Messenger; Transcription, Genetic; Tretinoin; Vitamin A Deficiency

To determine whether vitamin A is involved in pancreatic alpha cell function, we tested for (a) effects of vitamin A deficiency on glucagon release from perifused islets and perfused pancreases, and (b) the presence of cytosolic retinol-binding proteins (CRBP) and retinoic acid-binding proteins (CRABP), in the glucagon-secreting alpha cell line, ln-R1-G9. Arginine 19 mM plus glucose 2.8 mM-stimulated glucagon secretion was markedly impaired in islets and pancreases of vitamin A-deficient rats or rats that had at some time been cycled through vitamin A deficiency (ever A-def) despite repletion with retinoids for 2-4 weeks. Insulin secretion was impaired likewise. Repletion starting early in the development of vitamin A deficiency and for a longer period of time (18 or 60 days) did not restore glucagon secretion, but did normalize insulin secretion. CRBP and CRABP were present in ln-R1-G9 cells. We conclude that (a) vitamin A deficiency is associated with a defect in glucagon secretion; (b) The defect in secretion occurs early in the course of vitamin A deficiency; (c) The defect persists despite repletion; and (d) The requirement of vitamin A for secretion and the presence of CRBP and CRABP in glucagon-secreting cells support a physiologic role for vitamin A at the alpha cell level.

Topics: Animals; Arginine; Cell Line; Diterpenes; Female; Glucagon; Glucose; Insulin; Insulin Secretion; Islets of Langerhans; Pregnancy; Radioimmunoassay; Rats; Rats, Sprague-Dawley; Receptors, Retinoic Acid; Retinoids; Retinol-Binding Proteins; Retinol-Binding Proteins, Cellular; Retinyl Esters; Tretinoin; Vitamin A; Vitamin A Deficiency

Retinoic acid (RA) stimulated the production of erythropoietin (Epo) in a human hepatoma cell line, HepG2 cells. The stimulation was due to the accumulation of Epo mRNA. The Epo production in HepG2 cells was also dependent on O2 tension for cell culture but the enhancement of Epo production by RA was independent of O2 tension, indicating that RA exerts its effect through a pathway different from O2. Oral administration of RA to the vitamin A-depleted rats elevated the concentration of Epo in serum. These results suggest that RA up-regulates EPO production in vivo as well as in vitro.

Topics: Animals; Base Sequence; DNA; Erythropoietin; Humans; Liver Neoplasms, Experimental; Molecular Sequence Data; Rats; Tretinoin; Tumor Cells, Cultured; Up-Regulation; Vitamin A Deficiency

In hypovitaminosis A, Ab-mediated immunity is severely impaired. We reported that Trichinella spiralis infection stimulates a strong Th2 cell response in control mice but in vitamin A-deficient mice it stimulates a strong Th1 cell response. Here we investigated the immunobiologic mechanisms underlying this shift from a Th2- to a Th1-dominated response. A kinetic analysis showed that the Th1 cells developed first and IFN-gamma secretion predominated in deficient mice, whereas the Th2 cells developed later and IL-5 and IL-10 secretion predominated in control mice. The IFN-gamma-secreting cell frequencies were the same but cells from deficient mice secreted IFN-gamma sixfold faster than cells from control mice, and retinoic acid addition in vitro decreased that rate 50%. In contrast, the IL-5-secretion rates were the same but the IL-5-secreting cell frequency was lower in deficient mice than in controls, and retinoic acid addition in vitro doubled this frequency independently of its inhibitory effect on IFN-gamma. The APC from deficient mice stimulated greater IFN-gamma release than control APC and retinoic acid addition in vitro decreased this activity 50%. Together these results identify at least three vitamin A activities that balance Th1 and Th2 functions, down-regulating Th1 cell IFN-gamma secretion directly, decreasing activated APC function, and promoting Th2 cell growth and/or differentiation. In this system and perhaps others, the imbalance between regulatory Th1 and Th2 cells is one mechanism underlying poor Ab-mediated immunity in hypovitaminosis A.

Topics: Animals; Antigen-Presenting Cells; Cells, Cultured; Female; Interferon-gamma; Interleukin-10; Interleukin-5; Male; Mice; Mice, Inbred BALB C; T-Lymphocytes, Helper-Inducer; Tretinoin; Vitamin A Deficiency

We describe an animal model to induce the histogenesis of squamous metaplasia of the cervical columnar epithelium, a condition usually preceding cervical neoplasia. This model is based on dietary retinoid depletion in female mice. Control sibling mice fed the same diet but with all-trans-retinoic acid (at 3 micrograms/g diet) showed the normal endocervical epithelial and glandular columnar morphology, typical of a simple epithelium without subcolumnar reserve cells. The stratified squamous ectocervical epithelium of these mice fed all-trans retinoic acid showed intense immunohistochemical staining in basal and suprabasal cells with mono-specific antibodies against keratins K5, K14, K6, K13, and, suprabasally, with antibodies specific for K1 and K10. At the squamocolumnar junction, the adjacent columnar epithelium (termed "suprajunctional") did not show staining for K5, K14, K6, K13, K1, and K10 but specifically stained for keratin K8, typical of simple epithelia and absent from the adjacent ectocervical squamous stratified lining (termed "subjunctional"), in striking contrast. Sections of the squamocolumnar junction from mice kept on the vitamin A-deficient diet for 10 weeks showed suprajunctional isolated patches of reserve cells, proximal and distal to the junction. These cells were detected prior to any symptoms of vitamin A deficiency, such as loss of body weight or respiratory discomfort. The subcolumnar reserve cells induced by vitamin A deficiency displayed positive staining for K5 and K14. As deficiency became severe, the reserve cells occupied the entirety of the suprajunctional basement membrane. This epithelium eventually became stratified and squamous metaplastic, the squamocolumnar junction was no longer discernible, and the entire endocervical epithelium and the endometrial glands lost K8 positivity, while acquiring K5, K14, K6, K13, K1, and K10 keratins typical of the ectocervix under normal conditions of vitamin A nutriture. Vitamin A deficiency also altered keratin expression and localization in squamous subjunctional epithelium. In situ hybridization studies for K1 and K5 mRNA showed their major site of expression at the basal (K5) and immediately suprabasal (K1) cell layers. The localization of both K5 and K1 proteins in these same cell layers, and above, is consistent with transcriptional regulation of these keratins. Early vitamin A deficiency caused the appearance of single subcolumnar reserve cells expressing K5 mRNA. After these ce

Topics: Animals; Carcinoma, Squamous Cell; Cervix Uteri; Diet; Disease Models, Animal; Epithelium; Female; Immunohistochemistry; In Situ Hybridization; Keratins; Metaplasia; Mice; Mice, Inbred BALB C; Mice, Nude; Phenotype; Precancerous Conditions; Retinoids; RNA, Messenger; Tretinoin; Uterine Cervical Neoplasms; Vitamin A Deficiency

The expression of cornifin, a putative cross-linked envelope precursor, was investigated in several squamous differentiating tissues by in situ hybridization and immunohistochemical analysis. Cornifin mRNA and protein, which are absent in the normal mucociliary tracheal epithelium, are induced in the suprabasal layers of the squamous metaplastic tracheal epithelium of vitamin A-deficient hamsters. Similar to the induction of squamous metaplasia in vivo, culture of rabbit tracheal cells in the absence of retinoids results in squamous differentiation and expression of cornifin. This induction of cornifin expression is suppressed by retinoic acid and several of its analogs. Cornifin mRNA and protein are also detected in the suprabasal layers of the squamous epithelium of rabbit esophagus and tongue. The distribution of cornifin in human epidermis was compared with that of two other crosslinked envelope precursor proteins, involucrin and loricrin. The localization of cornifin and involucrin is very similar. Both are induced in the spinous layer and appear at an earlier stage during epidermal differentiation than loricrin. The expression of cornifin is greatly increased in psoriatic skin. Cornifin mRNA is barely detectable in normal epidermis, whereas it is present at relatively high levels in the suprabasal layers of psoriatic epidermis. Topical treatment with RA results in thickening of the skin and increases the level of cornifin mRNA and protein in the upper spinous layers of mouse skin. Cornifin expression correlates generally with squamous differentiation in a variety of tissues and is abnormally regulated in psoriatic skin and in skin treated topically with retinoic acid.

Topics: Administration, Topical; Animals; Cells, Cultured; Cornified Envelope Proline-Rich Proteins; Cricetinae; Esophagus; Gene Expression; Humans; Male; Membrane Proteins; Mice; Protein Precursors; Psoriasis; Rabbits; RNA, Messenger; Skin; Trachea; Tracheal Neoplasms; Tretinoin; Vitamin A Deficiency

Retinol, or one of its metabolites such as retinoic acid (RA), is an important factor in the differentiation and maintenance of integrity of lung epithelium. Retinol deficiency in rats induces morphologic changes in respiratory tract epithelial cells that are histologically similar to those found in human premature infants with bronchopulmonary dysplasia. The exact mechanism of retinoid action in cellular growth and differentiation is not understood, but recently investigators have focused on mechanisms mediated by nuclear RA receptors (RAR). The role of these RAR as regulators of retinoid function is being studied in adult animal tissues and malignant cell lines, but little is known about RAR in developing fetal lung tissue. The purpose of this study was to determine the effect of gestational age and vitamin A deficiency on fetal rat lung nuclear RAR. RAR were also assayed in vitamin A control and vitamin A-deficient adult rat lung. A competitive binding assay and size exclusion HPLC separation were used to quantitate total RAR-specific binding. Binding analysis revealed a single class of receptor binding sites with high affinity (kd approximately 10(-9) M) for RA and RAR saturation at 2-5 nM RA. Specific binding of lung RAR in rat fetuses at 18 d gestation was two to three times greater than in fetuses at 20-21 d gestation, newborn pups, or adults. Western blot analysis revealed a predominance of RAR-beta receptors in fetal lung. Lungs from vitamin A-deficient fetuses demonstrated up-regulation of nuclear RAR.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Carrier Proteins; Cell Nucleus; Female; Fetal Organ Maturity; Fetus; Gestational Age; Kinetics; Lung; Pregnancy; Rats; Rats, Sprague-Dawley; Receptors, Retinoic Acid; Tretinoin; Up-Regulation; Vitamin A Deficiency

Null mutant mice for retinoic acid receptor gamma 2 (RAR gamma 2) or all RAR gamma isoforms were generated. RAR gamma 2 mutants appeared normal, whereas RAR gamma mutants exhibited growth deficiency, early lethality, and male sterility due to squamous metaplasia of the seminal vesicles and prostate. These defects were previously observed in vitamin A-deficient animals and could be prevented by RA administration, demonstrating that RAR gamma mediates some of the retinoid signal in vivo. Congenital defects included Harderian gland agenesis, tracheal cartilage malformations, and homeotic transformations along the rostral axial skeleton, establishing a direct link between RA and patterning of the axial skeleton. We also show that in utero RA-induced lumbosacral truncations are mediated by RAR gamma. The observed RAR gamma null phenotype suggests a high degree of functional redundancy among the RARs. The variable penetrance of some of the observed defects is discussed in light of this redundancy and stochastic variation of gene activity.

Topics: Abnormalities, Multiple; Animals; Base Sequence; Bone and Bones; Carrier Proteins; Cartilage; Embryonic and Fetal Development; Female; Genes, Lethal; Growth Disorders; Homozygote; Male; Mice; Mice, Mutant Strains; Molecular Sequence Data; Receptors, Retinoic Acid; Recombination, Genetic; Tretinoin; Vitamin A Deficiency

The activity of the hepatic enzyme lecithin: retinol acyltransferase (LRAT), thought to catalyze the esterification of retinol for storage, was previously shown to vary directly with the vitamin A (retinol) status of the rat [Randolph, R. K., and Ross, A. C. (1991) J. Biol. Chem. 266, 16453-16457]. The present studies were designed to determine whether liver LRAT activity is regulated in vivo by retinoic acid, a principal active metabolite of retinol. LRAT activity was negligible in the livers of vitamin A-deficient rats. Following treatment with a single 2-micrograms dose of retinoic acid, LRAT activity increased significantly while, following treatment with a single 20-micrograms dose, liver LRAT activity equalled that of vitamin A-sufficient adult rats. Retinoic acid was more effective than an equimolar quantity of retinol in restoring LRAT activity in the vitamin A-deficient rat liver. The increase in hepatic LRAT activity after administration of retinoic acid occurred rapidly, reaching a maximum within 12-16 h but declining again after 48-72 h. Studies were conducted in vivo to gain insight into the level of regulation of LRAT by retinoic acid. The increase in LRAT activity by retinoic acid in the vitamin A-deficient rat was blocked completely by both actinomycin D and cycloheximide. The ability of liver to esterify retinol in vivo was correlated with the in vitro activity of LRAT after retinoic acid induction. We conclude that retinoic acid, an important end product of retinol metabolism, regulates a key aspect of hepatic retinol metabolism through its regulatory activity on liver LRAT.

Topics: Acyltransferases; Animals; Cycloheximide; Dactinomycin; Dose-Response Relationship, Drug; Enzyme Induction; Esterification; Female; Liver; Male; Microsomes, Liver; Phosphatidylcholines; Protein Biosynthesis; Rats; Rats, Inbred Lew; Transcription, Genetic; Tretinoin; Vitamin A; Vitamin A Deficiency

Dietary deficiency in the retinoid precursors of the visual pigment chromophore 11-cis retinal results in the synthesis of photoreceptor outer segments containing opsin in excess of the vitamin A available for rhodopsin regeneration. This suggests that vitamin A-free opsin may be incorporated into newly synthesized outer segment disc membranes. If this opsin is functionally intact, it should be possible convert it to rhodopsin in vivo by providing the appropriate retinoids, and the resulting rhodopsin should should be able to mediate visual transduction. Experiments were conducted to evaluate this possibility and to identify the rate-limiting steps in photoreceptor recovery from retinoid depletion. Rates were maintained on diets either containing or lacking retinoid precursors of 11-cis retinal for 23 weeks, at which time outer segment opsin content greatly exceeded the availability of visual cycle retinoids in the retina. The retinoid-deprived animals were then each given a single intramuscular injection of all-trans retinol. At various time intervals after retinol administration, electroretinograms (ERGs) were recorded on some rats, and retinal rhodopsin contents were determined in others. At similar time intervals, blood and retinal pigment epithelial (RPE) retinoid levels and photoreceptor outer segment size were also determined. No significant increase in retinal rhodopsin content was observed up to 8 hr after injection, despite the fact that by 3 hr, blood retinol levels had recovered to more than 30% of normal. By 1 day after injection, however, rhodopsin levels had recovered to 30% of normal and ERG responses showed increases in visual sensitivity commensurate with the recovery of rhodopsin. The lag in rhodopsin recovery was apparently due to delayed uptake of retinol from the blood by the RPE. Photoreceptor outer segment size was reduced by over 50% in the retinoid- deprived rats and did not begin to recover by 1 day. By 1 week, however, outer segment size had returned to an average of 65% of normal. Commensurate with this regrowth of the outer segments, both rhodopsin levels and visual sensitivity increased between 1 and 7 days after vitamin A administration. Because the rates of recovery in rhodopsin levels and visual sensitivity greatly exceeded the normal rate of new opsin synthesis at short time intervals after vitamin A repletion, it appears that the opsin incorporated into the disc membranes of retinoid-deprived rats is able to form functi

Topics: Animals; Electroretinography; Male; Photoreceptor Cells; Pigment Epithelium of Eye; Rats; Rats, Inbred F344; Retina; Rhodopsin; Rod Cell Outer Segment; Rod Opsins; Tretinoin; Vitamin A; Vitamin A Deficiency

The effects of vitamin A nutritional status on the levels of expression of retinoic acid nuclear receptors (RAR), and the retinoic acid-responsive gene, tissue transglutaminase, were determined in rats. Weanling male Sprague-Dawley rats fed a vitamin A-deficient diet for approximately 7 wk developed vitamin A deficiency, as confirmed by the depletion of liver retinol and retinyl palmitate. Controls were fed the same diet supplemented with 24 mg/kg retinyl acetate. The levels of expression of RAR beta mRNA were approximately 80% lower in bladder, brain, liver, lung and trachea and those of RAR gamma mRNA were approximately 50% lower in bladder, lung and trachea of rats fed the vitamin A-deficient diet than in controls. The levels of expression of RAR alpha mRNA were approximately 90% lower in brain and approximately 30% greater in liver, kidney, intestine and lung of rats fed the vitamin A-deficient diet. Vitamin A deficiency also resulted in reduced expression of tissue transglutaminase in the bladder, lungs and trachea, which paralleled the effects observed for RAR beta and RAR gamma. When vitamin A-deficient rats were subsequently fed a retinol-deficient diet supplemented with retinoic acid for 4 wk, the expression of RAR (beta and gamma) and tissue transglutaminase returned to the control levels. These results indicate that vitamin A nutritional status in rats influences the expression of both RAR and tissue transglutaminase in certain tissues.

Topics: Animals; Blotting, Northern; Carrier Proteins; DNA Probes; Gene Expression Regulation; Male; Nucleic Acid Hybridization; Rats; Rats, Sprague-Dawley; Receptors, Retinoic Acid; RNA; Transglutaminases; Tretinoin; Viscera; Vitamin A Deficiency

Topics: Animals; Female; Immunoenzyme Techniques; Isotretinoin; Kidney; Plasminogen Activator Inhibitor 1; Rats; Rats, Inbred BN; Time Factors; Tissue Plasminogen Activator; Tretinoin; Urokinase-Type Plasminogen Activator; Vitamin A; Vitamin A Deficiency

In this study, we examined effects of retinol deficiency and three retinoids, all-trans-retinoic acid, 13-cis-retinoic acid, and etretin, the aromatic derivative of retinoic acid on nuclear retinoic acid receptor isoforms alpha, beta, and gamma mRNA in rat adipose tissue. Retinol deficiency caused an almost 50% decrease in isoform beta mRNA levels of adipose tissue, whereas little change occurred in the abundance of alpha and gamma isoforms transcripts in this tissue. Intragastric administration of all-trans-retinoic acid to retinol-deficient rats increased the adipose tissue retinoic acid receptor isoforms beta and gamma mRNA levels sixfold and twofold, respectively, in 4 h as compared to adipose tissue of retinol-deficient rats that were administered cottonseed oil. In contrast to this, 13-cis-retinoic acid and etretin at equimolar doses were not effective in inducing beta or gamma isoforms mRNA in retinol-deficient rats. These results show that adipose tissue isoform beta responds to retinol deficiency, and all-trans-retinoic acid rapidly induces beta and gamma mRNA isoforms in this tissue. Thus, retinoic acid may regulate expression of specific genes through its interaction with retinoic acid receptors in adipocytes.

