transforming-growth-factor-beta and Colitis

The most effective way to control severity and mortality rate of the novel coronavirus disease (COVID-19) is through sensitive diagnostic approaches and an appropriate treatment protocol. We aimed to identify the effect of adding corticosteroid and Tocilizumab to a standard treatment protocol in treating COVID-19 patients with chronic disease through hematological and lab biomarkers.. This study was performed retrospectively on 68 COVID-19 patients with chronic disease who were treated by different therapeutic protocols. The patients were categorized into four groups: control group represented the patients' lab results at admission before treatment protocols were applied; group 1 included patients treated with anticoagulants, Hydroxychloroquine, and antibiotics; group 2 comprised patients treated with Dexamethasone; and group 3 included patients treated with Dexamethasone and Tocilizumab.. The study paves the way into the effectiveness of combining Dexamethasone with Tocilizumab in treatment COVID-19 patients with chronic diseases.. La forma más eficaz de controlar la gravedad y la tasa de mortalidad de la enfermedad del nuevo coronavirus (COVID-19) es mediante enfoques de diagnóstico sensibles y un protocolo de tratamiento adecuado. Nuestro objetivo fue identificar el efecto de agregar corticosteroides y tocilizumab a un protocolo de tratamiento estándar en el tratamiento de pacientes con COVID-19 con enfermedad crónica a través de biomarcadores hematológicos y de laboratorio.. Este estudio se realizó de forma retrospectiva en 68 pacientes COVID-19 con enfermedad crónica que fueron tratados por diferentes protocolos terapéuticos. Los pacientes se clasificaron en cuatro grupos: el grupo de control representaba los resultados de laboratorio de los pacientes en el momento de la admisión antes de que se aplicaran los protocolos de tratamiento; el grupo 1 incluyó a pacientes tratados con anticoagulantes, hidroxicloroquina y antibióticos; el grupo 2 estaba compuesto por pacientes tratados con dexametasona; y el grupo 3 incluyó a pacientes tratados con dexametasona y tocilizumab.. El estudio allana el camino hacia la eficacia de la combinación de dexametasona con tocilizumab en el tratamiento de pacientes con COVID-19 con enfermedades crónicas.. The Child-Mother Index constitutes a potential useful risk factor indicator for statistical analyses on data after birth. The value of the Child-Mother Index based on the estimated fetal weight before birth deserves evaluation.. Six ceria supports synthesized by various synthesis methodologies were used to deposit cobalt oxide. The catalysts were thoroughly characterized, and their catalytic activity for complete methane oxidation was studied. The supports synthesized by direct calcination and precipitation with ammonia exhibited the best textural and structural properties as well as the highest degree of oxidation. The remaining supports presented poorer textural properties to be employed as catalytic supports. The cobalt deposited over the first two supports presented a good dispersion at the external surface, which induced a significant redox effect that increased the number of Co. Some studies show that children with obesity are more likely to receive a diagnosis of depression, anxiety, or attention-deficit hyperactivity disorder (ADHD). But this does not necessarily mean obesity causes these conditions. Depression, anxiety, or ADHD could cause obesity. A child's environment, including family income or their parents' mental health, could also affect a child's weight and mental health. Understanding the nature of these relationships could help scientists develop better interventions for both obesity and mental health conditions. Genetic studies may help scientists better understand the role of the environment in these conditions, but it's important to consider both the child's and their parents’ genetics in these analyses. This is because parents and children share not only genes, but also environmental conditions. For example, families that carry genetic variants associated with higher body weight might also have lower incomes, if parents have been affected by biases against heavier people in society and the workplace. Children in these families could have worse mental health because of effects of their parent’s weight, rather than their own weight. Looking at both child and adult genetics can help disentangle these processes. Hughes et al. show that a child's own body mass index, a ratio of weight and height, is not strongly associated with the child’s mental health symptoms. They analysed genetic, weight, and health survey data from about 41,000 8-year-old children and their parents. The results suggest that a child's own BMI does not have a large effect on their anxiety symptoms. There was also no clear evidence that a child's BMI affected their symptoms of depression or ADHD. These results contradict previous studies, which did not account for parental genetics. Hughes et al. suggest that, at least for eight-year-olds, factors linked with adult weight and which differ between families may be more critical to a child's mental health than a child’s own weight. For older children and adolescents, this may not be the case, and the individual’s own weight may be more important. As a result, policies designed to reduce obesity in mid-childhood are unlikely to greatly improve the mental health of children. On the other hand, policies targeting the environmental or societal factors contributing to higher body weights, bias against people with higher weights, and poor child mental health directly may be more beneficial.. The development of an efficient photocatalyst for C2 product formation from CO. Оценка антиастенического эффекта последовательной терапии левокарнитином (ЛК) и ацетилкарнитином (АЛК) пациентов с артериальной гипертензией и/или ишемической болезнью сердца (ИБС) с астеническим синдромом (АС).. В открытое сравнительное исследование были включены 120 пациентов в возрасте 54—67 лет с артериальной гипертензией и/или ИБС с АС. Пациенты 1-й группы (. У больных 1-й группы отмечено статистически значимое уменьшение различных проявлений АС. Отличия носили достоверный характер по сравнению как с исходным уровнем, так и со 2-й группой. Установлено эндотелийпротективное действие ЛК и АЛК.. Полученные результаты свидетельствуют, что у таких коморбидных пациентов использование ЛК и АЛК уменьшает выраженность проявлений АС, а установленные эндотелиотропные свойства препаратов позволяют рекомендовать их в составе комплексной персонифицированной терапии пациентов с сердечно-сосудистыми заболеваниями.. Naproxen sodium 440 mg/diphenhydramine 50 mg combination demonstrated improvement in sleep maintenance (WASO) vs. naproxen sodium 550 mg and higher efficiency in average daily pain reduction compared with the comparison groups. The treatment was well tolerated There were no serious or unexpected adverse events reported in the study.. Сравнительный анализ эффективности и безопасности новой комбинации напроксена натрия и дифенгидрамина у пациентов с неспецифическим болевым синдромом в пояснично-крестцовом отделе спины (M54.5 «Боль внизу спины») и нарушением сна (G47.0 «Нарушения засыпания и поддержания сна [бессонница]»).. Проведено проспективное многоцентровое рандомизированное открытое сравнительное в параллельных группах клиническое исследование. Пациенты были рандомизированы в 3 группы. Больные 1-й группы получали напроксен натрия (440 мг) и дифенгидрамин (50 мг), 2-й — напроксен натрия (550 мг), 3-й — парацетамол (1000 мг) и дифенгидрамин (50 мг). Исследуемые препараты пациенты принимали однократно перед сном в течение 3 дней. Все пациенты также принимали 275 мг (1 таблетка) напроксена натрия в качестве препарата фоновой терапии. Первичным критерием эффективности было общее время бодрствования после наступления сна (WASO), измеряемое методом актиграфии. Также использовались критерии оценки продолжительности и качества сна и выраженности боли.. Анализ эффективности проведен для ITT популяции (. Применение комбинации напроксена натрия (440 мг) и дифенгидрамина (50 мг) характеризовалось более выраженным поддержанием сна по сравнению с напроксеном натрия 550 мг и более высокой эффективностью в отношении снижения интенсивности боли по сравнению со 2-й и 3-й группами. Отмечена хорошая переносимость препарата, серьезных нежелательных явлений зарегистрировано не было.

Topics: Acetaminophen; Acetylcarnitine; Acetylcholinesterase; Acids; Acinetobacter baumannii; Acinetobacter Infections; Adaptation, Psychological; Adolescent; Adsorption; Adult; Aged; Alcohol Drinking; Alzheimer Disease; Amikacin; Ammonia; Anaerobiosis; Animals; Anorexia; Anti-Bacterial Agents; Anti-Infective Agents; Anti-Inflammatory Agents; Anti-Inflammatory Agents, Non-Steroidal; Antineoplastic Agents; Anxiety; Aptamers, Nucleotide; Asthenia; Attention Deficit Disorder with Hyperactivity; Bacterial Proteins; Beryllium; beta-Lactamases; Biofuels; Biomass; Biosensing Techniques; Bismuth; Blister; Body Mass Index; Body Surface Area; Boronic Acids; Brain; Breast Neoplasms; Butyrylcholinesterase; Cannabis; Carbapenems; Carbonyl Cyanide m-Chlorophenyl Hydrazone; Carboxylic Acids; Carcinoma, Hepatocellular; Cardiovascular Diseases; Carnitine; Case-Control Studies; Catalysis; Cell Cycle Proteins; Cell Line, Tumor; Cell Proliferation; Child; China; Cholinesterase Inhibitors; Clarithromycin; Clostridioides; Clostridioides difficile; Clostridium Infections; Cohort Studies; Colistin; Colitis; Colon; Coloring Agents; Coronary Artery Bypass; Creatinine; Crystalloid Solutions; Cytokines; Depression; Dextran Sulfate; Dextrans; Diabetes Mellitus, Type 2; Diabetic Retinopathy; Diarrhea; Dietary Supplements; Diphenhydramine; Disease Models, Animal; Disease Outbreaks; Double-Blind Method; Doxorubicin; Drosophila; Drug Tapering; Dysbiosis; Electrons; Escherichia coli; Extracellular Vesicles; Fatigue; Female; Fermentation; gamma-Cyclodextrins; Gastrointestinal Microbiome; Glucose; Graft Survival; Graft vs Host Disease; Head and Neck Neoplasms; Heart Arrest, Induced; Hematopoietic Stem Cell Transplantation; High-Intensity Interval Training; Hippocampus; Humans; Hydrogen-Ion Concentration; Hypertension; Incidence; Interferon-gamma; Italy; Kinetics; Klebsiella Infections; Klebsiella pneumoniae; Lab-On-A-Chip Devices; Lactoferrin; Larva; Length of Stay; Lignin; Liver; Liver Neoplasms; Liver Transplantation; Living Donors; Low Back Pain; Lung; Lung Volume Measurements; Macrophages; Male; Melphalan; Men; Mendelian Randomization Analysis; Meropenem; Methane; Mice; Mice, Inbred C57BL; Microbial Sensitivity Tests; Mitochondrial Proteins; Molecular Docking Simulation; Molecular Structure; Mothers; Motivation; Mycoplasma; Mycoplasma hominis; Mycoplasma Infections; NAD; Nanocomposites; Nanoparticles; Nanotubes, Carbon; Naproxen; Neovascularization, Pathologic; Neurons; Nitrates; Nucleolin; Opuntia; Paratyphoid Fever; Phenotype; Phosphatidylinositol 3-Kinases; Phytochemicals; Plant Extracts; Pregnancy; Prevalence; Prospective Studies; Proto-Oncogene Proteins c-akt; Pulmonary Disease, Chronic Obstructive; Rats; Rats, Wistar; Resveratrol; Retrospective Studies; Rifampin; Risk Factors; RNA, Messenger; Selenium; Sleep; Social Behavior; Soil; Soil Pollutants; Squamous Cell Carcinoma of Head and Neck; Staphylococcus aureus; Structure-Activity Relationship; Suicidal Ideation; Suicide; Superoxide Dismutase-1; Surveys and Questionnaires; Swimming; Syndrome; Tannins; Temperature; Transforming Growth Factor beta; Transplantation Conditioning; Treatment Outcome; Triple Negative Breast Neoplasms; Troponin T; Tumor Microenvironment; United Kingdom; Ureaplasma; Ureaplasma urealyticum; Urinary Tract Infections; Viscum; Waste Disposal Facilities; Wastewater; Water; Water Pollutants, Chemical; Wolfiporia; Young Adult