Topics: Adipose Tissue; Animals; Carrier Proteins; Cell Nucleus; Male; Rats; Rats, Inbred Strains; Receptors, Retinoic Acid; Retinoids; RNA, Messenger; Stereoisomerism; Tretinoin; Vitamin A Deficiency

Recently, we have reported that retinoic acid (RA), similarly to retinol acetate, is able to reinitiate spermatogenesis in vitamin A-deficient rats. Here, we investigated the expression of RA receptors RAR alpha, RAR beta, RAR gamma, and retinoid X receptor RXR alpha by Northern blot analysis of poly(A)+ RNA of testes of vitamin A-deficient rats before and after reinitiation of spermatogenesis induced by injection of retinol acetate or RA and testes of 21-day-old and 10-week-old normal rats. In the testis of vitamin A-deficient rats 1.9-, 2.8-, and 3.8-kilobase (kb) transcripts of RAR alpha; 2.8- and 3.3-kb transcripts of RAR beta; 1.8-, 2.8-, and 3.4-kb transcripts of RAR gamma; and two transcripts of RXR alpha of 2.5 and 4.8 kb are expressed. When vitamin A-deficient rats receive RA or retinol acetate, a 3-fold increase in the amount of poly(A)+ RNA per testis can be observed after 8 h, while the amounts of glyceraldehyde-3-phosphate dehydrogenase and sulfated glycoprotein-1 mRNA hardly change. Also, the expression of several transcripts of each RAR type is significantly increased from 1.8- up to 3.6-fold. Moreover, additional transcripts of RAR beta and RXR alpha (1.8 and 1.0 kb, respectively) can be detected. In the testes of 21-day-old rats, three transcripts of each RAR type and two RXR alpha transcripts are expressed. In contrast, in the normal adult rat testis the expression of all RARs, if present, is lower than that in the 21-day-old rat testis or the adult vitamin A-deficient rat testis. The expression of all transcripts of each RAR in the testis of 21-day-old rats shows great similarity with the expression in the testis of the vitamin A-deficient rat after replacement of retinol acetate or RA. These changes in expression indicate that RARs and RXR alpha may play a role in the process of proliferation and differentiation of A spermatogonia, which is induced in vitamin A-deficient rats shortly after replacement of RA or retinol acetate.

Topics: Animals; Blotting, Northern; Carrier Proteins; Diterpenes; Gene Expression; Male; Rats; Rats, Inbred Strains; Receptors, Retinoic Acid; Retinoids; Retinyl Esters; RNA, Messenger; Spermatogenesis; Testis; Transcription, Genetic; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Animals; Carrier Proteins; Gene Expression Regulation; Rats; Receptors, Retinoic Acid; Retinoids; RNA, Messenger; Tretinoin; Vitamin A; Vitamin A Deficiency

Alveolar Type II cells were isolated from control and vitamin A deficient rats and allowed to form a monolayer in plastic dishes for 16-18 hours. The vitamin A content (retinol plus retinyl palmitate) of deficient cells was 50-75% less than in control cells on a per mg protein basis. Isolated Type II cells took up [3H]-retinol, synthesized [3H]-retinyl palmitate, and after 4 hours, 24% of the radioactivity in the Type II cells was [3H]-retinoic acid. Deficiency did not appear to alter retinoic acid synthesis. Phosphatidylcholine (PC) and disaturated phosphatidylcholine (DSPC) synthesis, were slightly less in deficient cells compared to control (95 and 85% respectively). In addition, 10(-6) M and 10(-5) M retinoic acid in the reaction media stimulated both PC and DSPC synthesis by 120-140% in control cells. The stimulating effect of retinoic acid was present in deficient cells as well, but less pronounced (120% with 10(-5) M). Vitamin A deficient Type II cells also had less basal levels of both tissue transglutaminase and epidermal transglutaminase activity than control cells.

Topics: Animals; In Vitro Techniques; Male; Phosphatidylcholines; Pulmonary Alveoli; Rats; Rats, Inbred Strains; Transglutaminases; Tretinoin; Vitamin A; Vitamin A Deficiency

We induced vitamin A depletion to define early and late changes during the histogenesis of squamous metaplasia of hamster tracheal epithelium. An early change is the "minimal morphological change" (MMC), in which the mucociliary epithelium is separated from the basement membrane by a continuous layer of basal cells. Immunohistochemistry showed an exclusive localization of the keratins K5 and K14 in basal cells of normal and MMC epithelia. At the MMC stage no staining was observed above the basal layer with antibodies to K5, but upon progression of the lesion to a squamous focus all cells from basal to terminally differentiated were positive for K5 and K14. In contrast, when we used antibodies to the keratins K6 or K13 all cells were negative in the normal epithelium and in the MMC epithelium. Successive layers of suprabasal squamous cells found in squamous metaplasia failed to express normal epidermal differentiation marker keratins K1 and K10 but expressed the proliferation marker keratin K6 and the internal stratified epithelium keratin K13, not normally found in the epidermis or in the trachea. Hamster tracheal epithelial cells could be maintained in culture in serum-free medium for at least 4 weeks in the presence of retinoic acid (RA). In non-RA-containing medium, cells from vitamin A-deficient hamsters showed markedly reduced growth and an increase in the expression of keratins K5, K6, K13, and K14. Since our previous work had implicated retinoids in the control of cell adhesiveness, we were interested to find out whether changes in cell adhesion occur in vitamin A-deficient hamster tracheal epithelial cells, compared to normal cells. Functional assays demonstrated that hamster tracheal epithelial cells, obtained from non-RA-treated tracheas or maintained in culture, displayed reduced attachment to laminin, compared to RA-treated cells. Immunofluorescence studies did not show a decrease either in the alpha 6 integrin subunit, which was localized in the basal aspect of basal cells, or in basement membrane laminin. However, the expression of laminin-binding protein 37 decreased as the epithelium changed from pseudostratified to stratified. Therefore, a coordinated pattern of changes in keratin gene expression, as well as in the expression of laminin-binding protein 37, the precursor to the cell surface laminin receptor 67LR, and in adhesive properties takes place in tracheal epithelium when its phenotype changes from mucociliary to the preneoplastic s

Topics: Amino Acid Sequence; Animals; Antibodies; Blotting, Western; Cell Adhesion; Cell Adhesion Molecules; Cells, Cultured; Cricetinae; Epithelium; Fibroblasts; Immunohistochemistry; Keratins; Laminin; Male; Mesocricetus; Metaplasia; Molecular Sequence Data; Protein Binding; Trachea; Tretinoin; Vitamin A Deficiency

Using the vitamin A depletion-replacement rat model to obtain testicular synchrony, we examined the reproducibility and degree of synchronization obtained by two different protocols. In the original protocol (A), synchrony was achieved by use of retinol alone. In protocol B, retinoic acid was used during the final days of vitamin A depletion as a supplement to retinol. With protocol A, a total of 56 rats were analyzed by an adaptation of a previously published method for quantifying synchrony. Animals treated by protocol A demonstrated a reproducible degree of synchrony although variability was high among individual animals. A smaller group of animals treated with protocol B demonstrated a lower degree of synchrony. In contrast, the midpoint of synchrony (point in the cycle at which 50% of the stages are more advanced and 50% are less advanced) was a more constant value and was not different between the two treatments. The midpoints of synchrony obtained from both protocols were used to calculate a cycle duration of 300 h for our strain of Sprague-Dawley rats. Our results indicate that while the use of either protocol can reproducibly provide testicular synchrony, protocol A results in a higher degree of synchrony. The ability to synchronize testes to selected stages provides sufficient experimental material for the study of the molecular and cellular events of spermatogenesis.

Topics: Animals; Evaluation Studies as Topic; Male; Rats; Rats, Inbred Strains; Spermatogenesis; Testis; Tretinoin; Vitamin A; Vitamin A Deficiency

Vitamin A-deficient (A-) mice produce poor IgG antibody responses due to a helper T cell dysfunction. We performed retinoid repletion studies to determine the minimum dietary retinyl acetate dose and the most active retinoid for supporting immune function. Dietary retinyl acetate repletion at 2 (R2 group) or 4 (R4 group) microgram/g diet restored serum retinol in A- mice to vitamin A-sufficient (A+) control levels within 24 h. However, in R4 mice, liver retinyl palmitate was restored about twofold faster than in R2 mice; liver retinyl palmitate reached A+ control levels by d 30 in R4 mice but not in R2 mice. We challenged the mice with antigen 24 h post repletion; the R4 mice gave an IgG1 response equal to that of A+ controls, but the R2 mice were comparable with the A- controls. We also compared four retinoids for IgG1 response restoration in vitro; 1 nmol/L retinoic acid fully repleted A- cell IgG1 responses and helper T cell frequencies to the unsupplemented A+ control levels. Retinoic acid was at least 10-fold more active than retinyl acetate or retinaldehyde, and 100-fold more active than retinol. Collectively, our results suggest that retinoic acid is probably the physiologically important metabolite for sustaining IgG immune responses in vivo. We discuss the possible relationship between liver retinyl palmitate levels and availability of retinoic acid to support immune function.

Topics: Animals; Antibody Formation; B-Lymphocytes; Cells, Cultured; Diet; Diterpenes; Female; Immunoglobulin G; Male; Mice; Retinyl Esters; T-Lymphocytes; Tretinoin; Vitamin A; Vitamin A Deficiency

Some morphologic changes of the epidermis of psoriatic skin were observed in organ culture in the presence of absence of vitamin A. Normal and uninvolved and involved psoriatic skin areas, punch biopsied in 3-4-mm diameter specimens, were put in serum-free medium containing no vitamin A with or without delipidized fetal calf serum (FCS) and rotation cultured at 60 rpm under an atmosphere consisting of 95% O2 + 5% CO2. The involved psoriatic skin specimens showed well-developed granular layers after 1 d of culture. The values of labeling indices of 3H-thymidine (3H-TdR) incorporated into the epidermal layers of the cultured specimens of the psoriatic skin were nearly constant during culture for as long as 8 d, although the viable epidermal layer gradually became thinner. Addition of tretinoin to the culture caused uninvolved and involved psoriatic skin specimens to become parakeratotic at concentrations as low as about 2.0 x 10(-6) M and 4.0 x 10(-8) M, respectively, suggesting that the involved psoriatic epidermis is much more sensitive, in terms of keratinization, than uninvolved epidermis to the effect of tretinoin.

Topics: Culture Media; Humans; Male; Organ Culture Techniques; Psoriasis; Skin; Tretinoin; Vitamin A; Vitamin A Deficiency

Rats maintained on a diet deficient in retinol and retinoic acid were given a diet containing retinoic acid for 21-29 days after the start of weight loss. The testes of four of these rats were studied. Spermatogonia of all types were observed, though in lower numbers than in controls, and their mitotic activity was normal. Normal preleptotene spermatocytes were encountered, but no normal spermatocytes in further stages of development were seen. Pale cells that appeared to be in prophase were observed. It was concluded that, in retinol-deficient rats maintained on retinoic acid, the spermatogonial population is qualitatively normal, but quantitatively subnormal, while spermatocyte development is qualitatively and quantitatively abnormal. No evidence of spermatogonial arrest or any other form of synchronization was found in testes of these rats, but when the remaining rats were injected with retinol, the seminiferous epithelium did show stage synchronization at 36 and 128 days after the injection.

Topics: Animals; Diet; Growth; Male; Rats; Rats, Inbred Strains; Sperm Count; Spermatocytes; Spermatogenesis; Spermatogonia; Testis; Tretinoin; Vitamin A Deficiency

Previous investigations have shown that lipofuscin accumulation in the retinal pigment epithelium (RPE) is reduced greatly as a consequence of vitamin A deprivation. The mechanism by which vitamin A regulates RPE lipofuscin deposition remains to be determined. It is possible that retinoids are direct precursors of this substance. Alternatively, vitamin A deficiency may reduce the uptake and processing of other potential precursors. In retinas lacking photoreceptor cells, RPE lipofuscin accumulation is decreased substantially. This finding suggested that components of phagocytosed photoreceptor outer segments may be precursors for RPE lipofuscin. The effect of vitamin A deprivation on RPE lipofuscin content therefore could be the result of reduced outer segment phagocytosis by the RPE of vitamin A-deprived animals. To evaluate this possibility, experiments were conducted to determine whether vitamin A deprivation altered the phagosomal content of the RPE. Rats were fed diets containing or lacking retinoid precursors of 11-cis retinal. Retinoic acid was included in the diets of the vitamin A-deprived animals. After both 10 and 26 weeks, the RPE phagosomal contents were determined in animals from each dietary group. Photoreceptor cell densities also were measured in these rats. At both time points, the RPE phagosomal content was lower significantly in the retinoid-deprived animals than in those fed a vitamin A precursor of the visual pigment chromophore. This reduction was not the result of photoreceptor cell death; the density of these cells was not affected significantly by dietary vitamin A. Thus, it appears that retinoid deprivation reduces the rate of photoreceptor outer segment turnover and, consequently, outer segment phagocytosis by the RPE.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Cell Count; Image Processing, Computer-Assisted; Male; Phagocytosis; Phagosomes; Photoreceptor Cells; Pigment Epithelium of Eye; Rats; Rats, Inbred F344; Tretinoin; Vitamin A Deficiency

The multitude of biological effects of the vitamin A metabolite, retinoic acid, are mediated by nuclear retinoic acid receptors (RARs), which are members of the steroid/thyroid hormone receptor superfamily. RAR-alpha, -beta, and -gamma are encoded by three genes from which multiple isoforms can be generated. Recent studies suggest that the expression of at least some RAR isoforms can be regulated by retinoic acid in certain cell lines. Here we examined regulation of RAR expression in the adult animal. RARs were analyzed by Northern blots from lung, liver, and testes of retinol-deficient rats. Retinol deficiency caused a 65-70% decrease in the mRNA levels of lung and liver RAR-beta, whereas no change was observed in RAR-alpha and -gamma mRNA levels in these organs. In the testes of retinol-deficient animals, two transcripts, RAR-alpha 1 (3.7 kb) and RAR-alpha 2 (2.8 kb), were detected as compared with one RAR-alpha 1 (3.7 kb) transcript in retinol-sufficient testes. When retinol-deficient rats were orally administered 1 dose of retinoic acid (100 micrograms per rat), lung RAR-beta mRNA levels started to increase after 1 hr and reached a 16-fold higher level after 4 hr; after 4 hr these retinoic acid-fed rats also showed a 7-fold increase in liver RAR-beta mRNA levels as compared with levels in the retinol-deficient rats. In contrast, liver, lung, and testes RAR-alpha transcripts remained either unchanged or showed only a slight increase in response to retinoic acid. RAR-gamma was constitutively expressed in lung, and its mRNA levels were induced 2-fold by retinoic acid. These results show tissue diversity in the rapid induction of RAR-beta and RAR-gamma by retinoic acid in the adult animal and suggest distinct roles for the various receptor isoforms in the control of the retinoid response.

Topics: Animals; Blotting, Northern; Carrier Proteins; Gene Expression; Liver; Lung; Rats; Receptors, Retinoic Acid; RNA, Messenger; Tretinoin; Vitamin A Deficiency

All-trans retinoyl beta-glucose was chemically synthesized in good yield by reaction of retinoyl fluoride with glucose. Retinoyl glucose, which is soluble in water, shows growth-promoting activity similar to retinyl acetate in vitamin A-deficient rats. In metabolic studies, retinoyl glucose was found to be hydrolyzed to retinoic acid, but at a slower rate. The possible therapeutic uses of retinoyl glucose are discussed.

Topics: Animals; Body Weight; Chromatography, High Pressure Liquid; Glucose; Male; Rats; Rats, Inbred Strains; Retinoids; Tretinoin; Vitamin A Deficiency

We report the results of a histochemical study, using polyclonal antipeptide antibodies to the different TGF beta isoforms, which demonstrates that retinoic acid regulates the expression of TGF beta 2 in the vitamin A-deficient rat. Basal expression of TGF beta 2 diminished under conditions of vitamin A deficiency. Treatment with retinoic acid caused a rapid and transient induction of TGF beta 2 and TGF beta 3 in the epidermis, tracheobronchial and alveolar epithelium, and intestinal mucosa. Induction of TGF beta 1 expression was also observed in the epidermis. In contrast to these epithelia, expression of the three TGF beta isoforms increased in vaginal epithelium during vitamin A deficiency, and decreased following systemic administration of retinoic acid. Our results show for the first time the widespread regulation of TGF beta expression by retinoic acid in vivo, and suggest a possible mechanism by which retinoics regulate the functions of both normal and pre-neoplastic epithelia.

Topics: Animals; Colon; Epidermis; Female; Gene Expression Regulation; Immunohistochemistry; Intestinal Mucosa; Intestine, Small; Lung; Rats; Rats, Inbred Strains; Transforming Growth Factor beta; Tretinoin; Vagina; Vitamin A Deficiency

Dietary vitamin A deficiency in young rabbits caused advanced squamous metaplasia with keratinization of conjunctival epithelium and concomitant reduced paracellular permeability to 3H-mannitol. Both morphologic and permeability changes were reversed with systemic administration of vitamin A. In adult rabbits, vitamin A deficiency caused milder changes of goblet cell loss and increased cellular stratification in conjunction with reduced permeability in the conjunctiva-like epithelium that covers the vascularized cornea after chemical injury with n-heptanol. Topically applied retinoid (tretinoin 0.1%) did not affect the morphology and permeability of the normal corneal or conjunctival epithelium of rabbits that were not vitamin A deficient. These studies showed that altered permeability is associated with the epithelial abnormality during vitamin A deficiency and helped clarify the physiologic function of retinoids in the ocular surface epithelia in the nondeficient state.

Topics: Administration, Topical; Animals; Cell Differentiation; Conjunctiva; Cornea; Epithelium; Mannitol; Perfusion; Permeability; Rabbits; Tretinoin; Vitamin A Deficiency

T lymphocytes from vitamin A-deficient (A-) mice show a decreased ability to stimulate B lymphocytes for Ag-specific secondary IgG1 responses in vivo and in vitro. Experiments reported here traced the molecular basis for this functional defect to an overproduction of IFN-gamma by A- CD4+ T cells compared with cells from A-sufficient (A+) mice. Secretion of IL-2 and IL-4 by cells from A- and A+ mice was equivalent. Retinoic acid supplementation in vitro decreased IFN-gamma secretion from A- T cells, indicating that IFN-gamma production is retinoid-responsive. Adding IFN-gamma neutralizing antibodies to cultures established with cells from immune A- mice substantially increased IgG1 production, whereas IL-4 addition moderately increased IgG1 production. Adding retinoic acid to the cultures either at initiation, or 48 h later, fully restored IgG1 production by A- cultures to the level of A+ control cultures. These results are consistent with a role for vitamin A in negatively regulating IFN-gamma secretion.