Ubiquitination is a three-step enzymatic cascade for posttranslational protein modification. It includes the ubiquitin-activating enzyme (E1), ubiquitin-conjugating enzyme (E2), and ubiquitin ligase (E3). RING-type E3 ubiquitin ligases catalyse the posttranslational proteolytic and nonproteolytic functions in various physiological and pathological processes, such as inflammation-associated signal transduction. Resulting from the diversity of substrates and functional mechanisms, RING-type ligases regulate microbe recognition and inflammation by being involved in multiple inflammatory signalling pathways. These processes also occur in autoimmune diseases, especially inflammatory bowel disease (IBD). To understand the importance of RING-type ligases in inflammation, we have discussed their functional mechanisms in multiple inflammation-associated pathways and correlation between RING-type ligases and IBD. Owing to the limited data on the biology of RING-type ligases, there is an urgent need to analyse their potential as biomarkers and therapeutic targets in IBD in the future.

Topics: Animals; Autophagy; Biomarkers; Colitis; Humans; Immune System; Inflammation; Inflammatory Bowel Diseases; MAP Kinase Signaling System; Mice; Phosphatidylinositol 3-Kinases; Protein Processing, Post-Translational; RNA, Untranslated; Signal Transduction; Transforming Growth Factor beta; Ubiquitin; Ubiquitin-Protein Ligases; Ubiquitination

Dendritic cells (DCs) mediate tolerance to food antigens, limit reactivity to the gut microbiota and are required for optimal response to intestinal pathogens. Intestinal DCs are heterogeneous but collectively generate both regulatory and effector T cell responses. The balance of outcomes is determined by the activity of functionally distinct DC subsets and their modulation by environmental cues. DCs constantly sample luminal content to monitor for pathogens; the significance of the various pathways by which this occurs is incompletely understood. Intestinal DC have distinctive properties shaped by local host, dietary and microbial signals. These properties include the ability to produce all-trans retinoic acid (RA) and imprint gut tropism on T cells they activate. In the steady-state, subsets of intestinal DC are potent generators of inducible Treg, aided by their ability to activate TGFβ and produce RA. However, responses induced by steady-state intestinal DCs are not exclusively regulatory in nature; effector T cells with specificity for commensal bacterial can be found in the healthy mucosa and these can be locally controlled to prevent inflammation. The ability of intestinal DCs to enhance effector responses in infection or sustain inflammation in disease is likely to involve both modulation of the local DC population and recruitment of additional populations. Immune pathways in the pathogenesis of inflammatory bowel disease can be mapped to DCs and in inflamed intestinal tissue, DCs show increased expression of microbial recognition machinery, activation, and production of key immunological mediators. Intestinal DCs may be targeted for disease therapy or to improve vaccine responses.

Topics: Allergens; Animals; Cell Communication; Colitis; Dendritic Cells; Disease Models, Animal; Food Hypersensitivity; Gastrointestinal Microbiome; Humans; Immunity, Mucosal; Immunologic Factors; Intestinal Mucosa; Intraepithelial Lymphocytes; T-Lymphocytes, Cytotoxic; T-Lymphocytes, Regulatory; Transforming Growth Factor beta; Tretinoin

Acute inflammation is a response to an alteration induced by a pathogen or a physical or chemical insult, which functions to eliminate the source of the damage and restore homeostasis to the affected tissue. However, chronic inflammation triggers cellular events that can promote malignant transformation of cells and carcinogenesis. Several inflammatory mediators, such as TNF-α, IL-6, TGF-β, and IL-10, have been shown to participate in both the initiation and progression of cancer. In this review, we explore the role of these cytokines in important events of carcinogenesis, such as their capacity to generate reactive oxygen and nitrogen species, their potential mutagenic effect, and their involvement in mechanisms for epithelial mesenchymal transition, angiogenesis, and metastasis. Finally, we will provide an in-depth analysis of the participation of these cytokines in two types of cancer attributable to chronic inflammatory disease: colitis-associated colorectal cancer and cholangiocarcinoma.

Topics: Bile Duct Neoplasms; Bile Ducts, Intrahepatic; Carcinogenesis; Cholangiocarcinoma; Chronic Disease; Colitis; Colorectal Neoplasms; Gene Expression; Humans; Inflammation; Interleukin-10; Interleukin-6; Oxidative Stress; Transforming Growth Factor beta; Tumor Microenvironment; Tumor Necrosis Factor-alpha

Transforming growth factor-β (TGF-β) plays a central role in a wide array of cellular functions including control of cell growth and differentiation, embryonic development, wound healing, angiogenesis, and immune regulation. In the gastrointestinal tract, TGF-β can either promote or suppress inflammation and cancer formation. This report reviews recent data on the role of TGF-β in the pathogenesis of inflammatory bowel disease and how TGF-β might contribute to the cancer risk associated with chronic inflammation of the gut.

Topics: Animals; Cell Proliferation; Cell Transformation, Neoplastic; Colitis; Colonic Neoplasms; Humans; Inflammatory Bowel Diseases; Lymphocyte Activation; Mice; Th17 Cells; Transforming Growth Factor beta

In the human body, mucosal surfaces of the intestinal tract are the largest and one of the most complex parts of the immune system. These surfaces are covered by a layer of epithelial cells which allows efficient absorption of nutrients but also serves to separate the intestine from an environment loaded with potential harmful agents. Discrimination between beneficial commensal bacteria, harmless antigens and pathogenic microorganisms is a central issue in the role that gut immune cells play in maintaining the balance between immune response and tolerance. However, the basis of this discrimination in the mucosal immune system, where this occurs and how it can affect both local and systemic responses is not yet well understood. Nevertheless, antigen uptake and presentation seems to be a crucial factor in this issue. In this review, we will discuss the key role of immune intestinal cells in the development of mucosal immunity, tolerance and disease.

Topics: Animals; Bacteria; Colitis; Dendritic Cells; Epithelial Cells; Humans; Immune Tolerance; Immunity, Mucosal; Interleukin-10; Intestinal Mucosa; Intestinal Neoplasms; Intestines; Lymph Nodes; Macrophages; Peyer's Patches; T-Lymphocytes, Regulatory; Transforming Growth Factor beta; Tumor Escape

IL-6 plays a pivotal role in immune responses and certain oncologic conditions. The intense investigation of its biological activity and function led to the discovery of two different IL-6-driven signalling pathways. Binding to the membrane-bound IL-6 receptor (mIL-6R, CD126) causes the recruitment of two gp130 co-receptor molecules (CD130) and the activation of intracellular signalling cascades via gp130. Although this classical pathway is mainly limited to hepatocytes, neutrophils, monocytes/macrophages and certain other leukocyte populations, which express IL-6R on their surface, an alternative mechanism has also been described. Proteolytic cleavage of the mIL-6R protein or translation from alternatively spliced mRNA leads to the generation of a soluble form of the IL-6R (sIL-6R), which is likewise able to bind to IL-6. The resulting IL-6/sIL-6R complex is also capable of binding to gp130 and inducing intracellular signalling. Through this so-called 'trans-signalling' mechanism, IL-6 is able to stimulate cells that lack an endogenous mIL-6R. High levels of IL-6 and sIL-6R have been reported in several chronic inflammatory and autoimmune diseases as well as in cancer. Preclinical animal disease models have provided strong evidence that specific blockade of IL-6-regulated signalling pathways represents a promising approach for the therapy of these diseases. An optimised variant of the recently described fusion protein sgp30Fc is now heading towards its clinical evaluation.