Topics: Animals; Cells, Cultured; Immunoglobulin G; Interferon-gamma; Interleukin-2; Interleukin-4; Mice; T-Lymphocyte Subsets; T-Lymphocytes; Transcription, Genetic; Tretinoin; Vitamin A Deficiency

The effects of feeding retinoic acid for 2 and 6 days on the metabolism of labeled retinol in tissues of rats maintained on a vitamin A deficient diet was studied. The metabolites of retinol were analyzed by high performance liquid chromatography. Feeding retinoic acid for 2 days significantly reduced the blood retinol and retinyl ester levels without affecting the vitamin A content of the liver. In intestine and testis the content of labeled retinoic acid was decreased significantly by dietary retinoic acid. Addition of retinoic acid to the diet for 6 days resulted, in addition to decreased blood retinol and retinyl ester values, in an increase in the retinyl ester values in the liver. The accumulation of retinyl ester in the retinoic acid fed rat liver was accompanied by an absence of labeled retinoic acid. Kidney tissue was found to contain the highest levels of labeled retinoic acid, retinol, and retinyl esters; dietary retinoic acid did not alter the concentrations of these retinoids in the kidney during the experimental period. Since kidney retained more vitamin A when the liver vitamin A was low and also dietary retinoic acid did not affect the concentrations of radioactive retinoic acid in the kidney, it is suggested that the kidney may play a major role in the production of retinoic acid from retinol in the body.

Topics: Animals; Chromatography, High Pressure Liquid; Diet; Liver; Male; Rats; Rats, Inbred Strains; Tretinoin; Vitamin A; Vitamin A Deficiency

The effect of various doses of retinoic acid (RA) on the seminiferous epithelium in vitamin A-deficient rats has been studied. Although it was generally thought that RA was not able to reinitiate spermatogenesis in vitamin A-deficient rats, one injection of 5 mg RA strongly stimulated the proliferative activity of A-spermatogonia within 24 h, as evidenced by a 7-fold increase in the number of bromodeoxyuridine-labeled A-spermatogonia. Ten days after RA administration, B-spermatogonia or preleptotene spermatocytes were seen in most of the seminiferous tubules. After 15 days, zygotene spermatocytes were present. Hence, RA is able to induce a massive and synchronized development of A-spermatogonia into spermatocytes. When RA was given once, combined with a RA-containing diet, only few of the zygotene spermatocytes present on day 15 were able to develop into pachytene spermatocytes, which did not develop into spermatids. In subsequent epithelial cycles new B-spermatogonia and spermatocytes were formed, although in lower numbers than during the first cycle after RA injection. When RA was given once a week, the formation of B-spermatogonia and preleptotene spermatocytes continued at a higher level. Also, more pachytene spermatocytes were formed, some of which were able to develop into spermatids. Finally, when RA was injected twice a week, even more pachytene spermatocytes and round spermatids were found after 36 days, and after 49 days elongated spermatids were found in all animals. It is concluded that RA, similar to retinol, is able to induce synchronous proliferation and differentiation of A-spermatogonia. When repeated injections are given, RA is able to support the full development of spermatogenic cells into elongated spermatids.

Topics: Animals; Cell Survival; Diet; Dose-Response Relationship, Drug; Injections; Male; Rats; Rats, Inbred Strains; Seminiferous Tubules; Sertoli Cells; Spermatocytes; Spermatogenesis; Spermatogonia; Tretinoin; Vitamin A Deficiency

Eight-week-old male Sprague--Dawley rats were dosed by gavage with 90 mg/kg of Ro 23-2895, (all-E)-9-[2-(nonyloxy)phenyl]-2,4,6,8 nonatetraenoic acid, dissolved in Tween 80. Treated animals (n = 3--4) were sacrificed after 3, 7, 11 and 21 days of dosing. Control rats (n = 3) received an equal volume of Tween 80 and were sacrificed after 3 or 21 days. Cross sections of formalin fixed testes were embedded in glycolmethacrylate, sectioned at 3 microns, and stained with periodic acid-Schiff and hematoxylin. No morphologic alterations were observed in the control rats or in treated rats after 3 days. After 7 days of treatment, there were occasional tubules in which there was a delayed release of mature sperm and occasionally the retained sperm were being resorbed. The frequency and severity of these morphologic changes was increased after 11 days of treatment, and round spermatids were occasionally observed with marginated chromatin in their nuclei. After 21 days of treatment, there was a significant reduction in testicular weight accompanied by marked degenerative changes and in some cases almost a complete desquamation of the germinal epithelium. Multinucleated giant cells and germ cells with marginated chromatin in their nuclei were commonly observed and there was moderate to severe oligospermia in the tubules. Sertoli cell nuclei were swollen and showed lucent, vesiculated nucleoplasm. In a parallel 21-day study, treated rats (n = 10) showed an 80% reduction in plasma retinol and a 56% decrease in testicular retinol compared to vehicle-treated rats (n = 10). A 53% decrease in plasma testosterone levels was also observed in treated rats. The testicular lesions produced by treatment with Ro 23-2895 were similar to vitamin A deficiency, which supports the hypothesis that high doses of synthetic retinoids may cause testicular degeneration through interference of normal retinol homeostasis.

Topics: Administration, Oral; Animals; Anti-Inflammatory Agents, Non-Steroidal; Male; Rats; Rats, Inbred Strains; Spermatogenesis; Testis; Testosterone; Tretinoin; Vitamin A; Vitamin A Deficiency

Dietary deficiency in the retinoid precursors of the visual pigment chromophore 11-cis-retinal eventually results in selective degeneration of the photoreceptor cells of the vertebrate retina. Early effects of retinoid deficiency are depletion of rhodopsin from the retina and vesiculation of the photoreceptor outer segment disc membranes. Experiments were conducted to determine whether these early changes were accompanied by an alteration of the opsin content of the disc membranes. After being fed a retinoid-deficient diet containing retinoic acid for 26 weeks, the rhodopsin content of rat retinas was reduced by over 85%. Both the diameters and the lengths of the outer segments decreased significantly. However, immunocytochemical and freeze-fracture analyses indicated that retinoid deficiency did not lower opsin density in the outer-segment disc membranes. These findings indicate that in the rat, opsin synthesis and disc assembly are coordinated processes that remain coupled despite reduced availability of the vitamin A chromophore. The fact that disc size decreases and disc synthesis eventually ceases in retinoid-deprived rats indicates that specific retinoids are essential for disc morphogenesis. The mechanism by which these retinoids regulate disc assembly remains to be determined.

Topics: Analysis of Variance; Animals; Diet; Freeze Fracturing; Immunoenzyme Techniques; Male; Microscopy, Fluorescence; Rats; Rats, Inbred F344; Rhodopsin; Rod Cell Outer Segment; Tretinoin; Vitamin A Deficiency

The incorporation of [2-3H]mannose into dolichyl phosphate mannose and glycoproteins is markedly reduced in livers of vitamin A-deficient hamsters. To determine whether vitamin A deficiency selectively alters the level of mannose incorporation into sugar phosphates and sugar nucleotides, we studied the in vivo incorporation of [2-3H]mannose, [5-3H]glucose, and [4,5-3H]galactose into sugar phosphates and sugar nucleotides. Male hamsters fed either a vitamin A-depleted or a retinoic acid-supplemented (3 micrograms/g) diet were used at 4, 6 and 8 wk of age; the animals were killed at various time points after an intraperitoneal injection of the radiolabeled sugar. A two- to threefold increase in the amount of [2-3H]mannose was found in liver of hamsters fed a vitamin A-depleted diet for 4 wk, resulting in enhanced incorporation into mannosyl-phosphate and guanosine diphosphate (GDP) mannose. As deficiency progressed, there was a smaller increase in [2-3H]mannose and a significant decrease in [3H]mannose-phosphate and GDP-[3H]mannose, suggesting a decreased mannose kinase activity. [5-3H]Glucose-labeled livers showed no difference in the total uptake of the label or its incorporation into uridine diphosphate glucose and galactose-phosphate during the 8-wk study. However, the synthesis of glucosyl-phosphate was reduced by 50 to 90% at 6 and 8 wk of deficiency, suggesting an impaired gluco-kinase activity. In hamsters injected with [4,5-3H]galactose only [3H]glucose was found within 5 min in the free sugar fraction. In contrast, as much as 70% of the label in the sugar phosphate and sugar nucleotide fraction remained as [3H]galactose even at 60 min. These effects on sugar, sugar phosphate and sugar nucleotide formation in part may explain the effects of vitamin A deficiency on glycoconjugate biosynthesis.

Topics: Animals; Cricetinae; Diet; Galactose; Glucose; Guanosine Diphosphate Mannose; Liver; Male; Mannose; Mesocricetus; Models, Biological; Nucleoside Diphosphate Sugars; Sugar Phosphates; Time Factors; Tretinoin; Uridine Diphosphate Glucose; Vitamin A Deficiency

Topics: Animals; Cytochrome P-450 Enzyme System; Diet; Glucuronosyltransferase; Liver; Male; Rats; Rats, Inbred Strains; Tretinoin; Vitamin A; Vitamin A Deficiency

The modified relative-dose-response (MRDR) assay has been validated in rats as a function of vitamin A status and tested in a group of American children. In this study the MRDR assay was applied to West Javan children who are at risk of being vitamin A deficient. Of 86 children enrolled, 75 were tested. In a time-course study involving 22 children aged 3.7-5.3 y, blood samples were taken at different times after doses of 0.35 mumol 3,4-didehydroretinyl acetate/kg body wt. Generally, the ratio of dehydroretinol to retinol (DR-R ratio) peaked between 4 and 8 h. Thereafter, in a survey of 53 children aged 0.6-4.8 y, single blood samples were drawn 5 h after the dehydroretinyl acetate dose. The DR-R ratio ranged from 0.0028 to 0.169. With a DR-R ratio of 0.03 as the cutoff value, 62% of the children were judged to be of marginal vitamin A status.

Topics: Anthropometry; Child, Preschool; Dose-Response Relationship, Drug; Humans; Indonesia; Infant; Nutritional Status; Time Factors; Tretinoin; Vitamin A; Vitamin A Deficiency

Retinoids have chemopreventive activity for epithelial tumors in a variety of systems, including the two-stage tumorigenesis system of mouse skin in which only the promotion stage is inhibited. We asked whether dietary vitamin A deficiency could affect the skin tumorigenic response, prior to major changes in body weight or general health of the animals. Two regimens were tested to induce vitamin A deficiency. SENCAR mice were either (a) fed a vitamin A-deficient diet from 4 or 9 weeks of age or (b) their mothers were fed the diet from the time of birth of the experimental animals which were then weaned on the same diet. The latter regimen produced typical symptoms of vitamin A deficiency in the offspring by Weeks 12-14 and all the mice died by Week 19; the former regimen permitted sufficient accumulation of retinol and its esters to sustain life for up to 45 and 75 weeks, respectively, in the majority of mice. For our experiments, vitamin A depletion was produced by placing the mothers on the deficient diet at birth of the experimental animals. A single topical dose of 20 micrograms of 7,12-dimethylbenz(a)anthracene (DMBA) was used as the initiator at 3 weeks of age and 1 to 2 micrograms of 12-O-tetradecanoylphorbol-13-acetate (TPA) once weekly as the tumor promoter for 10 weeks (from Week 4 through 13 of the experiment). Fifty-five % of mice (n = 40) on Purina laboratory chow (mean body weight, 31.4 g) developed skin tumors (2.58 per mouse) at 12 weeks, versus 2.5% (0.05 papillomas per mouse) of mice (n = 40) kept on the purified vitamin A-deficient diet (mean body weight, 30.3 g), a 98% decrease in tumor/mouse. Retinoic acid (RA) (1-3 micrograms/g diet) supplementation after Week 12 caused a rapid tumorigenic response in 95% of the mice by week 22. This tumor response occurred to a reduced extent in the absence of continued TPA treatment up to Week 13. Even though tumor incidence increased within 1 week of RA and 95% of the mice showed the tumorigenic response, the number of tumors per mouse was about 50% of that observed in mice maintained on standard Purina diet. This was confirmed in an experiment in which the mice were maintained for life either on Purina or on the RA (3 micrograms/g) containing purified diet, the latter being the control group for the effect of vitamin A deficiency on skin tumorigenesis.(ABSTRACT TRUNCATED AT 400 WORDS)

Topics: 9,10-Dimethyl-1,2-benzanthracene; Animals; beta Carotene; Carotenoids; Cocarcinogenesis; Diet; Female; Liver; Male; Mice; Neoplasms, Experimental; Skin Neoplasms; Tetradecanoylphorbol Acetate; Tretinoin; Vitamin A Deficiency; Weight Loss

Administration of a single oral dose (10 micrograms/kg) of tetrachlorodibenzo-p-dioxin (TCDD) caused a 33% decrease in retinyl esters in the livers of male rats, but a 13-fold increase in retinyl esters in the kidney and a 3-fold increase in serum retinol. Liver and kidney microsomal uridine diphosphoglucuronosyltransferase (UDPGT) activity toward all-trans-retinoic acid was increased 3.7- and 2.6-fold, respectively, ten days following exposure to TCDD. Verification of the in vitro formation of [3H]retinoyl beta-glucuronide (RG) was by cochromatography with authenic RG on reversed phase high pressure liquid chromatography (HPLC), identification of retinoic acid as the hydrolysis product after beta-glucuronidase treatment, and the characterization of the all-trans-retinoyl glucuronide by negative fragment mass spectroscopy, fast atom bobardment. We conclude that increased retinoic acid glucuronidation may be a contributing factor to the hepatic depletion of vitamin A and the increased excretion of vitamin A metabolites following TCDD exposure.

Topics: Animals; Chromatography, High Pressure Liquid; Dioxins; Glucuronosyltransferase; Kidney; Kinetics; Liver; Male; Mass Spectrometry; Microsomes, Liver; Polychlorinated Dibenzodioxins; Rats; Rats, Inbred Strains; Reference Values; Tretinoin; Vitamin A; Vitamin A Deficiency

The effect of all-trans retinol and retinoic acid (RA) on the local and systemic immune system of broiler chicks was examined. Chicks fed diets supplemented with .2 micrograms retinol/g of diet had significantly greater serum immunoglobulin IgG, IgM, and IgA concentrations 5 wk after primary vaccination with live B1 strain of Newcastle disease virus (NDV) than did chicks fed 2 micrograms retinol/g of diet. Eight days after a second vaccination with an inactivated La Sota strain of NDV, serum, intestinal, and tracheal IgG and IgM concentrations were higher in chicks fed a diet without retinol or retinoic acid (RA) than in chicks fed either 2 micrograms of retinol or RA/g of diet, respectively. Despite an increase in serum immunoglobulin concentration, serum antibody titer in response to the second NDV vaccination was significantly lower in chicks fed a vitamin A-deficient diet than in chicks fed adequate retinol and RA-supplemented diets. Eight days after revaccination, IgA concentrations in bile were also significantly lower in vitamin A-deficient chicks than in controls. Serum IgG, IgM, IgA, biliary IgA, and antibody responses were greater in chicks fed diets supplemented with 2 micrograms of RA/g of diet than in chicks fed an equivalent amount of vitamin A in the form of retinol.

Topics: Animals; Chickens; Immunoglobulin A; Immunoglobulin G; Immunoglobulin M; Immunoglobulins; Male; Newcastle Disease; Tretinoin; Viral Vaccines; Vitamin A; Vitamin A Deficiency

An enzyme activity which converts retinal to retinoic acid was found in the cytosol of rat kidney. The oxidation of retinal was pH-, temperature-, time- and protein-dependent. Under the assay conditions employed, the oxidase activity had an apparent Km of 125 microM toward all-trans retinal. n-Propylgallate, butylated hydroxytoluene and quinacrine inhibited the reaction. The inhibition caused by quinacrine can be partly reversed by FAD. p-Hydroxymercuribenzoate, a sulfhydryl cross-linking agent, was a potent inhibitor. 4'-(9-Acridinylamino)methanesulfon-anisidide, an inhibitor of aldehyde oxidase, inhibited the reaction by 77% at a concentration of 3 mM. All-trans retinal reversed the inhibition caused by acetaldehyde and 2-aminobenzaldehyde. Retinol inhibited the reaction, but retinoic acid did not. The specific activity of the enzyme was increased by vitamin A deficiency. These data indicate that retinal-oxidizing enzyme activity found in the kidney is a sulfhydryl flavoprotein and its activity is dependent on the vitamin A levels of the tissues.

Topics: Animals; Cytosol; Flavin-Adenine Dinucleotide; Hydrogen-Ion Concentration; Kidney; Kinetics; Male; Oxidation-Reduction; Propyl Gallate; Quinacrine; Rats; Retinaldehyde; Retinoids; Temperature; Time Factors; Tretinoin; Vitamin A Deficiency

Ornithine decarboxylase (ODC, EC 4.1.1.17) activity was measured, without exogenous stimulation, in the liver, oesophagus and lung of Wistar rats which were vitamin A deficient or supplemented with retinol or retinoic acid. The enzyme basal activity in such deficiency conditions was higher, when compared with controls, in the oesophagus and especially in the lungs. Retinoic acid normalized enzyme activity only at high doses (300 micrograms/day). In the liver, initial retinol deficiency did not sensitively modify ODC activity, and retinoic acid then stimulated the enzyme abnormally. This phenomenon could not be observed at later stages of vitamin deficiency (but there again without cytological abnormalities or thymidine incorporation disturbances): liver ODC response then became comparable to that of other tissues. These results highlight the particular basal hyperactivity of pulmonary ODC during the initial stages of vitamin A deficiency, indicative of an enhanced tendency to cell proliferation. A special stimulating effect of retinoic acid on ODC, contemporary with early deficiency, was observed in the liver; this effect was not observed at a later stage in normally fed rats.

Topics: Animals; Autoradiography; Dose-Response Relationship, Drug; Esophagus; Liver; Lung; Male; Ornithine Decarboxylase; Rats; Rats, Inbred Strains; Tretinoin; Vitamin A; Vitamin A Deficiency

In this study, we examined, by ultrastructural autoradiography, the uptake and intracellular transport of [3H]all-trans-retinoic acid ([3H]RA) in the livers of vitamin A-deficient hamsters. Four-week-old animals were administered 25 microCi of [3H]RA by gavage, and, at different intervals thereafter, one animal was sacrificed. Their livers were excised and processed for autoradiography. Radioactive grains were observed to pass randomly through the plasma membrane by diffusion. No evidence of retinoid internalization by endocytosis was observed. Between 1 and 30 min after gavage, the radioactivity in parenchymal cells was associated mainly with rough endoplasmic reticulum (RER) and mitochondria. The labeling over nuclei was apparent at 1 min, remained relatively high up to 30 min, and subsequently decreased. At 2 and 5 hr, only a few grains were observed over nuclei, RER and mitochondria. At 24 hr, most of the labeling was associated with endothelial cells and sinusoidal spaces, indicating mobilization of [3H]RA from the liver. The results indicate that [3H]RA is transported through the plasma membrane by transmembrane diffusion without endocytosis and, after entering the cells, the ligand is rapidly translocated into nuclei.