Topics: Alternative Splicing; Animals; Anti-Inflammatory Agents; Antineoplastic Agents; Arthritis; Arthritis, Rheumatoid; Asthma; Autoimmune Diseases; Cell Line; Clinical Trials as Topic; Colitis; Colonic Neoplasms; Cytokine Receptor gp130; Disease Models, Animal; Drug Delivery Systems; Drug Evaluation, Preclinical; Humans; Inflammation; Interleukin-6; Leukocytes; Male; Mice; Neoplasms; Receptors, Interleukin-6; Recombinant Fusion Proteins; Signal Transduction; Solubility; Transforming Growth Factor beta

The gastrointestinal (GI) tract is the main interface where the body encounters exogenous antigens. It is crucial that the local response here is tightly regulated to avoid an immune reaction against dietary antigens and commensal flora while still mounting an efficient defense against pathogens. Faults in establishing intestinal tolerance can lead to disease, inducing local and often also systemic inflammation. Studies in human as well as in animal models suggest a role for regulatory T cells (Tregs) in maintaining intestinal homeostasis. Transfer of Tregs can not only prevent the development of colitis in animal models but also cure established disease, acting both systemically and at the site of inflammation. In this review, we discuss the major regulatory pathways, including transforming growth factor-beta (TGF-beta), interleukin-10 (IL-10), and cytotoxic T-lymphocyte antigen-4 (CTLA-4), and their role in Treg-mediated control of systemic and mucosal responses. In addition, we give an overview of the known mechanisms of lymphocyte migration to the intestine and discuss how CD103 expression can influence the balance between regulatory and effector T cells. Further understanding of the factors that control the activity of Tregs in different immune compartments may facilitate the design of strategies to target regulation in a tissue-specific way.

Topics: Animals; Antigens, CD; Antigens, Differentiation; Cell Movement; Colitis; CTLA-4 Antigen; Humans; Immunosuppression Therapy; Integrin alpha Chains; Interleukin-10; Intestinal Mucosa; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Transfer of CD4+ T cells to immune-deficient mice in the absence of the CD25+ subset leads to the development of colitis, indicating that regulatory cells capable of controlling a bacteria-driven inflammatory response are present in normal mice. Cells with this function are present in the thymus as well as in the periphery of germ-free mice, suggesting they may be reactive with self-antigen. These cells resemble CD4+CD25+ cells that inhibit organ-specific autoimmunity, suggesting that a similar subset of regulatory T cells may control responses to self and foreign antigens. Development of colitis is dependent on accumulation of activated CD134L+ dendritic cells (DC) in the mesenteric lymph nodes, which is inhibited by CD4+CD25+ cells, indicating that regulatory T cells may control DC activation in vivo. Whilst inhibition of T-cell activation in vitro by CD4+CD25+ cells does not involve interleukin-10 and transforming growth factor-beta, these cytokines are required for the suppression of colitis. It may be that control of responses that activate the innate immune system requires multiple mechanisms of immune suppression. Recently, we identified CD4+CD25+ cells with immune suppressive activity in the thymus and peripheral blood of humans, raising the possibility that dysfunction in this mechanism of immune regulation may be involved in the development of autoimmune and inflammatory diseases.

Topics: ADP-ribosyl Cyclase; ADP-ribosyl Cyclase 1; Animals; Antigens, CD; Antigens, Differentiation; Bacteria; Colitis; Dendritic Cells; Humans; Inflammatory Bowel Diseases; Interleukin-10; Leukocyte Common Antigens; Membrane Glycoproteins; NAD+ Nucleosidase; Receptors, Interleukin-2; T-Lymphocyte Subsets; Thymus Gland; Transforming Growth Factor beta

The above new findings concerning the immunological mechanisms governing mucosa, immune responses and oral tolerance in TCR-transgenic mice, as well as those operative in mice with experimental colitis, greatly expand our understanding of the processes that normally control mucosal inflammation and possibly other types of inflammation as well (Fig. 1). They indicate that, in the nondiseased mouse, ingested proteins evoke a Th1-cell (IFN-gamma) response in the mucosal follicles that is quickly counter-regulated by induction of T-cell anergy/deletion, if this Th1-cell response is inhibited (experimentally by anti-IL-12), TGF beta-producing cells appear, and these are capable of active immune suppression. This reciprocal relationship between IFN-gamma production and TGF-beta production is further supported in mouse models of mucosal inflammation. Thus, in the TNBS-colitis model, there is direct stimulation of the immune cells in the lamina propria as a result of diffuse haptenization of mucosal proteins, which leads to a massive Th1-cell response capable of overwhelming any suppressive counter-regulatory mechanisms normally generated in the PPs. This highly polarized Th1-cell response is controlled only by direct abrogation of IL-12 production with exogenous administration of anti-IL-12, or with indirect inhibition of this response via induction of oral tolerance and accompanying production of TGF-beta (Refs 6-8). The data obtained from this model are consistent with those obtained with another model of intestinal inflammation--inflammation in severe combined immunodeficiency (SCID) mice following lymphoid repletion with CD45Rbhi (naive) T cells. In this model, inflammation is again mediated by Th1 cells and is prevented by co-repletion with CD45Rbhi (memory) T cells, which appear to work by secreting TGF-beta (Refs 9, 10). Thus, a common feature of the various experimental models of intestinal inflammation studied to date is the Yin-Yang relationship of IFN-gamma and TGF-beta, with the former being proinflammatory and the latter anti-inflammatory. Is the IFN-gamma TGF-beta dichotomy that is evident both in the normal state and in models of inflammation simply a reflection of an underlying Th1 Th2 dichotomy? The answer to this important question is not yet known. Thus, while it is clear from the in vitro studies already discussed that IL-12 and/or IFN-gamma inhibit TGF-beta production, the role of IL-4 in this process is more elusive. These in vitro studies

Topics: Animals; Colitis; Humans; Interferon-gamma; Intestinal Mucosa; Transforming Growth Factor beta