Topics: Animals; Autoradiography; Cricetinae; Liver; Microscopy, Electron; Tretinoin; Vitamin A Deficiency

When an [35S] labeled mixture of methionine and cysteine was injected intratesticularly into retinol-deficient rats, two hours later more than 980 cytosolic proteins were detected by computer aided two dimensional gel electrophoresis. Furthermore, two hours after oral refeeding retinyl acetate as the source of retinol to retinol deficient rats, synthesis of 286 proteins was inhibited and that of 101 proteins was activated. Refeeding with retinoic acid leads in two hours to even higher inhibition of protein synthesis and the labeling patterns of proteins are not identical when compared to retinol refed rats. The results indicate that retinol or retinoic acid quickly influence expression of many proteins and suggest that retinol action in the testes is not identical to that of retinoic acid.

Topics: Animals; Cytosol; Electrophoresis, Polyacrylamide Gel; Male; Protein Biosynthesis; Rats; Software; Testis; Tretinoin; Vitamin A; Vitamin A Deficiency

3,4-Didehydroretinol (vitamin A2, DR, dehydroretinol), a naturally occurring analogue of retinol (vitamin A1, R), is active in vision, growth and cellular differentiation but is converted to retinol in very small amounts, if at all. When vitamin A-depleted rats were given 500 micrograms of R acetate, a naturally occurring mixture of 480 micrograms DR ester and 20 micrograms R ester or 500 micrograms DR acetate orally in corn oil, serum levels of all administered retinoids peaked between 3.5 and 5 h and then declined. When an oral dose of 600 micrograms DR/kg body wt was administered to rats with various liver reserves of vitamin A, the serum ratio of DR to R at 3.5 h was inversely related to the liver reserves of vitamin A below approximately 2 micrograms/g liver. Because the administration of DR does not affect serum R values, a single blood sample taken at 3.5 h might provide information analogous to that obtained from two blood samples in the conventional relative dose-response method.

Topics: Animals; Chromatography, High Pressure Liquid; Female; Liver; Rats; Rats, Inbred Strains; Tretinoin; Vitamin A; Vitamin A Deficiency

These studies were performed to follow a spectrum of relevant parameters in male Syrian golden hamsters fed either a vitamin A-deficient diet or the same diet supplemented with retinoic acid at 3 micrograms/g diet. Body weight and life span were not affected by the vitamin A-deficient diet until after 6-7 wk. Squamous metaplastic lesions of the Formalin-fixed tracheas were not generally observed in the hamsters fed the deficient diet until 6-7 wk, at which time blood retinol and liver retinyl palmitate levels had also decreased. Blood glucose levels remained normal (90 mg/dl) until about 7 wk but declined to about 40% of normal at 9 and 10 wk. Dietary retinoic acid supplementation of the vitamin-deficient diet (3 micrograms/g diet) inhibited the loss of retinol from blood and of retinyl palmitate from the liver so that these compounds were still present at 10 wk, but were not detectable in hamsters fed the vitamin A-deficient diet without retinoic acid.

Topics: Animals; Blood Glucose; Cricetinae; Diet; Diterpenes; Epithelium; Growth; Life Expectancy; Liver; Male; Mesocricetus; Metaplasia; Retinyl Esters; Trachea; Tretinoin; Vitamin A; Vitamin A Deficiency

We studied the effects of vitamin A deficiency and repletion on rat insulin release and islet cellular retinol binding protein (CRBP) and cellular retinoic acid binding protein (CRABP). Biphasic insulin release from vitamin A-deficient perifused islets was markedly impaired. Release remained impaired with retinoic acid (RA) repletion, 2 micrograms/g diet compared to release from islets of rats repleted with retinol in the form of retinyl palmitate, 4 micrograms/g diet. Release normalized with RA, 8 micrograms/g diet. Vitamin A deficiency did not affect islet insulin content, cell size, number or structure. In vivo, vitamin A-deficient rats had impaired glucose-induced acute insulin release and glucose intolerance, which improved with repletion. Normal islets had greater concentrations of CRBP than CRABP; vitamin A deficiency reduced CRBP but not CRABP levels. We conclude retinol is required for normal insulin secretion. Retinoic acid may substitute for retinol in this function.

Topics: Animals; Blood Glucose; Body Weight; Carrier Proteins; Energy Intake; In Vitro Techniques; Insulin; Insulin Secretion; Islets of Langerhans; Proteins; Rats; Receptors, Retinoic Acid; Retinol-Binding Proteins; Retinol-Binding Proteins, Cellular; Tretinoin; Vitamin A; Vitamin A Deficiency

A state of pure vitamin A deficiency, without any clinical manifestations, rapidly induces a stimulation of ornithine decarboxylase (ODC) activity and some oesophageal mucosal abnormalities (hyperkeratosis, dyskeratosis, cytonuclear abnormalities) in rats treated with N-nitrosobenzylmethylamine (NBMA). Retinol deficient rats fed with retinoïc acid (all-trans) show a mucosal ODC induction, but no morphological lesion. The association of retinoïc acid + retinol, as does retinol alone, prevents simultaneously histological lesions and enzymatic induction. In the liver of vitamin A deficient rats treated with NBMA, a stimulation of the ODC system, without any macroscopical lesions, has been observed.

Topics: Animals; Dimethylnitrosamine; Enzyme Induction; Esophageal Neoplasms; Keratosis; Neoplasms, Experimental; Ornithine Decarboxylase; Rats; Tretinoin; Vitamin A; Vitamin A Deficiency

The regulation of Sertoli cell transferrin and sulfated glycoprotein-2 (SGP-2) mRNA levels by vitamin A was studied in vitamin A deficient rats. Vitamin A deficiency differentially affected the levels of these mRNAs. Sertoli cells from vitamin A deficient rats contained 3-fold less transferrin mRNA and 1.8-fold more SGP-2 mRNA than Sertoli cells from normal rats. Vitamin A deficiency did not result in changes in the amount of transferrin mRNA per liver cell. When vitamin A deficient rats were fed a retinol supplemented diet, Sertoli cell transferrin and SGP-2 mRNA levels returned to normal. In contrast, dietary retinoic acid did not support recovery of the levels of the two mRNAs. The regulation of transferrin and SGP-2 mRNA levels was further examined in vitamin A deficient rats that had received subcapsular testicular injections of vitamin A and in vitamin A supplemented cultures of Sertoli cells from vitamin A deficient rats. Under these conditions, retinol and retinoic acid both stimulated transferrin mRNA levels but did not affect SGP-2 mRNA levels. Retinol did not inhibit the turnover of transferrin mRNA in Sertoli cell cultures suggesting that the stimulation of transferrin mRNA levels by retinol is due to increased transcription of the transferrin gene. A mathematical correlation between the Sertoli cell transferrin mRNA levels and the weight of the testes or the number of germinal cells was observed.

Topics: Animals; Cells, Cultured; Clusterin; Glycoproteins; Male; Molecular Chaperones; Organ Size; Rats; Rats, Inbred Strains; RNA, Messenger; Sertoli Cells; Testis; Transferrin; Tretinoin; Vitamin A; Vitamin A Deficiency

This study was designed to determine the effect of polyprenoic acid on wound healing in rats after thermal injury and to compare the effect with healing in vitamin A deficient controls and in retinol-fed vitamin A deficient rats. Both polyprenoic acid and retinol visibly accelerated wound healing after thermal injury. Both enhanced the induction of platelets in the peripheral blood but neutrophils were not affected by polyprenoic acid. Histologically, in the hypo-vitamin A rat, polyprenoic acid administration increased both capillary formation and also the production of fibroblasts and collagen deposition in the wound by comparison with control vitamin A deficient animals; similar effects were produced by retinol feeding. These results indicate that both polyprenoic acid and retinol can facilitate wound healing. 3H-thymidine incorporation into dermal tissues showed that in polyprenoid-fed rats capillaries multiplied 1.3 times, epithelial cells 2.1 times, and fibroblasts 2.0 times faster than those in vitamin A-deficient animals; 3H-thymidine incorporation was greater after polyprenoid feeding than after retinol feeding. Polyprenoid thus stimulates both collagen deposition and neo-vascularization within burns, and so accelerates healing.

Topics: Animals; Burns; Capillaries; Collagen; Fibroblasts; Leukocyte Count; Mitosis; Neutrophils; Platelet Count; Rats; Rats, Inbred Strains; Skin; Thymidine; Tretinoin; Vitamin A; Vitamin A Deficiency; Wound Healing

Ceruloplasmin, a copper-containing acute phase plasma protein, has been shown to be regulated by 13-cis retinoic acid in rats. Ceruloplasmin activity was significantly increased within 24 h and remained elevated for at least 72 h after a single injection of 13-cis retinoic acid. With daily injections of retinoic acid, the ceruloplasmin activity continued to increase for at least 4 d. After 4 d, the activity was four times control levels. In copper-deficient rats, the ceruloplasmin activity did not increase in response to retinoic acid unless copper was also given to these rats 8 h after retinoic acid. Actinomycin D blocked the retinoic acid-induced stimulation of ceruloplasmin activity in copper-sufficient rats, but in copper-deficient rats only about half of the increase was blocked when the rats were given copper or copper and retinoic acid. By use of pulse-labeling techniques, ceruloplasmin synthesis was shown to increase 1.5-fold after retinoic acid and this increase was blocked by actinomycin D. When vitamin A-deficient rats were repleted with 13-cis retinoic acid for 3 or 5 d, both the ceruloplasmin activity and synthesis were significantly stimulated when compared to the nonrepleted, deficient rats. Therefore, the dietary components, copper and vitamin A, play an important role in the regulation of plasma ceruloplasmin levels.

Topics: Animals; Ceruloplasmin; Copper; Dactinomycin; Kinetics; Male; Rats; Tretinoin; Vitamin A Deficiency

Earlier work from our laboratory had shown that vitamin A-deficient rats had increased levels of fibronectin in their serum. To explain this result, we investigated the mechanism whereby vitamin A deficiency affects the production of fibronectin by liver and hepatocytes, since liver is the known source of plasma fibronectin. By use of cDNA specific for rat liver fibronectin, we showed that livers of vitamin A-deficient rats had a 2-4-fold increase in the level of fibronectin mRNA and also a higher transcription rate. Rate of synthesis and secretion of fibronectin was found to be increased 2-fold in primary cultures of hepatocytes of deficient animals. Exogenous addition of retinyl acetate or retinoic acid to the media reversed this increase to control levels. The increase was paralleled by an increase in fibronectin mRNA, also reversed by exogenous retinoic acid. At least 12 h were needed for this reversal to take place. Thus, vitamin A appears to regulate the synthesis of fibronectin through its action on fibronectin mRNA transcription. This represents the first reported observation of an action of vitamin A at the genomic level on the synthesis of a specific protein in liver.

Topics: Animals; Cells, Cultured; Diterpenes; Fibronectins; Gene Expression Regulation; Kinetics; Liver; Male; Rats; Retinyl Esters; RNA, Messenger; Transcription, Genetic; Tretinoin; Vitamin A; Vitamin A Deficiency

We have examined RNA synthesis by nuclei isolated from testes of rats of varying vitamin A status. Nuclei from retinol-deficient animals showed substantially decreased RNA synthesis by polymerase II when compared to nuclei from normal animals. Within 4 hours after oral administration of retinyl acetate (as the source of retinol) to deficient animals, RNA synthesis by polymerase II had significantly increased. Administration of retinoic acid had a similar but lesser effect. Nucleoside analysis after alkaline hydrolysis of the RNA synthesized by the endogenous polymerase II suggested that the increased activity was due to a greater number of actively transcribing polymerase II molecules on the DNA. Further, when the template capacity of testicular chromatin isolated from deficient and retinyl acetate refed animals was compared, the number of sites recognized by E. coli RNA polymerase was increased twofold after retinyl acetate administration. We conclude that these retinol-induced changes in transcription are due at least in part to changes in chromatin structure.

Topics: Animals; Chromatin; Diterpenes; Male; Rats; Retinyl Esters; RNA; RNA Polymerase II; Testis; Transcription, Genetic; Tretinoin; Vitamin A; Vitamin A Deficiency

Cellular retinoic acid-binding protein (CRABP), a potential mediator of retinoic acid action, enables retinoic acid to bind in a specific manner to nuclei and chromatin isolated from testes of control and vitamin A-deficient rats. The binding of retinoic acid was followed after complexing [3H]retinoic acid with CRABP purified from rat testes. The binding was specific, saturable, and temperature dependent. If CRABP charged with nonlabeled retinoic acid was included in the incubation, binding of radioactivity was diminished, whereas inclusion of free retinoic acid, or the complex of retinol with cellular retinol binding protein (CRBP) or serum retinol binding protein had no effect. Approximately 4.0 X 10(4) specific binding sites for retinoic acid were detected per nucleus from deficient animals. The number of binding sites observed was influenced by vitamin A status. Refeeding vitamin A-deficient rats (4 h) with retinoic acid lowered the amount of detectable binding sites in the nucleus. CRABP itself did not remain bound to these sites, indicating a transfer of retinoic acid from its complex with CRABP to the nuclear sites. Further, CRBP, the putative mediator of retinol action, was found to enable retinol to be bound to testicular nuclei, in an interaction similar to the binding of retinol to liver nuclei described previously.

Topics: Animals; Binding, Competitive; Biological Transport; Carrier Proteins; Cell Nucleus; Chromatin; Male; Protein Binding; Rats; Receptors, Retinoic Acid; Temperature; Testis; Time Factors; Tretinoin; Vitamin A; Vitamin A Deficiency

A study was conducted to explore the effects of retinoic acid, fed to retinol-deficient rats, on the tissue distribution and levels of cellular retinol-binding protein (CRBP) and cellular retinoic acid-binding protein (CRABP). Sensitive and specific radioimmunoassays were employed to measure the levels of both CRBP and CRABP. Two groups of six male rats each were fed a purified retinoid-deficient diet supplemented with either: i) retinyl acetate (control group); or ii) retinoic acid (30 mg/kg diet) (retinol deficient-retinoic acid group). The retinoic acid supplementation was begun after 38 days on the retinoid-deficient diet alone, and was continued for 52-54 days. Analysis of the data indicated that only the CRBP level of the proximal epididymis in the retinol-deficient/retinoic acid group differed significantly from (was lower than) the corresponding control level, at the 1% confidence level. CRABP tissue levels did not differ significantly between the two groups. Thus, a moderately large intake of retinoic acid, as the only source of retinoids, had very little effect on the tissue distribution or levels of either its own cellular binding protein (CRABP) or of CRBP. This study provides further information showing that the tissue levels of the cellular retinoid-binding proteins are highly regulated and maintained in rats, even in the presence of marked changes in retinoid nutritional status.

Topics: Animals; Carrier Proteins; Genitalia, Male; Liver; Male; Rats; Rats, Inbred Strains; Receptors, Retinoic Acid; Retinol-Binding Proteins; Retinol-Binding Proteins, Cellular; Tissue Distribution; Tretinoin; Vitamin A Deficiency

All-trans-retinoic acid is metabolized in vitro to a biologically active metabolite, retinoyl-beta-glucuronide. We have studied the synthesis of this metabolite in vitro. The identity of the product was established by cochromatography on reverse-phase high-performance liquid chromatography, beta-D-glucuronidase hydrolysis, and fast atom bombardment and collisionally activated decomposition/fast atom bombardment mass spectrometry. The formation of retinoyl-beta-glucuronide is catalyzed by a UDP-glucuronosyltransferase with apparent Km's of 54.7 microM for all-trans-retinoic acid and 2.4 mM for UDP-glucuronic acid. The reaction requires enzyme, UDP-glucuronate, and no other factor. It is strongly inhibited by millimolar concentrations of coenzyme A. The specific activity of UDP-glucuronosyltransferase is greatest in the liver and least in the kidney of those tissues examined. The specific activity of the enzyme is increased by vitamin A deficiency. The increased specific activity observed in the vitamin A-deficient rat liver is uncharacteristic of retinoic acid inactivation enzymes; therefore, retinoyl-beta-glucuronide may be of functional importance.

Topics: Animals; Chromatography, High Pressure Liquid; Coenzymes; Female; Glucuronosyltransferase; Hydrogen-Ion Concentration; In Vitro Techniques; Kinetics; Male; Microsomes, Liver; Rats; Temperature; Tretinoin; Vitamin A Deficiency

We have examined the effect of in vivo vitamin A status on subsequent rat third molar formation and mineralization in an in vitro organ culture system. Vitamin A deficiency imposed during an eight-day in vitro period caused effects very similar to those of vitamin A deficiency imposed on rats in vivo. Analysis of the data also demonstrates that retinoic acid is capable of reversing the interference in mineralization of third molars induced by vitamin A deficiency in the organ culture system.

Topics: Animals; Calcium; In Vitro Techniques; Molar, Third; Odontogenesis; Phosphorus; Proteins; Rats; Tretinoin; Vitamin A; Vitamin A Deficiency

The temporal effects of retinoic acid supplementation on hepatic cytochrome P-450-dependent enzymes were studied on the rat. Four groups of male weanling rats were fed semi synthetic diets: two groups containing 0 or 4.4 mg retinol equivalents per kg diet as retinyl palmitate (A- RA- and A+ RA- groups) and two similar groups supplemented with all trans retinoic acid (12 mg/kg diet) (A- RA+ and A+ RA+ groups). After five or ten weeks of feeding, the rats were killed, liver microsomes were prepared and assayed for aniline hydroxylase, aminopyrine N demethylase activities and cytochrome P-450 levels. Whereas no change was observed between the four groups after 5 weeks, the following modifications appeared after 10 weeks: Vitamin A deficiency decreased hepatic drug metabolism by phase I enzymes (hydroxylase and N demethylase) but only when liver storage pool was not detectable. Vitamin A concentration as low as 4 micrograms/g is sufficient to avoid any perturbation of these enzymes. Parallel to a sparing effect on liver reserves of vitamin A, retinoic acid maintained a normal activity of enzymes of xenobiotic metabolism. However, retinoic acid treatment produced an alteration of phase I enzymes in vitamin A supplemented group (A+ RA+). As this was accompanied by a doubling of vitamin A liver reserves, compared to A+ RA- group, it is suggested that this might result from a liver vitamin A overloading, leading to membrane damage perturbing microsomal enzymes. These results indicate the need for a more careful use of retinoids as a therapeutic agent.

Topics: Aminopyrine N-Demethylase; Aniline Hydroxylase; Animals; Cytochrome P-450 Enzyme System; Diterpenes; Liver; Male; Rats; Rats, Inbred Strains; Retinyl Esters; Tretinoin; Vitamin A; Vitamin A Deficiency

There is reported the first four members of heteroarotinoids, the names of which are ethyl (E)-p-[2-(4,4-dimethylthiochroman-6-yl)propenyl]benzoate (1b), ethyl (E)-p-[2-(4,4-dimethylchroman-6-yl)propenyl]benzoate (1c), ethyl (E)-p-[2-(4,4-dimethyl-1-oxothiochroman-6-yl)propenyl]benzoate (1d), and (E)-p-[2-(4,4-dimethylchroman-6-yl)propenyl]benzoic acid (1e). IR, 1H NMR and 13C NMR data have been recorded for each compound and support the structural assignments. To provide a firm basis for comparison purposes of future analogues, an X-ray analysis was performed on a single crystal of ethyl (E)-p-[2-(4,4-dimethylthiochroman-6-yl)propenyl]benzoate (1b) and a precursor 4,4-dimethylthiochroman-6-yl methyl ketone 1,1-dioxide (18). These data for the heteroarotinoid 1b revealed that the two aryl ring systems were nearly perpendicular in each of the two molecules present in the unit cell (86.37 degrees and 84.17 degrees, respectively). The space group for both molecules was P1 in triclinic systems. Unit cell dimensions (at 15 degrees C) are as follows: for 1b, a = 20.568 (6) A, b = 14.760 (3) A, c = 7.679 (2) A, alpha = 113.33 (2) degrees, beta = 79.45 (2) degrees, gamma = 79.98 (2) degrees, Z = 4; for 18, a = 9.292 (5) A, b = 9.291 (5) A, c = 7.951 (3) A, alpha = 102.16 (3) degrees, beta = 77.49 (3) degrees, gamma = 79.60 (4) degrees, Z = 2. The sulfur-containing ring is in a distorted half-chair in 1b and the methyl carbon C(12) is shown to be trans to H(13) at the C(11)-C(13) bond. The biological activity of these arotinoids was determined in the tracheal organ culture assay and compared with trans-retinoic acid for ability to reverse keratinization in vitamin A deficient hamsters. The ester 1b displayed activity about one-half log unit less than that of the reference while 1c and 1e had activity nearly one log until less than trans-retinoic acid. The sulfoxide was the least active of the heteroretinoids.

Topics: Animals; Benzopyrans; Chromans; Cricetinae; Keratins; Magnetic Resonance Spectroscopy; Models, Molecular; Organ Culture Techniques; Trachea; Vitamin A Deficiency; X-Ray Diffraction

A large body of evidence suggests that retinoids modulate the phenotypic expression of epithelial cells of skin and mucous membranes. The purpose of this study was to investigate the effect of retinoic acid on keratinocyte-mediated collagen breakdown. Keratinocytes derived from the ventral surfaces of the tongues of 4-6 week-old male Wistar rats were established in culture under conditions which are restrictive to growth of fibroblasts, and they were eventually cloned by limiting dilution. The cells were seeded (100,000 cells/cm2) in dishes coated with 3H-labeled, reconstituted type I collagen fibrils and incubated in serum-free medium over a 3-5 day period. Dissolution of the collagen fibrils was monitored by the release of radioactivity to the culture medium. Unstimulated cells metabolized the collagen rather slowly, but addition of retinoic acid in concentrations from 10(-6)M to 10(-8)M resulted in marked acceleration of the degradative process, with complete solubilization of the collagen fibrils in four or five days. The effect of 10(-6)M retinoic acid was of the same order of magnitude as that obtained by addition of a proteolytic activating system either in the form of plasmin or of plasminogen, which is converted to catalytic plasmin by endogenous activators. The effects of retinoic acid and plasminogen/plasmin, however, were not additive. Keratinocytes rendered vitamin A-deficient by cultivation in sera from deficient rats were clearly less effective in degrading the collagen substrate than were "sufficient" cells. Addition of retinoic acid (10(-7)M) enhanced collagen breakdown in both sets of cultures and partially restored the collagenolytic activity of the deficient cells.

Topics: Animals; Cells, Cultured; Collagen; Epithelial Cells; Epithelium; Fibrinolysin; Male; Plasminogen; Rats; Rats, Inbred Strains; Solubility; Tongue; Tretinoin; Tritium; Vitamin A Deficiency

Two synthetic retinoids were examined for their ability to support growth in male vitamin A-deficient rats. One of the compounds, (E)-4-[2-(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-naphthalenyl)-1 -propenyl]-benzoic acid (TTNPB), was found to be highly effective; it was 35-fold more active than all-trans-retinoic acid. Thus, the in vivo results were in agreement with the in vitro activity of this compound published by previous investigators, and support the view that this compound may be useful in determining the molecular mechanism of action of the retinoids. Another analog, 4,4-difluororetinoic acid, was only 12% as effective as retinoic acid. However, the possible instability of this compound and the electronegativity of the fluoro groups prohibited conclusions concerning the biological function of metabolic modification on the 4 position of retinoic acid.

Topics: Animals; Benzoates; Body Weight; Growth; Male; Rats; Retinoids; Tretinoin; Vitamin A Deficiency

The metabolism of all-trans-[15-14C]retinoic acid in vitamin A-adequate or vitamin A-deficient rats fed retinoic acid was studied following intratesticular injection. Analysis of testicular metabolites by HPLC at 1, 6, and 24 h demonstrated that the retinoic acid was isomerized to 13-cis-retinoic acid and metabolized to polar metabolites in both groups of rats at all time periods. However, polar metabolites predominated in the testes of the vitamin A-deficient rats. At 24 h the total radioactivity remaining in the testis was much lower in the testes of vitamin A-deficient rats than in normal animals suggesting a faster rate of metabolism of all-trans-retinoic acid in these rats.

Topics: Animals; Chromatography, High Pressure Liquid; Kinetics; Male; Rats; Stereoisomerism; Testis; Tretinoin; Vitamin A Deficiency

The kinetics and metabolism of physiological doses of all-trans-retinoic acid were examined in blood and small intestinal mucosa of vitamin A-depleted rats. A major portion of intrajugularly injected retinoic acid is rapidly (within 2 min) sequestered by tissues; subsequently 13-cis-retinoic acid and polar metabolites are released into circulation. All-trans-retinoic acid appears in small intestinal epithelium within 2 min after dosing and is the major radioactive compound there for at least 2 h. Retinoyl glucuronide and 13-cis-retinoic acid are early metabolites of all-trans-retinoic acid in the small intestine of bile duct-cannulated rats. Retinoyl glucuronide, the major metabolite of retinoic acid intestinal epithelium, in contrast to other polar metabolites, was not detected in circulation. An examination of [3H]retinyl acetate metabolites under steady state conditions in vitamin A-repleted rats demonstrates the occurrence of all-trans-retinoic acid and 13-cis-retinoic acid in circulation and in intestinal epithelium, in a pattern similar to that found after injection of retinoic acid into vitamin A-depleted rats. Our data establish that all-trans-retinoic acid, 13-cis-retinoic acid, and retinoyl glucuronide are physiological metabolites of vitamin A in target tissues, and therefore are important candidates as mediators of the biological effect of the vitamin.

Topics: Animals; Chromatography, High Pressure Liquid; Diterpenes; Intestinal Mucosa; Intestine, Small; Isomerism; Kinetics; Male; Rats; Rats, Inbred Strains; Retinoids; Retinyl Esters; Structure-Activity Relationship; Tretinoin; Tritium; Vitamin A; Vitamin A Deficiency

Corneas of normal and vitamin A-deficient rabbits were treated topically with [11, 12-3H] retinol or [11, 12-3H] all-trans retinoic acid. Methanol extracts of these corneas were analyzed by high pressure liquid chromatography. Radiolabeled compounds were extracted from the corneas which co-migrated chromatographically with known retinoid standards. In agreement with studies on other tissues and organs, retinol was metabolized to retinoic acid and more polar compounds by corneas of normal and vitamin A-deficient rabbits. All-trans retinoic acid was isomerized to 13-cis retinoic acid in normal rabbit corneas; however, this trans-cis isomerization did not occur in vitamin A-deficient, xerophthalmic corneas.

Topics: Animals; Chromatography, High Pressure Liquid; Cornea; Isomerism; Rabbits; Tretinoin; Vitamin A; Vitamin A Deficiency; Xerophthalmia

Topics: Animals; Antibody-Producing Cells; Diterpenes; Leukocyte Count; Lymphocyte Activation; Lymphocytes; Male; Rats; Retinyl Esters; Tretinoin; Vitamin A; Vitamin A Deficiency

In order to study the metabolism of 13-cis-retinoic acid (13-cis-RA) in animal sebaceous glands and analogues, preputial glands from normal and vitamin A-deficient male rats were incubated with [3H]13-cis-RA for up to 24 h; vitamin A-normal hamster costovertebral glands (flank organs) were incubated for 24 h as well. High-performance liquid chromatography was used to identify the metabolites. [3H]13-cis-RA was rapidly converted to a less polar compound, [3H]all-trans-retinoic acid, by the preputial glands from both normal and deficient rats. In normal preputial glands, the level of [3H]all-trans-RA decreases and two more polar compounds, metabolite I and [3H]4-keto-13-cis-RA appear. In contrast, [3H]all-trans-RA is not metabolized further by the preputial glands from deficient rats, while [3H]13-cis-RA in the hamster costovertebral glands remains intact for up to 24 h. The major metabolite of [3H]13-cis-RA in rat preputial glands is [3H]4-keto-13-cis-RA. Initially, [3H]13-cis-RA is converted to [3H]all-trans-RA. In vitamin A-deficient rats the preputial glands fail to further metabolize [3H]13-cis-RA to the more polar [3H]13-cis-RA derivatives. This may be due to the reduced level of P-450 enzyme in vitamin A-deficient rat preputial glands.

Topics: Animals; Cricetinae; Male; Rats; Rats, Inbred Strains; Sebaceous Glands; Stereoisomerism; Tretinoin; Vitamin A Deficiency

A series of conformationally restricted retinoids was synthesized and screened in two assays used to measure the ability of retinoids to control cell differentiation, namely, the reversal of keratinization in tracheal organ culture from vitamin A deficient hamsters and the inhibition of the induction of mouse epidermal ornithine decarboxylase by a tumor promoter. These compounds had bonds corresponding to selected bonds of the E-tetraene chain of retinoic acid (1) held in a planar cisoid conformation by inclusion in an aromatic ring. The meta-substituted analogue 3 of 4-[(E)-2-methyl-4-(2,6,6-trimethylcyclohexenyl)-1,3-butadienyl+ ++]benzoic acid (2) was far less active than 2 in both assays. In contrast, the vinyl homologue of 2 (4) and the 7,8-dihydro and 7,8-methano analogues (5 and 6) had activity comparable to that of 2. Analogues of 4-[(E)-2-(1,1,4,4-tetramethyl-1,2,3,4-tetrahydro-6-naphthyl)propenyl] benzoic acid (7) were also screened. Replacement of the tetrahydronaphthalene ring of 7 by a benzonorbornenyl group (9) significantly reduced activity, as did removal of the vinylic methyl group from 9 (10). Replacement of the propenyl group of 9 by a cyclopropane ring (12) also reduced activity. Replacement of the tetrahydronaphthalene ring of 7 by 4,4-dimethyl-3,4-dihydro-2H-1-benzopyran and -benzothiopyran rings (13 and 14) also decreased activity. Inclusion of the 7,9 double bond system of 1 in an aromatic ring (15 and 16) reduced activity, whereas inclusion of the 5,7 double bond system in an aromatic ring enhanced activity (7 and 19). Inclusion of the 11,13 and 9,11,13 double bond systems in aromatic rings (2 and 18) also reduced activity below that of 1. Retinoic acid, 7, 13, 14, and 19 inhibited papilloma tumor formation in mice. Toxicity testing indicated that 7 was more toxic than 1, 13, 14, and 19, 19 was more toxic than 1, and 13 and 14 were less toxic than 1.

Topics: Animals; Cell Differentiation; Chemical Phenomena; Chemistry; Female; Keratins; Mice; Organ Culture Techniques; Ornithine Decarboxylase Inhibitors; Papilloma; Retinoids; Skin; Structure-Activity Relationship; Trachea; Vitamin A Deficiency

Vitamin A is essential for growth and development, reproduction and vision in humans. Chemical modification of vitamin A has yielded compounds showing therapeutic promise in skin and neoplastic diseases. The medicinal use of retinoids (vitamin A and its derivatives) is limited by the toxicity associated with this group of compounds. One retinoid, 13-cis-retinoic acid, has proved to be quite effective in the treatment of severe recalcitrant cystic acne under conditions in which toxicity is manageable.

Topics: Acne Vulgaris; Adolescent; Humans; Hypervitaminosis A; Isotretinoin; Male; Retinoids; Tretinoin; Vitamin A Deficiency

Vitamin A deficiency (A-) is known to cause morphologic changes in tooth structures. However, its effects on glycosaminoglycan (GAG) distribution in dental pulp, and the role of retinoic acid (RA) in altering these effects are not clearly defined. Tissue changes induced by vitamin A deficiency and RA administration were evaluated histologically in incisors of rats fed on one of 3 different diets: a) vitamin A sufficient (A+); b) vitamin A deficient (A-); and c) vitamin A deficient supplemented with retinoic acid (A-/RA). Four weeks after the onset of vitamin A deficiency, all rats were killed and their 4 continuously erupting incisors evaluated histologically. A- rats had altered dentine and pulp with disrupted histodifferentiation of pulpal mesenchymal cells to normal odontoblasts. The frequency of these abnormalities in dentine and pulp was lower in A-/RA rats. The enamel organ was unremarkable in the 4-week deficient period. Using special stains, we noted that pulpal GAG accumulation in A- and A-/RA rats was limited to the lingual area, while in A+ rats, GAG were distributed throughout. These data suggest that vitamin A deficiency affects histodifferentiation of pulpal mesenchymal cells to odontoblasts, as well as GAG distribution in pulp. RA administration reduces the A- changes and therefore, appears to have some activity in dentinogenesis.

Topics: Animals; Cell Differentiation; Dental Pulp; Dentin; Enamel Organ; Glycosaminoglycans; Incisor; Male; Rats; Rats, Inbred Strains; Time Factors; Tretinoin; Vitamin A Deficiency; Weaning

The seminiferous tubules of rat testes contain a protease and heat sensitive factor capable of stimulating (3H)-thymidine incorporation into quiescent cultures of NIH 3T3 mouse fibroblast cells. This mitogenic factor activity was only 4% in the testes of vitamin A deficient-retinoic acid maintained rats as compared to that of normal rats. Supplementation of retinyl acetate to these vitamin A deficient rats for 4 days resulted in a 56% recovery in the mitogenic factor activity while by day 16, the recovery was 80 percent. Incorporation of (3H)-thymidine into DNA of the seminiferous tubules of vitamin A deficient rats was only 46% of that of normal rats which was restored to normal levels by retinyl acetate supplementation for 24 days.

Topics: Animals; Cell Division; Cells, Cultured; DNA Replication; Growth Substances; Kinetics; Male; Mice; Rats; Rats, Inbred Strains; Seminiferous Tubules; Testis; Tretinoin; Vitamin A Deficiency

Topics: Chemical Phenomena; Chemistry; Humans; Isotretinoin; Neoplasms; Precancerous Conditions; Retinaldehyde; Skin Diseases; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Animals; Blood Vessels; Cell Differentiation; Cell Division; Cell Line; Cell Transformation, Neoplastic; Chick Embryo; Cricetinae; Gene Expression Regulation; Humans; Macromolecular Substances; Mice; Organ Culture Techniques; Phenotype; Rats; Skin; Staining and Labeling; Tretinoin; Vitamin A; Vitamin A Deficiency

Cytokinetic and ultrastructural studies were carried out to elucidate mechanisms involved in the reversal of squamous metaplasia (SM) by beta-retinoic acid in organ cultures of tracheas derived from vitamin A-deficient hamsters. Tracheal cultures exhibiting focal areas of SM were treated with the retinoid for up to 7 days. The retinoid significantly inhibited [3H]-thymidine labeling indices in the basal cells and stimulated the labeling indices in mucous cells. At the ultrastructural level the retinoid induced marked remodeling alterations in the metaplastic epithelium that included: (a) disruption of desmosomes and widening of intercellular spaces; (b) extensive vacuolation and degeneration of the metaplastic cells; (c) extrusion of the degenerated cells; (d) aggregation of keratin filaments; and (e) differentiation of certain basal cells into secretory cells. Consequently most degenerated metaplastic cells were extruded and the epithelium repopulated as a result of differentiation of basal cells into mucous cells and hyperplasia of the pre-existing mucous cells. The degenerative effects of the retinoid were limited to the metaplastic foci since the uninvolved epithelium adjoining metaplastic foci were not significantly altered. The results suggest that the restoration of normal tracheal epithelium following the retinoid treatment of explants exhibiting focal areas of squamous metaplasia is associated with the enhanced proliferation of the mucous cells. The inhibition of proliferation of basal cells further prevented hyperplasia and restored cell replication within the normal range.

Topics: Animals; Cell Differentiation; Cell Division; Cricetinae; Epithelium; Keratins; Metaplasia; Microscopy, Electron; Organ Culture Techniques; Trachea; Tretinoin; Vitamin A Deficiency

Topics: Animals; Follicle Stimulating Hormone; Luteinizing Hormone; Male; Rats; Rats, Inbred Strains; Spermatogenesis; Testis; Testosterone; Tretinoin; Vitamin A; Vitamin A Deficiency

The effects of all-trans retinol and retinoic acid (RA) on the humoral and cell-mediated immune response and on lymphoid organ weights of broiler chicks were examined. Chicks were fed diets supplemented with retinol or RA at 0, 0.2 and 2.0 micrograms/g diet. The diets were fed continuously from day of hatch or after depletion of hepatic vitamin A reserves. Rapid vitamin A deficiency was induced in chicks by initially feeding the diet containing 2.0 micrograms RA/g diet and subsequently feeding diets containing 0 or 0.2 microgram RA/g diet. Serum antibody hemagglutination titers, in response to intravenous injection of human serum albumin (HSA), were not affected by level or chemical form of vitamin A supplemented in diets. In vitamin A-deficient chicks, there was an inverse relationship observed between the number of peripheral blood lymphocytes obtained per milliliter of blood and their ability to proliferate in response to mitogenic stimulation. Growth of the bursa of Fabricius and the thymus was impaired in chicks fed a vitamin A-free diet. A partial deficiency of vitamin A adversely affected relative bursa weight but not that of thymus. In general, RA was inferior to retinol in maintenance of lymphoid tissue.

Topics: Animals; Antibody Formation; Chickens; Immunity, Cellular; Isomerism; Lymphoid Tissue; Male; Organ Size; Tretinoin; Vitamin A; Vitamin A Deficiency

The effects of a vitamin A-deficient diet and the subsequent supplementation with retinoic acid (10 micrograms/g dry diet) on serum thyroid hormones in rats were studied. In vitamin A-deficient animals there are increased levels of thyroxine and triiodothyronine and changes in serum transport of these hormones. Retinoic acid was able to restitute both levels and serum transport of hormones as in control rats.