The most effective way to control severity and mortality rate of the novel coronavirus disease (COVID-19) is through sensitive diagnostic approaches and an appropriate treatment protocol. We aimed to identify the effect of adding corticosteroid and Tocilizumab to a standard treatment protocol in treating COVID-19 patients with chronic disease through hematological and lab biomarkers.. This study was performed retrospectively on 68 COVID-19 patients with chronic disease who were treated by different therapeutic protocols. The patients were categorized into four groups: control group represented the patients' lab results at admission before treatment protocols were applied; group 1 included patients treated with anticoagulants, Hydroxychloroquine, and antibiotics; group 2 comprised patients treated with Dexamethasone; and group 3 included patients treated with Dexamethasone and Tocilizumab.. The study paves the way into the effectiveness of combining Dexamethasone with Tocilizumab in treatment COVID-19 patients with chronic diseases.. La forma más eficaz de controlar la gravedad y la tasa de mortalidad de la enfermedad del nuevo coronavirus (COVID-19) es mediante enfoques de diagnóstico sensibles y un protocolo de tratamiento adecuado. Nuestro objetivo fue identificar el efecto de agregar corticosteroides y tocilizumab a un protocolo de tratamiento estándar en el tratamiento de pacientes con COVID-19 con enfermedad crónica a través de biomarcadores hematológicos y de laboratorio.. Este estudio se realizó de forma retrospectiva en 68 pacientes COVID-19 con enfermedad crónica que fueron tratados por diferentes protocolos terapéuticos. Los pacientes se clasificaron en cuatro grupos: el grupo de control representaba los resultados de laboratorio de los pacientes en el momento de la admisión antes de que se aplicaran los protocolos de tratamiento; el grupo 1 incluyó a pacientes tratados con anticoagulantes, hidroxicloroquina y antibióticos; el grupo 2 estaba compuesto por pacientes tratados con dexametasona; y el grupo 3 incluyó a pacientes tratados con dexametasona y tocilizumab.. El estudio allana el camino hacia la eficacia de la combinación de dexametasona con tocilizumab en el tratamiento de pacientes con COVID-19 con enfermedades crónicas.. The Child-Mother Index constitutes a potential useful risk factor indicator for statistical analyses on data after birth. The value of the Child-Mother Index based on the estimated fetal weight before birth deserves evaluation.. Six ceria supports synthesized by various synthesis methodologies were used to deposit cobalt oxide. The catalysts were thoroughly characterized, and their catalytic activity for complete methane oxidation was studied. The supports synthesized by direct calcination and precipitation with ammonia exhibited the best textural and structural properties as well as the highest degree of oxidation. The remaining supports presented poorer textural properties to be employed as catalytic supports. The cobalt deposited over the first two supports presented a good dispersion at the external surface, which induced a significant redox effect that increased the number of Co. Some studies show that children with obesity are more likely to receive a diagnosis of depression, anxiety, or attention-deficit hyperactivity disorder (ADHD). But this does not necessarily mean obesity causes these conditions. Depression, anxiety, or ADHD could cause obesity. A child's environment, including family income or their parents' mental health, could also affect a child's weight and mental health. Understanding the nature of these relationships could help scientists develop better interventions for both obesity and mental health conditions. Genetic studies may help scientists better understand the role of the environment in these conditions, but it's important to consider both the child's and their parents’ genetics in these analyses. This is because parents and children share not only genes, but also environmental conditions. For example, families that carry genetic variants associated with higher body weight might also have lower incomes, if parents have been affected by biases against heavier people in society and the workplace. Children in these families could have worse mental health because of effects of their parent’s weight, rather than their own weight. Looking at both child and adult genetics can help disentangle these processes. Hughes et al. show that a child's own body mass index, a ratio of weight and height, is not strongly associated with the child’s mental health symptoms. They analysed genetic, weight, and health survey data from about 41,000 8-year-old children and their parents. The results suggest that a child's own BMI does not have a large effect on their anxiety symptoms. There was also no clear evidence that a child's BMI affected their symptoms of depression or ADHD. These results contradict previous studies, which did not account for parental genetics. Hughes et al. suggest that, at least for eight-year-olds, factors linked with adult weight and which differ between families may be more critical to a child's mental health than a child’s own weight. For older children and adolescents, this may not be the case, and the individual’s own weight may be more important. As a result, policies designed to reduce obesity in mid-childhood are unlikely to greatly improve the mental health of children. On the other hand, policies targeting the environmental or societal factors contributing to higher body weights, bias against people with higher weights, and poor child mental health directly may be more beneficial.. The development of an efficient photocatalyst for C2 product formation from CO. Оценка антиастенического эффекта последовательной терапии левокарнитином (ЛК) и ацетилкарнитином (АЛК) пациентов с артериальной гипертензией и/или ишемической болезнью сердца (ИБС) с астеническим синдромом (АС).. В открытое сравнительное исследование были включены 120 пациентов в возрасте 54—67 лет с артериальной гипертензией и/или ИБС с АС. Пациенты 1-й группы (. У больных 1-й группы отмечено статистически значимое уменьшение различных проявлений АС. Отличия носили достоверный характер по сравнению как с исходным уровнем, так и со 2-й группой. Установлено эндотелийпротективное действие ЛК и АЛК.. Полученные результаты свидетельствуют, что у таких коморбидных пациентов использование ЛК и АЛК уменьшает выраженность проявлений АС, а установленные эндотелиотропные свойства препаратов позволяют рекомендовать их в составе комплексной персонифицированной терапии пациентов с сердечно-сосудистыми заболеваниями.. Naproxen sodium 440 mg/diphenhydramine 50 mg combination demonstrated improvement in sleep maintenance (WASO) vs. naproxen sodium 550 mg and higher efficiency in average daily pain reduction compared with the comparison groups. The treatment was well tolerated There were no serious or unexpected adverse events reported in the study.. Сравнительный анализ эффективности и безопасности новой комбинации напроксена натрия и дифенгидрамина у пациентов с неспецифическим болевым синдромом в пояснично-крестцовом отделе спины (M54.5 «Боль внизу спины») и нарушением сна (G47.0 «Нарушения засыпания и поддержания сна [бессонница]»).. Проведено проспективное многоцентровое рандомизированное открытое сравнительное в параллельных группах клиническое исследование. Пациенты были рандомизированы в 3 группы. Больные 1-й группы получали напроксен натрия (440 мг) и дифенгидрамин (50 мг), 2-й — напроксен натрия (550 мг), 3-й — парацетамол (1000 мг) и дифенгидрамин (50 мг). Исследуемые препараты пациенты принимали однократно перед сном в течение 3 дней. Все пациенты также принимали 275 мг (1 таблетка) напроксена натрия в качестве препарата фоновой терапии. Первичным критерием эффективности было общее время бодрствования после наступления сна (WASO), измеряемое методом актиграфии. Также использовались критерии оценки продолжительности и качества сна и выраженности боли.. Анализ эффективности проведен для ITT популяции (. Применение комбинации напроксена натрия (440 мг) и дифенгидрамина (50 мг) характеризовалось более выраженным поддержанием сна по сравнению с напроксеном натрия 550 мг и более высокой эффективностью в отношении снижения интенсивности боли по сравнению со 2-й и 3-й группами. Отмечена хорошая переносимость препарата, серьезных нежелательных явлений зарегистрировано не было.

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Patients suffering from inflammatory bowel diseases (IBDs) express increased mucosal levels of transforming growth factor (TGF)-β compared with non-IBD controls. SMAD7 negatively regulates TGF-β signaling. An earlier study aiming to target Smad7 showed a lack of clinical benefit. It remains unknown whether inhibition of SMAD7 is beneficial in specific settings of IBD. We evaluated the effect of Smad7 deficiency on inflammation, fibrogenesis, and wound healing.. For the initiation of fibrosis in Smad7-/- (Smad7Δex-I) CD-1 mice, the dextran sodium sulfate-induced chronic colitis model and the heterotopic transplantation model of fibrosis were used. Wound closure of fibroblasts from Smad7-/- mice was determined using culture inserts and electric cell-substrate impedance sensing in vitro.. In dextran sodium sulfate-induced chronic colitis, Smad7 deficiency was associated with ameliorated inflammation, as evidenced by decreased clinical score, histological score, and myeloperoxidase activity. Absence of SMAD7 decreased T-cell accumulation in colonic tissue and tumor necrosis factor (TNF) mRNA expression levels. Smad7-/- mice showed a significant increase in hydroxyproline and collagen content, as well as ColIVa1 mRNA expression. Wild type mice transplanted with terminal ileum from Smad7-/- mice in the heterotopic animal model for intestinal fibrosis showed a significant increase in collagen content and protein expression of α-smooth muscle actin.. Smad7 deficiency is associated with a decrease in intestinal inflammation and an increase in fibrosis. Targeting SMAD7 constitutes a potential new treatment option for IBD; progression of disease-associated fibrosis should be considered.. We evaluated the effect of Smad7 deficiency on inflammation and fibrogenesis. Smad7 deficiency was associated with ameliorated inflammation and increased collagen deposition. When targeting Smad7 as therapeutic strategy in IBD, potential initiation or aggravation of fibrosis should be considered.

Topics: Animals; Colitis; Collagen; Dextrans; Fibrosis; Inflammation; Mice; Mice, Inbred C57BL; RNA, Messenger; Smad7 Protein; Transforming Growth Factor beta

Gut fibrosis occurs under chronic inflammation. This study examined the effects of different cyclooxygenase (COX) inhibitors on fibrosis in the inflamed colon.. Colitis was induced by 2,4-dinitrobenzenesulfonic acid (DNBS) in albino male Sprague-Dawley rats. After 6, 12 and 18 days, macroscopic and microscopic damage, collagen and elastic fibre content were examined. At day 6, pro-fibrotic factors (collagen I and III, hydroxyproline, fibronectin, matrix metalloproteinase-2 and -9), transforming growth factor-beta (TGF-β) signalling [TGF-β, Ras homolog gene family member A (RhoA), phosphorylated small mother against decapentaplegic (pSMAD)-2 and -6] and peristalsis were assessed, and the effects of indomethacin, SC-560 or celecoxib were tested.. Six days after DNBS administration, significant histopathological signs of fibrotic remodelling were observed in rats. At day 6, pro-fibrotic factors were up-regulated and colonic peristalsis was altered. COX inhibitors reversed the histochemical, molecular and functional changes in the fibrotic colon. COX inhibition reduced TGF-β expression, SMAD2 phosphorylation and RhoA, and increased SMAD6 expression.. Colonic fibrosis is associated with altered bowel motility and induction of profibrotic factors driven by TGF-β signalling. COX-1 and COX-2 inhibition counteracts this fibrotic remodelling by the modulation of TGF-β/SMAD signalling, mainly via SMAD6 induction and reduction in SMAD2 phosphorylation.

Topics: Animals; Colitis; Collagen; Disease Models, Animal; Fibrosis; Male; Matrix Metalloproteinase 2; Prostaglandin-Endoperoxide Synthases; Rats; Rats, Sprague-Dawley; Smad Proteins; Transforming Growth Factor beta

Mesenchymal stem cell (MSC) therapy has been shown to have some therapeutic effects in rodent models and patients with IBD; however, its role in colon tumor models is controversial. In this study, the potential role and mechanisms of bone marrow-derived MSCs (BM-MSCs) in colitis-associated colon cancer (CAC) were investigated.. The CAC mouse model was established with azoxymethane (AOM) and dextran sulfate sodium (DSS). The mice were administered an intraperitoneal injection of MSCs once weekly for different periods. The progression of CAC and the cytokine expression in tissues was assessed. Immunofluorescence staining was used to detect MSCs localization. Levels of immune cells in the spleen and lamina propria of the colon were detected using flow cytometry. A co-culture of MSCs and naïve T cells was performed to determine the effect of MSCs on naïve T cell differentiation.. Early administration of MSCs inhibited the occurrence of CAC, while late administration promoted the progression of CAC. The inhibitory effect of early injection in mice was characterized by the expression of inflammatory cytokines in colon tissue was decreased, and induction of T regulatory cells (Tregs) infiltration via TGF-β. The promotive effect of late injection was characterized by a shift of T helper (Th) 1/Th2 immune balance toward a Th2 phenotype through IL-4 secretion. IL-12 can reverse this shift to Th2 accumulation in mice.. MSCs can curb the progression of colon cancer by inducing Treg accumulation via TGF-β at the early stage of inflammatory transformation but promote the progression of colon cancer by inducing a shift in Th1/Th2 immune balance to Th2 through IL-4 secretion at the late stage. And the immune balance of Th1/Th2 influenced by MSCs could be reversed by IL-12.