Topics: Animals; Blood Proteins; Body Weight; Male; Protein Binding; Rats; Rats, Inbred Strains; Serum Albumin; Thyroxine; Tretinoin; Triiodothyronine; Vitamin A Deficiency

Vitamin A-deficient rats were given a single intrajugular injection of 1 mg all-trans-[11-3H]retinoic acid and 3 h later the rats were killed. The small intestines were extracted and chromatographed by high-performance liquid chromatography to yield distinct metabolites. These were quantitated using the assumption that the specific activity of the metabolite is equal to that of the parent [3H]retinoic acid. The biological activity of all discernible metabolites was determined in the vitamin A-deficient female rat by vaginal smear assay. Retinoic acid and retinoyl-beta-glucuronide from the preparation had equal activity while no activity was found for any of the other metabolite fractions. Thus, no evidence for an unknown metabolite having potent epithelial differentiating activity could be found in this target tissue of vitamin A action.

Topics: Animals; Dose-Response Relationship, Drug; Female; Intestinal Mucosa; Rats; Tretinoin; Vaginal Smears; Vitamin A Deficiency

Biliary metabolites from physiological doses of all-trans-[10-3H]retinoic acid were examined in normal and vitamin A-deficient rats. The bile from normal and vitamin A-deficient rats contained approximately 60% of the administered dose following a 24-h collection period. However, vitamin A-deficient rats show a 6-h delay in the excretion of radioactivity compared to normal rats. Retinoyl-beta-glucuronide excretion was particularly sensitive to the vitamin A status of the rats. In normal rats, retinoyl-beta-glucuronide reached a maximum concentration of 235 pmol/ml of bile 2 h following the dose and then rapidly declined. Vitamin A-deficient rats show a relatively constant concentration of this metabolite (100-150 pmol/ml of bile) over a 10-h collection period. Retinoic acid excretion was low in both normal and deficient rats. The concentration of retinotaurine, a recently identified biliary metabolite, was approximately equal to retinoyl-beta-glucuronide in normal rats and appeared in the bile 2 h later than the glucuronide.

Topics: Animals; Bile; Kinetics; Male; Rats; Tretinoin; Vitamin A Deficiency

A study was conducted on the incorporation of [11-3H]retinyl acetate into various retinyl esters in liver tissues of rats either vitamin A-sufficient, vitamin A-deficient or vitamin A-deficient and maintained on retinoic acid. Further, the metabolism of [11-3H]retinyl acetate to polar metabolites in liver tissues of these three groups of animals was investigated. Retinol metabolites were analyzed by high-performance liquid chromatography. In vitamin A-sufficient rat liver, the incorporation of radioactivity into retinyl palmitate and stearate was observed at 0.25 h after the injection of the label. The label was further detected in retinyl laurate, myristate, palmitoleate, linoleate, pentadecanoate and heptadecanoate 3 h after the injection. The specific radioactivities (dpm/nmol) of all retinyl esters increased with time. However, the rate of increase in the specific radioactivity of retinyl laurate was found to be significantly higher (66-fold) than that of retinyl palmitate 24 h after the injection of the label. 7 days after the injection of the label, the specific radioactivity between different retinyl esters were found to be similar, indicating that newly dosed labelled vitamin A had now mixed uniformly with the endogenous pool of vitamin A in the liver. The esterification of labelled retinol was not detected in liver tissues of vitamin A-deficient or retinoic acid-supplemented rats at any of the time point studied. Among the polar metabolites analyzed, the formation of [3H]retinoic acid from [3H]retinyl acetate was found only in vitamin A-deficient rat liver 24 h after the injection of the label. A new polar metabolite of retinol (RM) was detected in liver of the three groups of animals. The formation of 3H-labelled metabolite RM from [3H]retinyl acetate was not detected until 7 days after the injection of the label in the vitamin A-sufficient rat liver, suggesting that metabolite RM could be derived from a more stable pool of vitamin A.

Topics: Animals; Chromatography, High Pressure Liquid; Diterpenes; Female; Kinetics; Liver; Male; Rats; Rats, Inbred Strains; Retinyl Esters; Tretinoin; Tritium; Vitamin A; Vitamin A Deficiency

Topics: Animals; Diterpenes; DNA; Genes; Intestinal Mucosa; Liver; Male; Poly A; Protein Biosynthesis; Rats; Rats, Inbred Strains; Retinyl Esters; RNA; RNA, Messenger; Testis; Transcription, Genetic; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Animals; Carrier Proteins; Centrifugation, Density Gradient; Conjunctiva; Cornea; Cytosol; Rabbits; Receptors, Retinoic Acid; Retinol-Binding Proteins; Tretinoin; Vitamin A; Vitamin A Deficiency

The metabolites of all-trans-[3H]retinoic acid appearing in the intestines of bile duct-cannulated rats were compared to those of similarly treated intact rats. 2.4% of administered radioactivity was found in the small intestines of bile duct-cannulated rats 2 H after dose, while a much larger proportion of the dose (7.2%) was found in the intestines of the intact rats. All-trans- and 3-cis-retinoic acids predominate in the intestinal mucosa of bile duct-cannulated rats shortly after dosing. Retinoyl glucuronide was the major metabolite occurring as a mixture of the all-trans and 13-cis forms. Highly polar metabolites of retinoic acid appear in mucosa at all times in both intact and bile duct-cannulated rats demonstrating rapid metabolism of retinoic acid in intestine. The finding of a similar proportion of the 13-cis isomers in retinoic acids and retinoyl glucuronides suggests that injected all-trans-retinoic acid is isomerized in vivo probably prior to conjugated with glucuronic acid.

Topics: Animals; Bile; Glucuronates; Intestinal Mucosa; Intestine, Small; Isotretinoin; Kinetics; Male; Rats; Tretinoin; Tritium; Vitamin A Deficiency

A retinoic acid epoxidase is present in rat kidney homogenates. It is found in the particulate fraction, and is dependent on ATP, NADPH, and oxygen. It is stimulated by Fe2+ and inhibited by Mn2+, Zn2+, Cu2+, EDTA, and N,N'-diphenyl-para-phenylenediamine. Its properties are closest to those of microsomal lipid peroxidases previously studied. Epoxidase activity is not affected by the rat's vitamin A status, nor is it inducible by retinoic acid. In N,N'-diphenyl-para-phenylenediamine-feeding experiments, it was shown that blocking epoxidation in vivo does not inhibit the function of retinoic acid. It is concluded that retinoic acid epoxidation is not required for retinoic acid function.

Topics: Animals; Antioxidants; Cations, Divalent; Kidney; Kinetics; Magnesium; NADP; Oxidoreductases; Phenylenediamines; Rats; Substrate Specificity; Tretinoin; Vitamin A Deficiency

The mannosyl derivative of phosphorylated vitamin A, mannosylretinylphosphate, is synthesized in vivo and in vitro by most epithelial tissues, including mouse epidermal cells. This compound is but one component of the complex biosynthetic machinery responsible for the assembly of glycoproteins in mammalian membranes; its specific role remains to be understood, even though it has been established that it functions as a carrier of mannose mostly in a direct transfer to protein. The system of five conjugated double bonds appears necessary for this function, since upon saturation the resulting perhydroretinylphosphate is incapable of acting in mannosyl transfer to endogenous microsomal proteins. Using vitamin A - deficient hamsters and rats, retinoic acid was shown to be as active as retinol in restoring the in vivo in corporation of mannose into glycoproteins to normal levels within a short time, in liver and tracheal tissues. Retinoic acid was also active in increasing adhesive properties of spontaneously transformed mouse dermal fibroblasts (3T12 cells) in a reversible manner. The free carboxyl group appeared to be a requirement for this activity inasmuch as the lactonized form of 11-hydroxyretinoic acid was inactive. Even though retinoic acid, as well as retinol, was able to enhance adhesion of 3T12 cells, only the cellular retinoic acid-binding protein was detected, suggesting that the cellular retinol-binding protein is not a requirement for this activity. A metabolite of retinoic acid, compound X, different from retinol, was found incorporated into mannosylretinoidphosphate (MXP) by 3T12 cells. This derivative constitutes up to 40% of the total radioactive pool of derivatives of retinoic acid after 48 hours of labeling. These data suggest that retinol, as mannosylretinylphosphate, and retinoic acid, as mannosylretinoidphosphate, function as mannosyl carriers in biologic membranes.

Topics: Animals; Cell Line; Diterpenes; Dolichol Monophosphate Mannose; Epithelium; Fibroblasts; Hexosephosphates; Mannose; Mannosephosphates; Mice; Mice, Inbred BALB C; Microsomes, Liver; Polyisoprenyl Phosphate Monosaccharides; Polyisoprenyl Phosphate Sugars; Rats; Skin; Tretinoin; Vitamin A; Vitamin A Deficiency

The major metabolite in the small intestinal mucosa of vitamin A deficient rats dosed intrajugularly with 5,6-epoxy[3H]-retinoic acid has been identified as 5,6-epoxyretinoyl beta-glucuronide. The assignment was based on the metabolite's chemical, spectral, and chromatographic properties. Incubation of the metabolite with beta-glucuronidase released 5,6-epoxyretinoic acid. Incubation of 5,6-epoxyretinoic acid with rat liver microsomes in the presence of uridine-5'-diphospho-1 alpha-D-glucuronic acid produced the metabolite. 5,6-Epoxy[3H]retinoyl beta-glucuronide weas observed in the liver, small intestinal mucosa, and intestinal contents but not in kidney of vitamin a deficient rats. Its concentration was greatly diminished in liver and small intestinal mucosa, and it was not observed in kidney of vitamin A deficient rats dosed orally with retinoic acid for several days before administration of 5,6-epoxy[3H]retinoic acid. Generally, oral retinoic acid treatment accelerated 5,6-epoxyretinoic acid metabolism and enhanced accumulation of highly polar metabolites. Moreover, 5,6-epoxyretinoic acid metabolism was more rapid than that of retinoic acid and did not result in production of retinoic acid.

Topics: Animals; Chromatography, High Pressure Liquid; Intestinal Mucosa; Intestine, Small; Male; Microsomes, Liver; Rats; Rats, Inbred Strains; Tretinoin; Vitamin A Deficiency

A method is described for measuring the relative activities of vitamin A active substances based on their direct effect on the cornified vaginal epithelium of vitamin A-deficient rats. The results obtained agree well with those found in the tracheal organ culture assay. The relative activities found for several test compounds were: all-trans-retinoyl-beta-glucuronide greater than all-trans-retinoic acid greater than all-trans retinol greater than all-trans-5,6-epoxyretinoic acid. The assay is simple and inexpensive to perform, and should find use in laboratories where the equipment and personnel required for the tracheal organ culture assay are not available.

Topics: Animals; Biological Assay; Castration; Dose-Response Relationship, Drug; Epithelium; Female; Glucuronates; Rats; Tretinoin; Vaginal Diseases; Vaginal Smears; Vitamin A; Vitamin A Deficiency

The ability of the biliary metabolite retinotaurine to reverse the cornification of vaginal epithelial cells induced by vitamin A deficiency was assessed using a vaginal smear assay. Retinotaurine activity was examined following the intravaginal administration of 10(-12) mol to 10(-8) mol of this compound per vagina. This metabolite exhibited no detectable activity at any dose tested. These results show that retinotaurine cannot be more than 1% as active as all-trans-retinoic acid since retinoic acid shows a response at concentrations of 10(-10) mol per vagina. The low biological activity of this recently identified biliary metabolite suggests that it represents an excretory form of retinoic acid.

Topics: Animals; Biological Assay; Castration; Cell Differentiation; Dose-Response Relationship, Drug; Epithelium; Female; Rats; Tretinoin; Vaginal Diseases; Vaginal Smears; Vitamin A Deficiency

Sample preparation and high-pressure liquid-chromatography separation methods useful for the study of retinoic acid metabolism are reported. The sample preparation procedure does not cause significant degradation of retinoic acid, and the gradient high-pressure liquid-chromatography separation method gives excellent separation of the major metabolites of retinoic acid. These methods were used to examine the metabolites of retinoic acid in blood, trachea and lung, testes, kidneys and small intestine of vitamin A-deficient rats dosed subcutaneously with 2 micrograms of [11,12-3H] retinoic acid. At 6h after dosing, a total of eight metabolites of retinoic acid produced in vivo were found in the tissues examined. Of these, four were found in most of the epithelial tissues examined, and therefore may be of interest as possible active metabolites in the epithelial functions of vitamin A.

Topics: Animals; Chromatography, High Pressure Liquid; Epithelium; Intestine, Small; Kidney; Lung; Male; Rats; Testis; Tissue Distribution; Trachea; Tretinoin; Vitamin A Deficiency

In vitro perfusion of corneas of normal and vitamin A-deficient rabbits provided a model in which to study the pharmacokinetics of corneal permeability and uptake of retinoic acid and retinol. The permeability coefficients of retinoic acid and retinol were 1.49 x 10(-5) and 0.61 x 10(-5) cm/s, respectively. Removal of the corneal epithelium did not affect the permeability of these lipid-soluble retinoids; however, diffusion through xerophthalmic, vitamin A-deficient corneas was significantly reduced. The corneal uptake of retinoic acid and retinol was reduced by 50% on removal of the epithelium, was nonspecific, and was not affected by xerophthalmia. High-performance liquid chromatography indicated that these retinoids were not metabolized during diffusion through the cornea. These results show that topical application of retinoids is a rational approach to the treatment of such corneal diseases as xerophthalmia and epithelial defects.

Topics: Animals; Cornea; Epithelium; In Vitro Techniques; Permeability; Rabbits; Tretinoin; Vitamin A; Vitamin A Deficiency; Xerophthalmia

The metabolism and bioactivity of N-(4-hydroxyphenyl)-all-trans-retinamide (HPR) and of various O-alkyl and ester derivatives of HPR were investigated in rodents. The principal metabolite of HPR in tissues is N-(4-methoxyphenyl)-all-trans-retinamide. N-(4-methoxyphenyl)all-trans-retinamide is equipotent to HPR in reversing keratinization of retinoid-deficient hamster trachea in vitro. Another nonpolar metabolite of HPR is also present in tissue and (although not positively identified) is thought to be a long-chain fatty acid ester of HPR. HPR is excreted into rat bile as numerous polar retinamides, including HPR-O-glucuronide. The rate of hydrolysis of HPR esters by rat serum and hepatic enzymes in vitro is inversely related to the length of the esterified acid side group. After a 30-min incubation at 37 degrees C in serum, percentage of hydrolysis for acetyloxy, propionyloxy, butyryloxy, pivaloyloxy and octanoyloxy esters of HPR is 41, 20, 7.5, 1.9 and 1.5, respectively. In contrast, hydrolysis by hepatic esterases is more rapid, particularly for the pivaloyloxy ester. Potency of the various HPR esters in the tracheal organ culture bioassay decreases as the length of the esterified side group increases; the acetyloxy ester is at least 5 times more potent than the octanoyloxy ester.

Topics: Animals; Biotransformation; Cricetinae; Fenretinide; Hydrolysis; In Vitro Techniques; Keratins; Liver; Male; Rats; Rats, Inbred Strains; Trachea; Tretinoin; Vitamin A Deficiency

Topics: Butyrates; Butyric Acid; Carcinoma; China; Croton Oil; Herpesvirus 4, Human; Humans; Nasopharyngeal Neoplasms; Plant Extracts; Plants, Medicinal; Stimulation, Chemical; Tretinoin; Virus Activation; Vitamin A Deficiency

The prevalence of goblet cells in duodenal crypts decreased to 60% of normal at day 4 of deficiency. Villus and crypt length, total mucosal cell number, cell cycle time, and migration rate were unaffected at day 8 of deficiency. The carbohydrate content, protein content, and molecular size of high molecular weight glycoproteins (peak I) were unaffected at day 8 of deficiency. The incorporation of L-[3H]threonine (G) into proteins of all peaks were reduced to 60% of normal at day 8 of deficiency. The incorporation of L-[1-14C]fucose into glycoprotein fell to 25% of normal in peak I, and to 70 to 90% of normal in other peaks at day 8 of deficiency. In vitamin A deficiency, a defect seems to exist either in the rate of differentiation of intestinal goblet cells at a critical period, or in the differentiation of a subclass of vitamin A-sensitive goblet cells. Some possible explanations for the decreased synthetic rate of some glycoproteins in the presence of normal steady-state concentrations in A- rats are discussed.

Topics: Animals; Cell Differentiation; Duodenum; Fucose; Glycoproteins; Intestinal Mucosa; Kinetics; Protein Biosynthesis; Rats; Threonine; Tretinoin; Vitamin A Deficiency

A description of the enzyme that produces 5,6-epoxyretinoic acid from all-trans-retinoic acid has been presented. This enzyme system is found in highest concentrations in the kidney followed by intestine, liver and spleen. The enzyme requires molecular oxygen, magnesium ions, ATP, and NADPH. In the kidney, it is found in the mitochondrial and microsomal fractions and has a Michaelis constant of 3.2 X 10(-6) M and 3.7 X 10(-6) M for 13-cis and all-trans-retinoic acid, respectively. The resultant product, 5,6-epoxyretinoic acid, has minimal activity in supporting growth of vitamin A-deficient rats, its activity estimated to be 0.5% that of retinoic acid. An investigation of the biliary excretion products of tritiated retinoic acid has revealed several unknown metabolites. A glucuronidase sensitive metabolite from these products has been isolated and identified as retinoyl-beta-glucuronide by ultraviolet absorption spectrometry and mass spectrometry. The retinoyl-beta-glucuronide originally discovered by Olson and collaborators accounts for only 12% of the total excreted biliary products of retinoic acid. At least four to six major unknown retinoic acid metabolites, in addition to retinoyl-beta-glucuronide, have been detected and will shortly be identified.

Topics: Animals; Bile; Chromatography, High Pressure Liquid; Epoxy Compounds; Glucuronates; Kidney; Rats; Tretinoin; Vitamin A Deficiency

Topics: Adipose Tissue; Animals; Chickens; Female; Kinetics; Liver; Male; Rats; Tretinoin; Vitamin A; Vitamin A Deficiency; Vitamin E

Topics: Animals; Humans; Neoplasms; Neoplasms, Experimental; Organ Culture Techniques; Papilloma; Rats; Skin Neoplasms; Stomach Neoplasms; Structure-Activity Relationship; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Animals; Biology; History, 20th Century; Massachusetts; Night Blindness; Rats; Retina; Tretinoin; United States; Vitamin A; Vitamin A Deficiency

Animals maintained on retinol (vitamin A-alcohol)-deficient diets exhibit testicular atrophy and loss of the germinal epithelium. Retinoic acid (vitamin A-acid), when fed to retinol-deficient animals, does not prevent these lesions and had thus been thought not to play a role in the tests. Serum testosterone (T) levels, determined by RIA, in retinol-deficient rats were determined to be significantly lower than in control rats. In contrast, retinoic acid-fed, retinol-deficient rats exhibited serum T concentrations similar to those of control rats. No difference in immunoreactive serum LH levels was observed in the three groups. The response of serum T to ip administration of LH in retinol-deficient animals relative to basal levels was similar to that observed in control as well as retinoic acid-fed, retinol-deficient rats. These results show that while basal T production in retinol-depleted rats is decreased, LH-stimulated T synthesis is unaffected. Furthermore, retinoic acid, in the absence of retinol, can support T production, suggesting that contrary to present dogma, retinoic acid plays a role in testis.

Topics: Animals; Luteinizing Hormone; Male; Organ Size; Rats; Testis; Testosterone; Tretinoin; Vitamin A Deficiency

Metabolites formed in vivo from [3H]retinoic acid dosed intrajugularly to vitamin A-deficient rats and to vitamin A-deficient rats supplemented orally with unlabeled retinoic acid were investigated. Extracts of liver, small intestinal mucosa, kidney, testes and serum were separated into charged uncharged fractions by DEAE-Sephadex. This allowed the direct application of 20-40% of the combined charged extracts from up to six organs to be loaded onto a high-performance liquid chromatography column. The large aliquot size plus the use of relatively high specific activity [3H]retinoic acid resulted in detection of nanomolar metabolite quantities. The substantial resolution achieved with the high-performance liquid chromatography gradient system aided in demonstrating the complexity, extent and variations of retinoic acid metabolism in vivo, The metabolic profiles changed with retinoic acid pretreatment, time after dose and tissue source. Some 3H-labeled metabolites were predominant in vitamin A-deficient animals; others appeared to be predominant in the retinoic acid-supplemented animals. The gross effect of retinoic acid supplementation was to both accelerate retinoic acid metabolism and cause an accumulation of more polar metabolites. It appears that retinoic acid metabolism in vivo is a complex process that occurs through multiple metabolites, which are, at least partially, tissue-specific.

Topics: Animals; Chromatography, High Pressure Liquid; Intestinal Mucosa; Intestine, Small; Kidney; Liver; Male; Rats; Rats, Inbred Strains; Testis; Tissue Distribution; Tretinoin; Vitamin A Deficiency

Topics: Animals; Cornea; Epithelium; Female; Germ-Free Life; Male; Microscopy, Electron, Scanning; Neovascularization, Pathologic; Rats; Tretinoin; Vitamin A Deficiency

The combined administration of follicle-stimulating hormone and testosterone to sexually mature male rats whose testes were in the early stages of damage from retinol deficiency did not prevent the continued degeneration of the germinal epithelium. It is concluded that the action of retinol on the testis is probably not mediated through, nor influenced by, the pituitary or steroid hormones.

Topics: Animals; Follicle Stimulating Hormone; Male; Rats; Spermatozoa; Testicular Diseases; Testis; Testosterone; Tretinoin; Vitamin A Deficiency

Experiments were conducted to evaluate critically the role of vitamin A in the maintenance and functional integrity of mucus-secreting goblet cells in rat small intestine. Essentially synchronous vitamin A deficiency was induced by the withdrawal of retinoic acid from mature, stringently-deficient male rats reared by feeding vitamin A-depleted weanlings diets first supplemented with and then lacking in 2 micrograms retinoic acid per gram diet in repeating 18 day:10 day cycles. Secondary inanition was minimized by force-feeding both deficient and control animals twice daily. Whereas the prevalence of oligomucus cells was unchanged, the number of goblet cells per duodenal crypt gland decreased abruptly to 60% of control values starting 2 to 3 days after the withdrawal of retinoic acid and then stabilized. The responses of mucus-secreting cells to atropine and pilocarpine were identical in vitamin A deficient and control animals. As studied with [3H]thymidine, the rate of division of epithelial cells and the migration rate of columnar and goblet cells out of the crypt gland and along the villus were also unaffected by vitamin A deficiency. We conclude that two populations of goblet cells exist in the intestine--one relatively insensitive and the other sensitive to vitamin A status. In vitamin A deficiency, the rate of differentiation of sensitive goblet cells from oligomucus cells and other precursor cells seems to be blocked.

Topics: Animals; Duodenum; Jejunum; Male; Mitosis; Pilocarpine; Rats; Tretinoin; Vitamin A Deficiency

Topics: Animals; Carbon Monoxide; Cricetinae; Kinetics; Male; Mesocricetus; Microsomes, Liver; NAD; NADP; Oxidation-Reduction; Tretinoin; Vitamin A Deficiency

Topics: Animals; Chromatography, High Pressure Liquid; Drug Stability; Rats; Retinaldehyde; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Animals; Carcinoma, Squamous Cell; Humans; Lipoproteins, VLDL; Rats; Skin Diseases; Tretinoin; Triglycerides; Vitamin A; Vitamin A Deficiency

5,6-Epoxyretinoic acid was detected in small intestine, kidney, liver, testes and serum of vitamin A-deficient rats 3 h after a single physiological dose of [3H]retinoic acid. The maximum concentration of 5,6-epoxide in intestinal mucosa was observed 3 h after intrajugular administration of retinoic acid. However, at 7 h post administration, no 5,6-epoxyretinoic acid was detected in mucosa, demonstrating the rapid intestinal metabolism or excretion of this metabolite. No 5,6-epoxy[3H]retinoic acid was detected in mucosa, liver or serum of retinoic acid-repleted rats 3 h after administration of 2 micrograms of [3H]retinoic acid.

Topics: Animals; Chromatography, High Pressure Liquid; Epoxy Compounds; Kinetics; Male; Rats; Tissue Distribution; Tretinoin; Vitamin A Deficiency

The biosynthesis of corneal epithelial proteins and glycoconjugates and their response to vitamin A were measured in vitamin A-deficient and normal rats. The synthesis of specific high-molecular-weight epithelial glycoconjugates was directly related to the levels of vitamin A administered. Protein synthesis, however, remained unaltered. Since vitamin A is known to reverse the keratinization process, the vitamin A-regulated high-molecular-weight glycoconjugates may play a role in modulating the expression of the keratinizing phenotype.

Topics: Animals; Autoradiography; Cornea; Electrophoresis, Polyacrylamide Gel; Epithelium; Glycoproteins; Molecular Weight; Rats; Scintillation Counting; Tretinoin; Vitamin A Deficiency; Xerophthalmia

The functional integrity of the local immune system in vitamin A-deficient (A-) rats was investigated. Secretory IgA levels in the intestinal fluid of A- rats were significantly lower than in controls. This and the decrease in intensity of immunofluorescent staining for secretory component (SC) in the intestinal cells was related to the duration of vitamin A deprivation. IgG levels in the intestinal fluid, and serum IgA and IgG levels were unaffected in deficiency. Moreover, when the response of animals to DNP50-BGG was evaluated, the local anti-DNP response in the intestine was markedly depressed. These defects may result from impaired synthesis of SC by epithelial cells. On the other hand, the serum antibody response in deficient animals was not noticeably different from that of the controls; if any, htere was a slight reduction in the affinity of antibody.

Topics: Animals; Antibody Formation; Immunoglobulin A; Immunoglobulin A, Secretory; Immunoglobulin G; Immunoglobulins; Intestinal Secretions; Intestines; Male; Rats; Tretinoin; Vitamin A Deficiency

Vitamin A and its derivaties (retinoids) are necessary for the maintenance of normal phenotypic expression. An attempt at understanding the biochemical role of vitamin A had led to the demonstration of a new pathway for retinol. In this pathway, vitamin A is phosphorylated to retinylphosphate (RP), which is then glycosylated to retinylphosphatemannose (MRP). These two derivatives have been found in a variety of tissues in vivo and in vitro and appear to be ubiquitous components of cellular membranes. The suggestion has been made that MRP may mediate specific cellular interactions by functioning as a lipid intermediate in the biosynthesis of specific glycoconjugates. A study on spontaneously-transformed mouse fibroblasts (Balb/c 3T12-3 cells) has shown that retinoids are active in increasing the adhesive properties of these cells as measured in an EDTA-mediated detachment assay. Various retinoids were tested for their activity in the adhesion test, and this activity was found to correlate well with their biological activity in maintaining the expression of normal epithelial differentiation in other systems. Retinoic acid, 5,6-epoxyretinol, and 5,6-epoxyretinoic acid were the most active compounds. Retinoids without biological activity in other systems were also inactive in inducihg adhesive properties of 3T12-3 cells. Among these were the synthetic derivatives of retinol, anhydroretinol, and 4,5-monoeneperhydroretinol, and the phenyl derivative of retinoic acid. Beta-Ionone, abscisic acid, and juvenile hormone, which are devoid of vitamin A activity in other systems, were also inactive in this system. Retinoid-induced changes in cell surface proteins were investigated but no difference in 125I-fibronectin (MW 220,000) was detectable between retinoid-treated and untreated cells. However, these cells synthesized retinylphosphatemannose and the incorporation of 2-3H-mannose into a specific glycoprotein (gp 180) was found to be enhanced specifically by retinoid treatment. Investigations of the involvement of gp 180 in adhesion are in progress.

Topics: Animals; Benzopyrenes; Cell Adhesion; Cell Membrane; Cells, Cultured; Cricetinae; Epithelium; Fibroblasts; Glycoproteins; Mice; Structure-Activity Relationship; Trachea; Tretinoin; Vitamin A; Vitamin A Deficiency

The potential of all-trans-5,6-epoxyretinoic acid to promote growth in vitamin A-deficient rats was evaluated over a 25-fold dose range using single daily intraperitoneal injections. 5,6-Epoxyretinoic acid was found to be 0.5% as active as all-trans-retinoic acid. The discrepancies between the present results and those obtained previously by other workers are discussed.

Topics: Administration, Oral; Animals; Dose-Response Relationship, Drug; Epoxy Compounds; Growth; Injections, Subcutaneous; Male; Rats; Tretinoin; Vitamin A Deficiency

When Wistar and Sprague-Dawley rats fed from weaning on a vitamin A and provitamin A deficient diet are supplemented with retinoic acid, they mature into adults whose general physical conditions and weight are comparable with those of retinol supplemented animals. However, the adrenal cortex, testes and ovaries delta 5-3 beta hydroxysteroid deshydrogenases-delta 5-delta 4-3-oxosteroide isomerases are in all cases strongly decreased compared with those of retinol supplemented animals. These results are reproducible only if the mothers' reserves in vitamin A are about 50 gamma/g of hepatic tissue and if the animals are weaned between the 19th and 21st day. These results obtained under these strictly defined conditions demonstrate the importance of the "biochemical lesion" which causes steroidogenesis perturbations induced by the vitamin A deficiency. The experimental data are discussed in the light of the recent findings concerning the direct role of the vitamin A in the transglycosylation reactions (Luigi M. de Luca, Vitamins and Hormones, 1977, 35, 1-57).

Topics: 3-Hydroxysteroid Dehydrogenases; Adrenal Glands; Animals; Female; Male; Organ Size; Ovary; Rats; Steroids; Testis; Tretinoin; Vitamin A; Vitamin A Deficiency

The synethesis of a new retinoid, N-(4-hydroxyphenyl)-all-trans-retinamide, which has useful biological properties, is described. This retinoid was more potent than retinyl acetate in reversing keratinization caused by retinoid deficiency in tracheal organ culture. It was markedly less toxic than retinyl acetate when fed p.o. to rats over 2-week or 6-month periods. It was an effective agent for inhibition of the development of breast cancer induced in rats by N-nitroso-N-methylurea, although it was not as potent as retinyl acetate in this regard. However, the lesser toxicity of 4-hydroxyphenylretinamide makes it a superior agent for prevention of breast cancer. High-pressure liquid chromatographic analyses of liver and breast extracts from rats treated for 6 months with retinoids show the pharmacokinetic basis for the superiority of 4-hydroxyphenylretinamide; this retinoid and its metabolites were found in high concentrations in breast tissue, without any measurable accumulation in the liver or evident liver toxicity. In contrast, chronic feeding of retinyl acetate caused marked deposition of retinyl esters in the liver and severe hepatotoxicity. Whole mounts of rat mammary glands, made after chronic feeding of 4-hydroxyphenylretinamide, showed that it had a marked antiproliferative effect on mammary epithelium.

Topics: Animals; Female; Liver; Mammary Glands, Animal; Mammary Neoplasms, Experimental; Methylnitrosourea; Organ Culture Techniques; Rats; Trachea; Tretinoin; Vitamin A; Vitamin A Deficiency

Animal studies have shown. a) an association between vitamin A and cancers of epithelial origin, and b) that vitamin A and its analogues delay tumour appearance, retard tumour growth and regress tumours induced by carcinogenic polycyclic aromatic hydrocarbons. Human epidemiological and biochemical studies suggest that cancers of epithelial origin may be associated with vitamin A deficiency. Vitamin A and its analogues may have a prophylactic and a therapeutic role in cancers of epithelial origin.

Topics: Animals; Bronchial Neoplasms; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; Humans; Lung Neoplasms; Mouth Neoplasms; Neoplasms, Experimental; Pharyngeal Neoplasms; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Animals; Chromatography, High Pressure Liquid; Chromatography, Ion Exchange; Intestinal Mucosa; Male; Rats; Tretinoin; Vitamin A; Vitamin A Deficiency

Incubation of [3H]retinoic acid in the presence of hamster liver 10000g supernatant produces several metabolites that are more polar than the parent compound. Two of these metabolites are identical with synthetic all-trans-4-hydroxyretinoic acid and all-trans-4-oxoretinoic acid both in ultraviolet absorption and mass spectral characteristics and in migration rates on two different reverse-phase high-pressure liquid chromatographic systems. The metabolites produced in a cell-free liver incubation reaction also migrate on a high-pressure liquid chromatography column together with metabolites isolated from a tracheal organ culture system. Both the metabolites and the synthetic standards show less biological activity than the parent all-trans-retinoic acid in a tracheal organ culture assay.

Topics: Animals; Biological Assay; Chromatography, High Pressure Liquid; Cricetinae; Female; Liver; Male; Mass Spectrometry; Spectrophotometry, Ultraviolet; Trachea; Tretinoin; Vitamin A; Vitamin A Deficiency

Rats were fed diets deficient [-A] or sufficient [+A] (3 mg retinol equivalents/kg) in vitamin A, and without [-RA] or with [+RA] (12 mg/kg) retinoic acid supplementation for up to 33 days. Rats with plasma vitamin A levels ranging from 7 to 62 micrograms/dl were studied at intervals during progressive depletion of liver stores of vitamin A (expt. 2) and when liver stores were nearly exhausted (less than 10 micrograms/g) or replete (up to 100 micrograms/g) with vitamin A (expt. 1). A dose of retinyl acetate in corn oil (20 micrograms retinol equivalents) was administered by intubation directly into the stomach. The relative dose response (RDR), expressed as a percentage and defined as the absolute magnitude of the rise in plasma vitamin A levels 5 hours after the dose of retinyl acetate, divided by the plasma level of vitamin A attained after 5 hours, was determined for each rat and correlated over a wide range of vitamin A plasma and liver levels. An RDR above 50% invariably was associated with low plasma levels (10 to 30 micrograms/dl) and low liver stores (less than 10 micrograms/g) of vitamin A, whereas an RDR of less than 40% was associated with plasma levels above 30 micrograms/dl and liver stores ranging from 3 to 100 micrograms/g.

Topics: Animals; Body Weight; Dose-Response Relationship, Drug; Liver; Male; Rats; Tretinoin; Vitamin A; Vitamin A Deficiency

Rats were fed vitamin A-deficient diets either alone, supplemented with retinoic acid (RA), or of limited protein quality or quantity (7%rice or 7% casein protein); one group was fed 7% rice protein supplemented with vitamin A. Plasma and liver levels of vitamin A were determined serially. Plasma levels in rats fed otherwise adequate vitamin A-deficient diets remained above 30 micrograms/dl until liver reserves were below 10 micrograms/g tissue, at which point plasma levels decreased in some but not all rats while liver levels continued to decline (at a slower rate) to levels as low as 3 micrograms/g. Supplementation with RA caused an immediate and sustained reduction of 15 to 20 micrograms/dl in circulating vitamin A. At 7% dietary protein, plasma levels of vitamin A remained above 30 micrograms/dl when casein protein was fed or when the rice protein diet was supplemented with dietary vitamin A, but not when the rice protein diet was fed without an exogenous source of the vitamin. A scheme is proposed suggesting possible regulatory mechanisms that might control homeostatic levels of plasma vitamin A.

Topics: Animals; Body Weight; Caseins; Dietary Proteins; Liver; Male; Oryza; Plant Proteins; Rats; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Animals; Biological Assay; Chromatography, High Pressure Liquid; Cricetinae; Female; Intestinal Mucosa; Kinetics; Liver; Male; Mesocricetus; Organ Specificity; Subcellular Fractions; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Animals; Cricetinae; Female; Intestinal Mucosa; Kinetics; Liver; Male; Organ Culture Techniques; Organ Specificity; Structure-Activity Relationship; Testis; Trachea; Tretinoin; Vitamin A; Vitamin A Deficiency

Experiments were conducted to determine the sequence and reliability of appearance of key signs of vitamin A deficiency. Rapid and essentially synchronous vitamin A deficiency was induced by the withdrawal of retinoic acid from mature (190--210 g) stringently vitamin A-deficient male rats reared by feeding early growth plateau (60--70 g) vitamin A-deprived rats diets first supplemented with and then lacking in 2 micrograms retinoic acid per gram diet in repeating 18 day:10 day supplementation:deprivation cycles. Growth was depressed within 1 to 2 days of the withdrawal of retinoic aicid whether animals were force-fed or were fed ad libitum. Similar patterns were obtained when animals were fed 5 or 10 micrograms retinoic acid per gram diet. Appetite was depressed (1--2 days) whether animals were fed 18% casein diets, or were given 10% dextrose drinking solutions only. Decreased food intake was not due to impaired taste function or to poor palatability of the deficient diet. Bilateral electrolytic lesions in the ventromedial nucleus of the hypothalamus or anterior prepyriform cortex failed to prevent or to delay loss of appetite. Supplementation with antibiotics decreased body weight losses in the late stages of deficiency and increased survival time. Other signs of deficiency (days until onset following retinoate withdrawal; percent incidence) were: decreased intestinal goblet cell numbers (2--3; 80), decreased pilocarpine induced salivation (6--8; 80), tracheal metaplasia (6--8; 80), transient periocular porphyria (6--8; 60), altered salivary gland morphology (9--10; 80), decreased stomach emptying in force-fed animals (12; 70), twisting (12; 5) and leg crippling (12; 5). We conclude that the sequence of appearance of individual signs of deficiency following the induction of synchronous vitamin A deficiency is highly reproducible, and that the more general use of synchronously deficient animals would materially assist studies of cause-effect relationships in vitamin A deficiency.

Topics: Animals; Appetite; Body Weight; Chlortetracycline; Hypothalamus; Male; Rats; Taste; Tretinoin; Vitamin A Deficiency

Topics: Animals; Body Composition; Body Weight; Feedback; Liver; Male; Rats; Retinol-Binding Proteins; Time Factors; Tissue Distribution; Tretinoin; Vitamin A; Vitamin A Deficiency

A retinoid was isolated by a multistep procedure from the small intestines of vitamin A-deficient rats given a single dose of retinoic acid. The compound, designated 8II, was pure, as demonstrated by four high-pressure liquid chromatographic procedures. It was positively identified as 5,8-oxyretinoic acid by ultraviolet spectrophotometry, mass spectrometry, and spectral and chromatographic comparison to known compounds. It is probable that 5,8-oxyretinoic acid was produced from 5,6-epoxyretinoic acid under the acidic conditions used in the isolation. It is highly probable, therefore, that the natural product is 5,6-epoxyretinoic acid.