Topics: Animals; Colitis; Colon; Colonic Neoplasms; Cytokines; Dextran Sulfate; Disease Models, Animal; Interleukin-12; Interleukin-4; Mesenchymal Stem Cell Transplantation; Mesenchymal Stem Cells; Mice; Mice, Inbred C57BL; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Fibrosis development in ulcerative colitis is associated directly with the severity of mucosal inflammation, which increases the risk of colorectal cancer. The transforming growth factor-β (TGF-β) signaling pathway is an important source of tissue fibrogenesis, which is stimulated directly by reactive oxygen species produced from nicotinamide adenine dinucleotide phosphate oxidases (NOX). Among members of the NOX family, NOX4 expression is up-regulated in patients with fibrostenotic Crohn's disease (CD) and in dextran sulfate sodium (DSS)-induced murine colitis. The aim of this study was to determine whether NOX4 plays a role in fibrogenesis during inflammation in the colon using a mouse model.. Acute and recovery models of colonic inflammation were performed by DSS administration to newly generated Nox4. Nox4. Nox4 protects against injury and plays a crucial role in fibrogenesis in DSS-induced colitis through canonical TGF-β signaling regulation, highlighting a new treatment target.

Topics: Animals; Colitis; Dextran Sulfate; Fibrosis; Inflammation; Mice; NADPH Oxidase 4; Reactive Oxygen Species; Transforming Growth Factor beta

Topics: Animals; Carcinogenesis; Cell Transformation, Neoplastic; Colitis; Colitis-Associated Neoplasms; Dextran Sulfate; Inflammation; Intestinal Mucosa; Mice; Transforming Growth Factor beta; Transforming Growth Factor beta1

Intestinal fibrosis is a common complication that affects more than 50% of Crohn´s Disease (CD) patients. There is no pharmacological treatment against this complication, with surgery being the only option. Due to the unknown role of P2X7 in intestinal fibrosis, we aim to analyze the relevance of this receptor in CD complications. Surgical resections from CD and non-Inflammatory Bowel Disease (IBD) patients were obtained. Intestinal fibrosis was induced with two different murine models: heterotopic transplant model and chronic-DSS colitis in wild-type and P2X7-/- mice. Human small intestine fibroblasts (HSIFs) were transfected with an siRNA against P2X7 and treated with TGF-β. A gene and protein expression of P2X7 receptor was significantly increased in CD compared to non-IBD patients. The lack of P2X7 in mice provoked an enhanced collagen deposition and increased expression of several profibrotic markers in both murine models of intestinal fibrosis. Furthermore, P2X7-/- mice exhibited a higher expression of proinflammatory cytokines and a lower expression of M2 macrophage markers. Moreover, the transient silencing of the P2X7 receptor in HSIFs significantly induced the expression of Col1a1 and potentiated the expression of Col4 and Col5a1 after TGF-β treatment. P2X7 regulates collagen expression in human intestinal fibroblasts, while the lack of this receptor aggravates intestinal fibrosis.

Topics: Animals; Colitis; Collagen; Crohn Disease; Fibroblasts; Humans; Intestines; Mice; Receptors, Purinergic P2X7; Transforming Growth Factor beta

Mesenchymal stem cells (MSCs) show promising therapeutic potential in treating inflammatory bowel disease (IBD), and intraperitoneal delivery of MSCs have become a more effective route for IBD treatment. However, the underlying mechanisms are still poorly understood. Here, we found that intraperitoneally delivered MSCs significantly alleviated experimental colitis. Depletion of peritoneal B cells, but not macrophages, clearly impaired the therapeutic effects of MSCs. Intraperitoneally delivered MSCs improved IBD likely by boosting the IL-10-producing B cells in the peritoneal cavity, and a single intraperitoneal injection of MSCs could significantly prevent disease severity in a recurrent mouse colitis model, with lower proinflammation cytokines and high level of IL-10. The gene expression profile revealed that thrombospondin-1 (THBS1) was dramatically upregulated in MSCs after coculture with peritoneal lavage fluid from colitis mice. Knockout of THBS1 expression in MSCs abolished their therapeutic effects in colitis and the induction of IL-10-producing B cells. Mechanistically, THBS1 modulates the activation of transforming growth factor-β (TGF-β), which combines with TGF-β receptors on B cells and contributes to IL-10 production. Blocking the interaction between THBS1 and latent TGF-β or inhibiting TGF-β receptors (TGF-βR) significantly reversed the THBS1-mediated induction of IL-10-producing B cells and the therapeutic effects on colitis. Collectively, our study revealed that intraperitoneally delivered MSCs secreted THBS1 to boost IL-10

Topics: Animals; B-Lymphocytes, Regulatory; Colitis; Dextran Sulfate; Disease Models, Animal; Inflammatory Bowel Diseases; Interleukin-10; Mesenchymal Stem Cells; Mice; Mice, Knockout; Receptors, Transforming Growth Factor beta; Transforming Growth Factor beta

Crohn's disease (CD) and ulcerative colitis (UC) are chronic inflammatory disorders of the gastrointestinal tract. Chronic inflammation is the main factor leading to intestinal fibrosis, resulting in recurrent stenosis, especially in CD patients. Currently, the underlying molecular mechanisms of fibrosis are still unclear. ZNF281 is a zinc-finger transcriptional regulator that has been characterized as an epithelial-to-mesenchymal transition (EMT)-inducing transcription factor, suggesting its involvement in the regulation of pluripotency, stemness, and cancer. The aim of this study is to investigate in vivo and in vitro the role of ZNF281 in intestinal fibrogenesis. Intestinal fibrosis was studied in vivo in C57BL/6J mice with chronic colitis induced by two or three cycles of administration of dextran sulfate sodium (DSS). The contribution of ZNF281 to gut fibrosis was studied in vitro in the human colon fibroblast cell line CCD-18Co, activated by the pro-fibrotic cytokine TGFβ1. ZNF281 was downregulated by siRNA transfection, and RNA-sequencing was performed to identify genes regulated by TGFβ1 in activated colon fibroblasts via ZNF281. Results showed a marked increase of ZNF281 in in vivo murine fibrotic colon as well as in in vitro human colon fibroblasts activated by TGFβ1. Moreover, abrogation of ZNF281 in TGFβ1-treated fibroblasts affected the expression of genes belonging to specific pathways linked to fibroblast activation and differentiation into myofibroblasts. We demonstrated that ZNF281 is a key regulator of colon fibroblast activation and myofibroblast differentiation upon fibrotic stimuli by transcriptionally controlling extracellular matrix (ECM) composition, remodeling, and cell contraction, highlighting a new role in the onset and progression of gut fibrosis.

Topics: Animals; Colitis; Colon; Crohn Disease; Dextran Sulfate; Fibroblasts; Fibrosis; Humans; Mice; Mice, Inbred C57BL; Repressor Proteins; RNA, Small Interfering; Transcription Factors; Transforming Growth Factor beta; Zinc

Topics: Amino Acids; Animals; Brain Neoplasms; Brain-Gut Axis; Carcinogenesis; Colitis; Colon; Cytokines; Dextran Sulfate; Disease Models, Animal; Hippocampus; Inflammation; Inflammatory Bowel Diseases; Interleukin-6; Lipids; Mice; Mice, Inbred C57BL; Neurogenesis; Sulfates; Thiamine; Transforming Growth Factor beta

The mechanism of the recovery of immune inflammation in the intestine remains to be investigated. The calcitonin-related protein (CGRP; neuropeptide) has immune regulatory capacity. We observed that lower levels of CGRP were found in the colon biopsies of UC patients. CGRP were negatively correlated to TNF-α, IL-1β and IFN-γ in biopsy samples. The levels of TGF-β were lower in the UC group than that of the normal control (NC) group, which were positively correlated with the CGRP levels. Blocking CGRP significantly delayed recovery from colitis inflammation. CGRP induced the TGF-β-expressing CD4

Topics: Animals; Calcitonin Gene-Related Peptide; Colitis; Inflammation; Intestines; Macrophages; Mice; Transforming Growth Factor beta

Inflammatory bowel disease (IBD), a progressive and unpredictable colorectal inflammatory disease, is a global health problem. Currently, therapeutic strategies for the management of the disease are limited. Results from our previous studies indicated that probiotic Lactobacillus plantarum exhibits therapeutic effects against IBD, and through screening, we obtained an active 61-amino-acid long protein, L. plantarum membrane protein 1 (LpMP-1). Based on druggability-guided strategies, the search for LpMPs with lower molecular weights and better bioactivities contributes to the development of new anti-inflammatory agents to overcome the limitations of existing therapies against IBD. We used amino-acid-truncation strategies to obtain modified LpMPs (LpMP-2 - LpMP-9) using LpMP-1 as the parent template. Furthermore, we systematically evaluated the anti-colitis pharmacodynamics of these LpMPs in terms of symptomatology, histopathology, and cytokine levels in DSS-induced ulcerative colitis mice. Their possible targets of action against IBD was investigated under an iTRAQ-based pharmacoproteomic system and a docking-guided receptor-ligand relationship frame. We found a new active protein, LpMP-8, which had a lower molecular weight than LpMP-1. LpMP-8 was found to exhibit anti-colitis activity following oral administration in vivo (50 μg/kg) by improving symptoms of colitis, colonic ulcerations, and cytokine disorders. TLRs and TGF-β were found to be involved in the action of LpMP-8 against colitis; LpMP-8 was to compete with TLR4-MD2-bound LPS and reverse TGF-β and Smad2/7 disorders. Our probiotic-derived LpMP-8 was shown to elicit oral anti-colitis activity, and its significant efficacy is probably associated with TLR4 and TGF-β.