Topics: Animals; Chromatography, High Pressure Liquid; Intestinal Mucosa; Intestine, Small; Rats; Spectrophotometry, Ultraviolet; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Animals; Complement System Proteins; Dietary Proteins; Hemolytic Plaque Technique; Male; Protein Deficiency; Rats; Time Factors; Tretinoin; Vitamin A Deficiency

Topics: Animals; Cell Differentiation; Female; Gestational Age; Litter Size; Necrosis; Placenta; Pregnancy; Rats; Tretinoin; Vitamin A Deficiency

A reverse phase high-pressure liquid chromatography system for rapid separation of various retinoids (vitamin A and its analogs) with little or no degradation is described. This method permits detection of as little as 22 pmol of retinoic acid. The procedure has been applied to the study of retinoic acid metabolism in vitamin A-deficient hamsters.

Topics: Animals; Chromatography, High Pressure Liquid; Cricetinae; Mesocricetus; Molecular Conformation; Retinaldehyde; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Animals; Cochlea; Ear, Inner; Rats; Retina; Saccule and Utricle; Semicircular Canals; Temporal Bone; Tretinoin; Vitamin A; Vitamin A Deficiency

Studies were conducted to investigate the in vivo and in vitro effects of retinol and retinoic acid on the synthesis of mannolipids and mannopeptides in rat liver. The incorporation of 14C-mannose into glycolipids and glycoproteins showed a decrease in vitamin A-depleted rats as compared with vitamin A-fed rats. By means of DEAE-cellulose, silicic acid and thin-layer chromatography, the mannose-containing lipids were separated into mannosyl retinyl phosphate (MRP, Rf 0.2) and dolichyl mannosyl phosphate (DMP, Rf 0.4), respectively. A rapid increase in the synthesis of labelled MRP was observed, exhibiting a peak between 25 and 60 min after intraperitoneal administration of retinol to vitamin A-depleted rats. Similarly, administration of retinoic acid brought about elevation of 14C-mannolipid (Rf 0.2) synthesis with a peak at 60 min after injection. On the other hand, the incorporation of 14C-mannose into DMP (Rf 0.4) remained unchanged by such treatment. In vitro addition of retinyl phosphate, but not retinoyl phosphate, markedly stimulated the synthesis of 14C-mannolipid (Rf 0.2), using crude membrane of rat liver and GDP-14C-mannose as the donor. These findings strongly suggest that not only retinol but also retinoic acid plays an important biological role in mannosyl transfer reaction in rat liver. However, the molecular participation of a metabolite of retinoic acid in the formation of a mannolipid and the structure of such a metabolite remain to be established.

Topics: Animals; Body Weight; Dolichol Monophosphate Mannose; Glycolipids; Glycopeptides; Glycoproteins; In Vitro Techniques; Liver; Male; Mannose; Mannosyltransferases; Polyisoprenyl Phosphate Monosaccharides; Polyisoprenyl Phosphate Sugars; Rats; Tretinoin; Vitamin A; Vitamin A Deficiency

A highly biologically active metabolite of retinoic acid (8III) has been isolated in pure form from intestinal mucosa of vitamin A deficient rats given [3H]retinoic acid. This metabolite has been positively identified as 5, 6-epoxyretinoic acid based on the ultraviolet absorption spectrum and mass spectrum of its methylated derivative. This identification was confirmed by cochromatography of the methylated metabolite and synthetic methyl 5, 6-epoxyretinoate on reverse-phase and straight-phase high-pressure liquid chromatography. The 5, 6-epoxyretinoic acid is a true in vivo generated metabolite of retinoic acid and not an artifact of the isolation procedure. In addition, 5, 8-oxyretinoic acid previously isolated in this laboratory from intestinal mucosa was probably generated from 5, 6-epoxyretinoic acid by the acidic conditions used in the extraction and isolation of the 5, 8-oxyretinoic acid.

Topics: Animals; Epoxy Compounds; Intestinal Mucosa; Male; Mass Spectrometry; Rats; Spectrophotometry, Ultraviolet; Tretinoin; Vitamin A; Vitamin A Deficiency

Eight children with corneal xerophthalmia (x2 or x3A) received standard high-protein diets and massive systemic vitamin A therapy. Retinoic acid, 0.1% in oil, was applied daily to one eye, and oil alone to the other. Topical retinoic acid proved safe and effective in speeding corneal healing, especially during the first critical days.

Topics: Administration, Topical; Animals; Child; Child, Preschool; Dietary Proteins; Female; Humans; Male; Rats; Tretinoin; Vitamin A; Vitamin A Deficiency; Xerophthalmia

The syntheses of the ring and four side-chain dihydroretinoic acids and/or their esters, 3-7, are described. The syntheses of several other retinoids containing a substituted aromatic ring are also included. The biological activity of the compounds was evaluated in vivo in a chemically induced mouse skin papilloma test and in vitro in two vitamin A deficient assays. The activity observed for 1a, 1c, and 2a in the former test was partially retained in the dihydro derivatives 4b, 4c, and 6b. Similar results were found in the in vitro assays.

Topics: Animals; Cells, Cultured; Cricetinae; Female; Mice; Neoplasms, Experimental; Organ Culture Techniques; Papilloma; RNA; Skin; Skin Neoplasms; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Animals; Carcinoma, Basal Cell; Humans; Neoplasms, Experimental; Papilloma; Precancerous Conditions; Skin Diseases; Skin Neoplasms; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Animals; Glycoproteins; Glycosides; Humans; Intestinal Mucosa; Liver; Mammals; Membranes; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Animals; Female; Progesterone; Rats; Tretinoin; Vagina; Vaginal Smears; Vitamin A; Vitamin A Deficiency

Testicular transfer RNA methyltransferase activity was examined in normal and vitamin A-deficient rats. The specific activity was reduced by 50% in the vitamin A-deficient rats. In addition, a 5-fold decrease in the extent of tRNA methylation was observed with enzyme preparations from deficient testes. Both the rate and extent of tRNA methylation returned to control levels in vitamin A-repleted rats. In contrast, retinoic acid repletion did not reverse the effect of vitamin A deficiency on testicular tRNA methyltransferase activity. The methylated nucleoside composition of tRNA methylated by extracts of vitamin A-deficient testes was altered dramatically compared to that of tRNA methylated by control testicular enzymes. Decreased testicular tRNA methyltransferase activity was noted in midly deficient rats before the onset of testicular degeneration suggesting that the decreased tRNA methyltransferase activity in the testes is primarily the result of vitamin A deficiency.

Topics: Animals; Kinetics; Male; Rats; Seminiferous Tubules; Subcellular Fractions; Testis; Tretinoin; tRNA Methyltransferases; Vitamin A; Vitamin A Deficiency

The half-life and metabolism of vitamin A were determined in a group of vitamin A-deficient retinoic acid supplemented rats after a single dose of 340 mug of [6,7-14C2] retinol. The total daily urinary radioactivity, plotted semilogarithmically as a function of days after injection, revealed three pools for retinol and/or metabolites in the rat: (1) a rapidly declining pool with half-life of 0.75 day; (2) a slowly declining pool with a constant rate of decrease and (3) a pool with a half-life of 13 days which begins at approximately 6 weeks after dose. The total daily fecal radioactivity also indicated three pools with half-lives of 2,28.5 and 11.5 days. The effect of retinoic acid feeding was observed on the fifth day after supplementation, as indicated by a decrease in the total daily urinary radioactivity. Thus, retinoic acid is probably in the metabolic pathway of retinol. The half-life and metabolism time of liver vitamin A in these rats were determined as 7 and 10 days, respectively. The specific activities of liver retinyl esters and retinol determined at different intervals after dose indicated continuous mixing of radioactive retinol with a pool of endogenous retinol. Blood retinol levels indicated normal values at 1 week after dose. However, they decreased at 2 weeks after dose and remained constant until the sixth week. The specific activity of blood retinol did not change indicating rapid equilibration after initial mixing and no further dilution from endogenous source.

Topics: Animals; Feces; Liver; Male; Rats; Time Factors; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Animals; Disease Models, Animal; Light; Membranes; Microscopy, Electron; Microscopy, Electron, Scanning; Phagocytosis; Photoreceptor Cells; Rats; Retinal Degeneration; Temperature; Tretinoin; Visual Fields; Vitamin A Deficiency

Topics: Amino Acids; Animals; Body Weight; Eye Proteins; In Vitro Techniques; Palmitates; Rats; Retina; Tretinoin; Trichloroacetic Acid; Vitamin A; Vitamin A Deficiency

Topics: Animals; Epithelium; Female; Humans; Lung Neoplasms; Mammary Neoplasms, Experimental; Neoplasms; Neoplasms, Experimental; Nutritional Requirements; Precancerous Conditions; Skin Neoplasms; Structure-Activity Relationship; Tretinoin; Urinary Bladder Neoplasms; Vitamin A; Vitamin A Deficiency

Both phospholipases A1 and A2 activities (EC 3.1.1.4) at pH 7.4 were found to be significantly decreased in retinol-deficient rat testes supplemented with retinoic acid as compared to retinol-fed controls using 1-acyl-2-[1-(14)C]-oleoyl-sn-glycero-3-phosphocholine as substrate. However, little or no difference was observed in phospholipase A1 activity at pH 3.0 in both groups of rats.

Topics: Animals; Hydrogen-Ion Concentration; Kinetics; Male; Phospholipases; Rats; Testis; Tretinoin; Vitamin A; Vitamin A Deficiency

Our study related to degenerescence of testes in vitamin A deficient rats led to the following observations : decrease of DNA, RNA on one hand and AMP, ADP and ATP on the other hand. These observations are considered as related to decrease of DNA polymerase activity.

Topics: Adenine Nucleotides; Animals; DNA; DNA Nucleotidyltransferases; Male; Organ Size; Rats; RNA; Testis; Tretinoin; Vitamin A Deficiency

Testes of rats contain two cellular binding proteins of interest in vitamin A metabolism. One protein binds retinoic acid with high specificity; the other binds retinol with high specificity. When the cellular retinol-binding protein was partially purified from rat testes, it exhibited fluorescence excitation and emission spectra similar to that of all-trans-retinol in hexane. Exposure of this preparation to UV light destroyed this fluorescence but spectra identical to the original were obtained after addition of retinol. Hexane extracts of the binding protein had fluorescence spectra identical to all-trans-retinol, suggesting that this compound is bound to the protein in vivo. Extracts of testes from retinol depleted rats were submitted to gel filtration but failed to show a retinol-like fluorescence at the elution position of retinol binding protein. This fluorescence was observed in the preparations from pair fed control animals. However, after addition of all-trans-retinol to the extracts from the depleted rats, fluorescence at that elution position was observed. This indicates that in testes of retinol depleted rats the cellular retinol binding protein is present but without bound retinol, in contrast to the non-depleted rats where 30-43% of the binding protein had bound retinol. The amounts of cellular retinol binding protein and retinoic acid binding protein in testes, as determined by sucrose gradient centrifugation, were found to be similar for retinol depleted and pair fed control rats.

Topics: Animals; Carrier Proteins; Male; Radiation Effects; Rats; Retinol-Binding Proteins; Retinol-Binding Proteins, Cellular; Structure-Activity Relationship; Testis; Tretinoin; Ultraviolet Rays; Vitamin A; Vitamin A Deficiency

Avitaminosis A, applied on deficient rats receiving retinoic acid leads to an important decrease of thymidine incorporation in testicular DNA in vivo as well as in vitro. On the contrary uridine incorporation in RNA is considerably increased in vitro as well as in vivo. The function of ribosomes, as measured by the aggregation ability in the polysomes, is not altered by vitamin A deficiency. From these results one can say that the degenerescence of rat testicle is accompagnied by a decrease of DNA anabolism and a stimulation of RNA anabolism due to an increase of its catabolism (compensatory synthesis).

Topics: Animals; DNA; Male; Organ Size; Polyribosomes; Rats; Ribosomes; RNA; Testis; Tretinoin; Vitamin A Deficiency

Topics: Animals; Body Weight; Liver; Male; Organ Size; Rats; Testis; Tretinoin; Vitamin A; Vitamin A Deficiency; Xerophthalmia

Topics: Animals; Growth; Isomerism; Rats; Retinol-Binding Proteins; Structure-Activity Relationship; Tretinoin; Vitamin A; Vitamin A Deficiency

Growth and development of the oestrogen-primed oviduct of immature chicks in terms of weight, length, total protein, total RNA, total DNA and total phospholipids are markedly arrested on deprivation of vitamin A; supplementation with retinyl acetate reversed the effect of the deficiency almost fully, whereas retinoic acid was only partly effective.

Topics: Animals; Animals, Newborn; Chickens; Estradiol; Female; Organ Size; Oviducts; Tretinoin; Vitamin A Deficiency

Up to date, the treatment of palmar/plantar hyperkeratoses presents a therapeutic problem. The known therapeutic procedures result in short-term improvement only, if any at all. In these investigations involving 68 patients suffering from palmar/plantar hyperkeratoses of different etiology, small doses of vitamin A acid locally applied, produced a striking improvement in hypertrophic lichen planus of palms or soles: the regression was complete and in most cases permanent. The skin texture of patients with genetic keratoses and callosities became normal within a few weeks: but this condition remained free of symptoms only as long as vitamin A acid was used as a maintenance dose once or twice weekly. In hyperkeratotic eczema, pityriasis rubra pilaris, and verrucae plantaris vitamin A acid locally applied was found to be unsuitable for treatment. The possible side effects of this treatment are mentioned. Several possibilities regarding the way of action of vitamin A acid are discussed.

Topics: Callosities; Eczema; Humans; Keratoderma, Palmoplantar; Lichen Planus; Long-Term Care; Occlusive Dressings; Ointments; Pityriasis Rubra Pilaris; Tretinoin; Vitamin A; Vitamin A Deficiency; Warts

Topics: Animals; Antibody Formation; Body Weight; Chickens; Immune Adherence Reaction; Liver; Male; Poultry Diseases; Tretinoin; Vitamin A; Vitamin A Deficiency

Studies were conducted to examine in detail the effects of vitamin A deficiency on fetal growth and development in the rat. The gradations of deficiency were examined in two studies. The first included total vitamin A depletion followed by retinoic acid supplements, and the second included three different levels of restricted intake of retinyl acetate (42, 16, or 8 mug of retinol equivalents/day/kg of body weight) in vitamin A-depleted rats. In the first study, extensive fetal resorption and death were observed in retinoic acid-fed females after day 14 of gestation. These findings confirmed the morphological studies of Thompson and associates (Proc. Roy. Soc. London, Ser. B 159, 510-535, 1964) who found the earliest Detectable histological lesions to be in the placentas at days 15-16 of pregnancy. Analyses were carried out of the total weight, the DNA, RNA, and protein contents of fetuses and placentas of different gestational ages in retinyl ester-fed and retinoic acid-fed females. Biochemical changes indicative of a reduced rate of cell division were observed in both fetus and placenta by day 14 in the retinoic acid-fed rats. The few live fetuses in this group maintained a growth rate of only 60-70% of that of the fetuses of retinyl ester-fed dams after day 14. By contrast, the growth rate of the placentas (of live fetuses) after day 14 of gestation was not as consistently affected by retinol deficiency. Restriction of retinyl acetate intake (in the second study) significantly reduced both the total litter size and the number of live pups per litter. Most of the females in the retinyl acetate-restricted groups delivered pups that had normal body weight and appeared normal on visual inspection. Significant differences from normal controls were seen only in the neonates from dams given 8 mug of retinol equivalents (per kg of body weight per day), which had smaller livers and kidneys than the control neonates. In contrast, the weights of the brains of the neonates in all three retinyl acetate-restricted groups showed no differences from control values. Vitamin A assays on maternal and neonatal sera and livers indicated that the transport of vitamin A across the placenta was well regulated, and suggested that this transport is maintained with high priority in the presence of maternal deficiency. The effects of vitamin A deficiency on fetal growth and development might reflect primary effects on the placenta, with secondary effects on the fetus, or primary d

Topics: Animals; Birth Weight; DNA; Female; Fetus; Growth; Litter Size; Maternal-Fetal Exchange; Organ Size; Placenta; Pregnancy; Pregnancy Complications; Proteins; Rats; RNA; Tretinoin; Vitamin A; Vitamin A Deficiency

A new analogue of vitamin A, viz., retinoic acid anhydride was prepared, for the first time, by the action of thionyl chloride on retinoic acid in benzene containing pyridine. The amhydride was charcterised by its chromatographic properties, elemental analysis, ultraviolet absorption, infrared and nuclear magnetic resonance spectral characteristics. The compound could be readily hydrolysed to retinoic acid both by acid and alkali treatments and reduced by lithium aluminium hydride to vitamin A alcohol (retinol). The spectral changes with antimony trichloride reagent were similar to those observed for retinoic acid. The metabolism of retinoic acid anhydride was found to be similar to that of retinoic acic. When administered either orally or intraperitoneally, the compound promotes growth in vitamin A-deficient rats. Time-course experiments revealed that retinoic acid anhydride is converted into retinoic acid by non-enzymatic hydrolysis and thereby exerts its biological activity. The biopotency of the anhydride was found to be nearly the same as that of the acid. A new method of preparing esters of retinoic acid employing retinoic acid anhydride as an intermediate, has been described.

Topics: Absorption; Animals; Body Weight; Esters; Male; Rats; Stimulation, Chemical; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Animals; Darier Disease; Humans; Rats; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Acanthosis Nigricans; Administration, Topical; Adolescent; Central Nervous System Diseases; Diverticulum; Female; Gastrointestinal Diseases; Humans; Skin; Sural Nerve; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Animals; Biological Assay; Body Weight; Carotenoids; Fatty Acids, Unsaturated; Intestinal Mucosa; Ketones; Liver; Magnetic Resonance Spectroscopy; Male; Oxidation-Reduction; Rats; Spectrophotometry, Infrared; Spectrophotometry, Ultraviolet; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Animals; Blindness; Chick Embryo; Esters; Meat; Pharmacology; Poultry; Pupil; Research; Toxicology; Tretinoin; Vitamin A Deficiency

Topics: Animals; Cerebrospinal Fluid; Cerebrospinal Fluid Pressure; Diet; Liver; Pharmacology; Research; Swine; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Alcohols; Animals; Diet; Female; Fetal Death; Histology; Placenta; Pregnancy; Pregnancy, Animal; Rats; Reproduction; Research; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Biochemical Phenomena; Biochemistry; Liver; Rats; Research; Sulfhydryl Compounds; Tretinoin; Vitamin A; Vitamin A Deficiency

Topics: Alcohols; Animals; Diet; Humans; Male; Rats; Reproduction; Testis; Tretinoin; Vitamin A; Vitamin A Deficiency