Topics: Animals; Anti-Inflammatory Agents; Colitis; Colonic Diseases; Cytokines; Dextran Sulfate; Inflammatory Bowel Diseases; Lactobacillus plantarum; Membrane Proteins; Mice; Toll-Like Receptor 4; Transforming Growth Factor beta

All-trans retinoic acid (ATRA) is a biologically active isomer of retinoic acid (RA). Topical ATRA (retin-a, retin-a micro, atralin, renova, and avita) is the active pharmaceutical ingredient for FDA-approved treatments for acne and skin wrinkles. Oral formulations (Vesanoid) treat acute promyelocytic leukemia, but oral dosing can induce severe side effects. Despite benefits in various rodent models of inflammatory bowel disease (IBD), toxicity and controversial clinical observations have diminished enthusiasm for ATRA IBD clinical trials. To circumvent these issues and to use ATRA's key role in maintaining gut tolerance, we developed a poly(lactic-co-glycolic acid) (PLGA) microsphere (MS) encapsulated ATRA formulation aimed at directing ATRA delivery to immune structures of the gut, limiting systemic exposure. Initially, ATRA MS was developed as a component of a combinatorial product (TreXTAM) that also contained encapsulated transforming growth factor (TGF)-β and ATRA in a 1:2 w/w ratio. Although the combination was optimal, benefit was also observed when ATRA MS was given alone in the CD4+ CD25-T-cell adoptive transfer (ACT) colitis model.. We used the ACT and DSS-induced murine models of colitis to expand on the dose-dependent effects of oral ATRA MS when given alone. The DSS model was also used to compare the efficacy of ATRA MS and soluble ATRA, while healthy animals were used to compare the pharmacokinetics of the two drugs.. In both the ACT and DSS-induced murine models of colitis, ATRA MS was observed to be effective in ameliorating disease. ATRA MS was also observed to be more effective than soluble ATRA in these models and displayed more favorable pharmacokinetics.. We suggest ATRA MS, as a standalone product, may attenuate IBD and perhaps limit fibrosis, while limiting systemic side effects.

Topics: Animals; Colitis; Inflammatory Bowel Diseases; Mice; Rodentia; Transforming Growth Factor beta; Tretinoin

Topics: Animals; Cell Differentiation; Colitis; Lactoferrin; Lymphocyte Activation; Mice, Inbred BALB C; Receptors, Transforming Growth Factor beta; Signal Transduction; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Colonization by gut microbiota in early life confers beneficial effects on immunity throughout the host's lifespan. We sought to elucidate the mechanisms whereby neonatal supplementation with p40, a probiotic functional factor, reprograms intestinal epithelial cells for protection against adult-onset intestinal inflammation.. p40 was used to treat young adult mouse colonic (YAMC) epithelial cells with and without deletion of a methyltransferase, su(var)3-9, enhancer-of-zeste and trithorax domain-containing 1β (Setd1β), and mice in early life or in adulthood. Anti-transforming growth factor β (TGFβ)-neutralizing antibodies were administered to adult mice with and without colitis induced by 2,4,6-trinitrobenzenesulfonic acid or dextran sulfate sodium. We examined Setd1b and Tgfb gene expression, TGFβ production, monomethylation and trimethylation of histone H3 on the lysine 4 residue (H3K4me1/3), H3K4me3 enrichment in Tgfb promoter, differentiation of regulatory T cells (Tregs), and the inflammatory status.. p40 up-regulated expression of Setd1b in YAMC cells. Accordingly, p40 enhanced H3K4me1/3 in YAMC cells in a Setd1β-dependent manner. p40-regulated Setd1β mediated programming the TGFβ locus into a transcriptionally permissive chromatin state and promoting TGFβ production in YAMC. Furthermore, transient exposure to p40 during the neonatal period and in adulthood resulted in the immediate increase in Tgfb gene expression. However, only neonatal p40 supplementation induced the sustained H3K4me1/3 and Tgfb gene expression that persisted into adulthood. Interfering with TGFβ function by neutralizing antibodies diminished the long-lasting effects of neonatal p40 supplementation on differentiation of Tregs and protection against colitis in adult mice.. Exposure to p40 in early life enables an epigenetic imprint on TGFβ, leading to long-lasting production of TGFβ by intestinal epithelial cells to expand Tregs and protect the gut against inflammation.

Topics: Animals; Colitis; Epigenesis, Genetic; Epithelial Cells; Female; Inflammation; Intestinal Mucosa; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Transgenic; Pregnancy; Prenatal Exposure Delayed Effects; Probiotics; Transforming Growth Factor beta

Inflammatory bowel disease (IBD) is a set of immunological disorders which can generate chronic pain and fatigue associated with the inflammatory symptoms. The treatment of IBD remains a significant hurdle with current therapies being only partially effective or having significant side effects, suggesting that new therapies that elicit different modes of action and delivery strategies are required. TGM1 is a TGF-β mimic that was discovered from the intestinal helminth parasite Heligmosomoides polygyrus and is thought to be produced by the parasite to suppress the intestinal inflammation response to help evade host immunity, making it an ideal candidate to be developed as a novel anti-inflammatory bio-therapeutic. Here we utilized the expression system of the edible green algae Chlamydomonas reinhardtii in order to recombinantly produce active TGM1 in a form that could be ingested. C. reinhardtii robustly expressed TGM1, and the resultant recombinant protein is biologically active as measured by regulatory T cell induction. When delivered orally to mice, the algal expressed TGM1 is able to ameliorate weight loss, lymphadenopathy, and disease symptoms in a mouse model of DSS-induced colitis, demonstrating the potential of this biologic as a novel treatment of IBD.

Topics: Administration, Oral; Animals; Chlamydomonas reinhardtii; Colitis; Disease Models, Animal; Mice; Nematospiroides dubius; Recombinant Proteins; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Inflammatory bowel disease (IBD) is a multifactorial intestinal disorder characterized by chronic intestinal inflammation. The etiology of IBD is still unclear, although genetic, environmental and host factors have been associated to the disease. Extra-virgin olive oil (EVO) is a central component of the Mediterranean diet and it decreases chronic inflammation by interfering with arachidonic acid and NF-κB signaling pathways. Specifically, the different components of EVO are able to confer advantages in terms of health in their site of action. For instance, oleic acid displays a protective effect in liver dysfunction and gut inflammation, whereas phenolic compounds protect colon cells against oxidative damage and improve the symptoms of chronic inflammation in IBD. Given the biological properties of EVO, we investigated whether its administration is able to confer protection in a mouse model of dextrane sodium sulfate (DSS)-induced colitis. Four EVO cultivars from the Apulian Region of Italy, namely Ogliarola (Cima di Bitonto), Coratina, Peranzana and Cima di Mola, respectively, were used. Administration of EVO resulted in reduced body weight loss in our colitis model. Furthermore, mice treated with Ogliarola, Coratina and Cima di Mola EVO displayed a reduction of rectal bleeding and IL-1β, TGFβ, IL-6 gene expression levels. Furthermore, Ogliarola, Coratina and Peranzana EVO administration ameliorated intestinal permeability and histopathological features of inflammation. Our data further validate the well-known positive effects of EVO supplementation in promoting human health and suggest the bona fide contribution of EVO in preventing onset and reducing progression of intestinal inflammation.

Topics: Administration, Oral; Animals; Body Weight; Colitis; Dextran Sulfate; Diet, Mediterranean; Dietary Supplements; Disease Models, Animal; Gene Expression; Inflammation; Interleukin-1beta; Interleukin-6; Intestinal Mucosa; Italy; Male; Mice, Inbred C57BL; Olive Oil; Permeability; Transforming Growth Factor beta

Inflammatory bowel disease (IBD) is an immunologically mediated chronic intestinal disorder. Growth hormone (GH) administration enhances mucosal repair and decreases intestinal fibrosis in patients with IBD. In the present study, we investigated the effect of cellular sensitivity to GH via suppressor of cytokine signaling 2 (SOCS2) deletion on colitis and recovery. To induce colitis, wild type and SOCS2 knockout (SOCS2-/-) mice were treated with 3% dextran sodium sulphate (DSS), followed by a recovery period. SOCS2-/- mice showed higher disease activity during colitis with increased mRNA expression of the pro-inflammatory cytokines nitric oxide synthase 2 (NOS2) and interleukin 1 β (IL1-β). At recovery time point, SOCS2-/- showed better recovery with less fibrosis measured by levels of α-SMA and collagen deposition. Protein and mRNA expressions of transforming growth factor beta β1 (TGF-β1) receptors were significantly lower in SOCS2-/- mice compared to wild-type littermates. Using an in vivo bromodeoxyuridine (BrdU) proliferation assay, SOCS2-/- mice showed higher intestinal epithelial proliferation compared to wild-type mice. Our results demonstrated that deletion of the SOCS2 protein results in higher growth hormone sensitivity associated with higher pro-inflammatory signaling; however, it resulted in less tissue damage with less fibrotic lesions and higher epithelial proliferation, which are markers of GH-protective effects in IBD. This suggests a pleiotropic effect of SOCS2 and multiple cellular targets. Further study is required to study role of SOCS2 in regulation of TGFβ-mothers against the decapentaplegic homolog (Smad) pathway.

Topics: Animals; Biomarkers; Colitis; Cytokines; Dextran Sulfate; Disease Models, Animal; Disease Susceptibility; Fibrosis; Gene Deletion; Humans; Inflammatory Bowel Diseases; Intestinal Mucosa; Male; Mice; Mice, Knockout; Signal Transduction; Suppressor of Cytokine Signaling Proteins; Transforming Growth Factor beta

IL-22 has dual functions during tumorigenesis. Short term IL-22 production protects against genotoxic stress, whereas uncontrolled IL-22 activity promotes tumor growth; therefore, tight regulation of IL-22 is essential. TGF-β1 promotes the differentiation of Th17 cells, which are known to be a major source of IL-22, but the effect of TGF-β signaling on the production of IL-22 in CD4+ T cells is controversial. Here we show an increased presence of IL-17+IL-22+ cells and TGF-β1 in colorectal cancer compared to normal adjacent tissue, whereas the frequency of IL-22 single producing cells is not changed. Accordingly, TGF-β signaling in CD4+ T cells (specifically Th17 cells) promotes the emergence of IL-22-producing Th17 cells and thereby tumorigenesis in mice. IL-22 single producing T cells, however, are not dependent on TGF-β signaling. We show that TGF-β, via AhR induction, and PI3K signaling promotes IL-22 production in Th17 cells.

Topics: Animals; Basic Helix-Loop-Helix Transcription Factors; Carcinogenesis; Cell Differentiation; Colitis; Colonic Neoplasms; Colorectal Neoplasms; Disease Models, Animal; Female; Humans; Interleukin-17; Interleukin-22; Interleukins; Lymphocytes, Tumor-Infiltrating; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Phosphatidylinositol 3-Kinases; Receptors, Aryl Hydrocarbon; Signal Transduction; Th17 Cells; Transforming Growth Factor beta; Transforming Growth Factor beta1

Mice deficient in the transcription factor Runx3 develop a multitude of immune system defects, including early onset colitis. This paper demonstrates that Runx3 is expressed in colonic mononuclear phagocytes (MNP), including resident macrophages (RM) and dendritic cell subsets (cDC2). Runx3 deletion in MNP causes early onset colitis due to their impaired maturation. Mechanistically, the resulting MNP subset imbalance leads to up-regulation of pro-inflammatory genes as occurs in IL10R-deficient RM. In addition, RM and cDC2 display a marked decrease in expression of anti-inflammatory/TGF β-regulated genes and β-catenin signaling associated genes, respectively. MNP transcriptome and ChIP-seq data analysis suggest that a significant fraction of genes affected by Runx3 loss are direct Runx3 targets. Collectively, Runx3 imposes intestinal immune tolerance by regulating maturation of colonic anti-inflammatory MNP, befitting the identification of RUNX3 as a genome-wide associated risk gene for various immune-related diseases in humans, including gastrointestinal tract diseases such as Crohn's disease and celiac.

Topics: Animals; beta Catenin; Cell Differentiation; Colitis; Colon; Core Binding Factor Alpha 3 Subunit; Disease Models, Animal; Humans; Mice; Mononuclear Phagocyte System; Receptors, Interleukin-10; Signal Transduction; Transforming Growth Factor beta; Up-Regulation

Eleutherine palmifolia (L.) Merr. extract (EPE) containing isoliquiritigenin and oxyresveratrol is believed to be an anticancer agent. This study evaluates colon histopathology, TNF-α, TGF-β, and hepatotoxicity on BALB/c mice colitis-associated colon cancer (CAC) model treated with EPE.. In vivo study was performed on BALB/c mice CAC model induced by 10 mg/kgBW AOM on the first day followed by administration that each cycle consisted of 5% DSS in water for seven days and regular water for seven days. The indicators of the formation of CAC were observed by a fecal occult blood test (FOBT) and serum amyloid α (SAA) test. The treatment was conducted once a week started from the seventh week up to the twentieth week with six treatment groups: I was administrated by regular water only (negative control), II was administrated by AOM and DSS only (positive control), III was administrated by doxorubicin,  IV-VI were treated by EPE (0.25 mg/kg BW, 0.50 mg/kg BW, and 1.00 mg/kg BW) respectively. The colon and liver's histopathology was observed using hematoxylin-eosin (HE) staining, TNF-α with immunohistochemistry (IHC), and level measurement of TGF-β colon with ELISA reader. The data were used one-way ANOVA followed by post hoc as statistical analysis.. The administration of EPE increased the expression of TNF-α, the total of goblet cells of the colon, and decreased the level of TGF-β. Administration of EPE 0.50 mg/20g BW decreased a liver histopathological score but induced a histopathological alteration of the liver at a dose of 1.00 mg/20g BW.. This study indicate that EPE could be recommended as a colon anticancer through increase the goblet cells, induce apoptosis through increase TNF-α, and decrease TGF-β.

Topics: Animals; Colitis; Colitis-Associated Neoplasms; Dextran Sulfate; Female; Goblet Cells; Iridaceae; Mice; Mice, Inbred BALB C; Plant Extracts; Toxicity Tests; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

Sesquiterpene lactones are organic compounds derived mainly from plants that exhibit anti-inflammatory and antitumor activities being one of the key mechanism of action of NF-kB pathway and synthesis of cytokines such as IL-1 and TNF- α.. The overall objective of the present study was to evaluate the anti-inflammatory action of a sesquiterpene lactone diacethylpiptocarphol (DPC) from Vernonia scorpioides (Lam.) Pers. and parthenolide (PTH) in Balb-c mice with DSS-induced colitis.. The anti-inflammatory effects of Intraperitonial administration of DPC (5 mg/kg/day) were evaluated in Balb/c mice with DSS-induced colitis, and further the body weight measurement, TNF-α and TGF-β level was determined.. After intraperitoneal treatment for one week, DSS-induced colitis was significantly reduced in mice treated with either of both sesquiterpenes lactones, as witnessed by reduced cellular infiltration, tissue damage, TNF-α production, and enhanced production of TGF-β.. Sesquiterpene lactone DPC, isolated from Vernonia scorpioides showed anti-inflammatory activity, in this experimental model of colitis the sesquiterpene lactones DPC and PTH exhibit equal anti-inflammatory activity.

Topics: Animals; Anti-Inflammatory Agents; Colitis; Colon; Dextran Sulfate; Flowers; Injections, Intraperitoneal; Lactones; Male; Mice, Inbred BALB C; Plant Leaves; Sesquiterpenes; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha; Vernonia

Regulatory T (T reg) cells are required for the maintenance of immune homeostasis. Both TGF-β signaling and epigenetic modifications are important for

Topics: Animals; CCAAT-Enhancer-Binding Proteins; Cell Differentiation; Colitis; DNA Methylation; Epigenesis, Genetic; Forkhead Transcription Factors; Gene Expression Profiling; Lymphocyte Activation; Lymphocyte Count; Mice; Mice, Knockout; Phosphorylation; Proteolysis; Signal Transduction; T-Lymphocytes, Regulatory; Transcriptome; Transforming Growth Factor beta; Ubiquitin-Protein Ligases

Regulatory T (Treg) cells play an essential role in maintaining immune homeostasis, but the suppressive function of Treg cells can be an obstacle in the treatment of cancer and chronic infectious diseases. Here, we identified the homeobox protein Hhex as a negative regulator of Treg cells. The expression of Hhex was lower in Treg cells than in conventional T (Tconv) cells. Hhex expression was repressed in Treg cells by TGF-β/Smad3 signaling. Retroviral overexpression of Hhex inhibited the differentiation of induced Treg (iTreg) cells and the stability of thymic Treg (tTreg) cells by significantly reducing Foxp3 expression. Moreover, Hhex-overexpressing Treg cells lost their immunosuppressive activity and failed to prevent colitis in a mouse model of inflammatory bowel disease (IBD).

Topics: Animals; Cell Differentiation; Colitis; CTLA-4 Antigen; Disease Models, Animal; Forkhead Transcription Factors; Gene Expression Regulation; Homeodomain Proteins; Inflammatory Bowel Diseases; Interleukin-2 Receptor alpha Subunit; Mice; Signal Transduction; Skin; Smad3 Protein; T-Lymphocytes, Regulatory; Transcription Factors; Transforming Growth Factor beta

Conjugated linoleic acid (CLA) has been shown to activate the nuclear receptor PPAR-γ and modulate metabolic and immune functions. Despite the worldwide use of CLA dietary supplementation, strong scientific evidence for its proposed beneficial actions are missing. We found that CLA-supplemented diet reduced mucosal damage and inflammatory infiltrate in the dextran sodium sulfate (DSS)-induced colitis model. Conditional deletion of PPAR-γ in macrophages from mice supplemented with CLA diet resulted in loss of this protective effect of CLA, suggesting a PPAR-γ-dependent mechanism mediated by macrophages. However, CLA supplementation significantly worsened colorectal tumor formation induced by azoxymethane and DSS by inducing macrophage and T-cell-producing TGF-β via PPAR-γ activation. Accordingly, either macrophage-specific deletion of PPAR-γ or in vivo neutralization of latency-associated peptide (LAP, a membrane-bound TGF-β)-expressing cells abrogated the protumorigenic effect of CLA. Thus, the anti-inflammatory properties of CLA are associated with prevention of colitis but also with development of colorectal cancer.

Topics: Aminosalicylic Acid; Animals; Carcinogenesis; Cells, Cultured; Colitis; Colorectal Neoplasms; Dextran Sulfate; Dietary Supplements; Disease Models, Animal; Female; Humans; Inflammatory Bowel Diseases; Linoleic Acids, Conjugated; Macrophages; Mice; Mice, Inbred C57BL; Mice, Knockout; PPAR gamma; T-Lymphocytes; Transforming Growth Factor beta

There is a major unmet clinical need to identify pathways in inflammatory bowel disease (IBD) to classify patient disease activity, stratify patients that will benefit from targeted therapies such as anti-tumor necrosis factor (TNF), and identify new therapeutic targets. In this study, we conducted global transcriptome analysis to identify IBD-related pathways using colon biopsies, which highlighted the coagulation gene pathway as one of the most enriched gene sets in patients with IBD. Using this gene-network analysis across 14 independent cohorts and 1800 intestinal biopsies, we found that, among the coagulation pathway genes, plasminogen activator inhibitor-1 (PAI-1) expression was highly enriched in active disease and in patients with IBD who did not respond to anti-TNF biologic therapy and that PAI-1 is a key link between the epithelium and inflammation. Functionally, PAI-1 and its direct target, the fibrinolytic protease tissue plasminogen activator (tPA), played an important role in regulating intestinal inflammation. Intestinal epithelial cells produced tPA, which was protective against chemical and mechanical-mediated colonic injury in mice. In contrast, PAI-1 exacerbated mucosal damage by blocking tPA-mediated cleavage and activation of anti-inflammatory TGF-β, whereas the inhibition of PAI-1 reduced both mucosal damage and inflammation. This study identifies an immune-coagulation gene axis in IBD where elevated PAI-1 may contribute to more aggressive disease.

Topics: Animals; Biological Factors; Blood Coagulation; Cell Proliferation; Citrobacter; Colitis; Colon; Cytokines; Inflammation; Inflammatory Bowel Diseases; Interleukin-17; Intestinal Mucosa; Mice; Plasminogen Activator Inhibitor 1; Severity of Illness Index; Small Molecule Libraries; Th17 Cells; Tissue Plasminogen Activator; Transcription, Genetic; Transforming Growth Factor beta

The clearance of apoptotic cells is an essential process to maintain homeostasis of immune system, which is regulated by immunoregulatory cytokines such as TGFβ. We show here that Extracellular Vesicles (EVs) were highly released from apoptotic cells, and contributed to macrophage production of TGFβ in vitro and in vivo. We further elucidated mechanistically that phosphatidylserine in EVs was a key triggering-factor, and transcription factor FOXO3 was a critical mediator for apoptotic EV-induced TGFβ in macrophages. Importantly, we found that macrophages pre-exposed to EVs exhibited an anti-inflammatory phenotype. More strikingly, administration of EVs in vivo promotes Tregs differentiation and suppresses Th1 cell response, and ameliorates experimental colitis. Thus, apoptotic-EV-based treatment might be a promising therapeutic approach for human autoimmune disease.

Topics: Animals; Apoptosis; Colitis; Disease Models, Animal; Extracellular Vesicles; Forkhead Box Protein O3; Gamma Rays; Homeodomain Proteins; Humans; Jurkat Cells; Macrophages; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Phosphatidylserines; RNA Interference; RNA, Small Interfering; Thymocytes; Transforming Growth Factor beta

TL1A was reported to contribute to the susceptibility to ulcerative colitis (UC). However, the molecular mechanisms of TL1A in UC development are poorly understood. We aimed to investigate the role of TL1A in colitis, and reveal the regulatory mechanism of TL1A in chronic colitis development.. Wild-type mice and transgenic mice with overexpressing TL1A in lymphocytes were used to construct chronic DSS colitis models. To investigate the molecular mechanism in vitro, CD4. The elevated TL1A expression in chronic DSS colitis models exacerbated intestinal inflammation. The differentiation of Th9 cells, IL-9 secretion and production of TGF-β, IL-4 and PU.1 was significantly enhanced in transgenic mice with TL1A overexpression. In vitro results showed that TL1A enhanced the Th9 cells, IL-9 and PU.1 production, while TL1A antibodies inhibited their production. In human translational studies, patients with ulcerative colitis with elevated TL1A expression also exhibited more serious inflammation with higher levels of Th9 cells, IL-9 and PU.1 expression.. We presented a possible mechanism of TL1A in UC development that TL1A may promote the differentiation of Th9 cells and enhanced IL-9 secretion by up-regulating the expression of TGF-β, IL-4 and PU.1, which provided a novel perspective to study the UC pathogenesis, and indicated that targeting of TL1A signal pathway may by a likely strategy for the treatment of chronic colitis.

Topics: Animals; Cell Differentiation; Colitis; Cytokines; Glutathione; Interleukin-17; Interleukin-1beta; Interleukin-9; Intestinal Mucosa; Intestines; Lymphocyte Activation; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Signal Transduction; T-Lymphocyte Subsets; T-Lymphocytes; Transforming Growth Factor beta; Tumor Necrosis Factor Ligand Superfamily Member 15; Tumor Necrosis Factor-alpha

We isolated two novel probiotics strains (s193 and s292) from Funazushi, which is a traditional Japanese fermented food, and evaluated its effects on DSS-induced colitis to determine the possible underlying mechanisms.. A single colony from homogenized Funazushi was isolated by its ability to suppress TNF-α in RAW 264.7. Effect of probiotics on colonic inflammation induced by DSS was evaluated. Effect of probiotics on Treg induction by CD11c. Two novel probiotics strains classified into the genus Lactobacillus were isolated (s193 and s292), and those strains showed stronger anti-inflammatory effects on DSS-induced colitis than those of L. gasseri isolated from the gut. mRNA expression β8 integrin in CD11c. Strains s193 and s292 demonstrate strong anti-inflammatory effects on DSS-induced colitis through induction of β8 integrin expression on DCs. Our results suggested that Japanese traditional fermented foods are valuable sources for probiotics that are effective for IBD therapy and treatment.

Topics: Adoptive Transfer; Animals; Anti-Inflammatory Agents; CD11 Antigens; Colitis; Dendritic Cells; Dextran Sulfate; Female; Fermented Foods; Integrin alphaV; Integrin beta Chains; Japan; Mice; Mice, Inbred C57BL; Probiotics; RAW 264.7 Cells; RNA, Messenger; T-Lymphocytes, Regulatory; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

Inflammatory bowel disease (IBD) is an autoimmune disease with an abnormal and persistent immune response. Iguratimod, a novel anti-rheumatic drug, exhibits anti-inflammatory effects and regulates immune response. The role of iguratimod in intestinal mucosal inflammation and immunity has not been examined. The aim of this study was to investigate whether iguratimod ameliorates dextran sulphate sodium (DSS)-induced murine colitis and its potential regulatory mechanism. Murine colitis was induced by administering 2.5% DSS for 5days. Some mice were administered iguratimod (5, 30mg/kg) by oral gavage once daily for 7days, beginning on the day 3 after colitis induction. Our study showed that iguratimod alleviates the symptoms of colitis and suppresses intestinal tissue damage, including macroscopic and histopathological manifestations. Moreover, iguratimod reduced interleukin (IL)-6, IL-17, and tumour necrosis factor-α levels, and increased the expression levels of IL-10 and TGF-β. In addition, iguratimod downregulated the proportion of Th17 cells, the level of transcription factor retinoic acid-related orphan receptor γt (RORγt), and the phosphorylation of signal transducer and activator of transcription-3 (STAT3), and upregulated the proportion of Treg cells, the level of transcription factor forkhead box p3 (Foxp3), and the phosphorylation of STAT5 in the colonic tissues. In conclusion, iguratimod plays a protective role in mice with DSS-induced colitis via anti-inflammatory effects and regulation of Th17/Treg cells. Therefore, use of iguratimod may serve as a novel therapeutic strategy for the treatment of IBD.

Topics: Animals; Chromones; Colitis; Dextran Sulfate; Disease Models, Animal; Drug Evaluation, Preclinical; Gene Expression Regulation; Immunosuppressive Agents; Inflammatory Bowel Diseases; Interleukins; Intestinal Mucosa; Lymph Nodes; Male; Mice; Mice, Inbred BALB C; Molecular Structure; Random Allocation; Specific Pathogen-Free Organisms; Spleen; Sulfonamides; T-Lymphocytes, Regulatory; Th17 Cells; Transcription Factors; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

A concomitant action of multiple profibrotic mediators appears crucial in the development and progression of fibrosis. Sphingosine kinase/sphingosine 1 phosphate and transforming growth factor-β/Smads pathways are both involved in pathogenesis of fibrosis in several organs by controlling differentiation of fibroblasts to myofibroblasts and the epithelial to-mesenchymal transition. However, their direct involvement in chronic colitis-associated fibrosis it is not yet known. In this study we evaluated the immunohistochemical expression of some proteins implicated in sphingosine kinase/sphingosine 1 phosphate and transforming growth factor-β/Smads pathways in Dextrane Sodium Sulphate (DSS)-induced colorectal fibrosis in mice. Compared to control mice, DSS-induced chronic colitis mice developed a marked intestinal fibrosis associated with a concomitant overexpression of TGF-β, p-Smad3, α-SMA, collagen I-III, SPHK1, RhoA, PI3K, Akt, p-Akt, p-mTOR. This study highlights the relationship between the two pathways and the possible role of SPHK1 in the intestinal fibrosis.  These results, if confirmed by in vitro studies, may have important clinical implications in the development of new therapeutical approaches in inflammatory bowel disease.

Topics: Animals; Colitis; Disease Models, Animal; Fibrosis; Immunohistochemistry; Intestines; Lysophospholipids; Mice; Mice, Inbred C57BL; Phosphotransferases (Alcohol Group Acceptor); Reference Standards; Signal Transduction; Smad3 Protein; Sphingosine; Transforming Growth Factor beta

Neutrophils are the first responders to sites of inflammation when the intestinal epithelial barrier is breached and the gut microbiota invade. Despite current efforts in understanding the role of neutrophils in intestinal homeostasis, the complex interactions between neutrophils and intestinal epithelial cells (IECs) is still not well characterized. In this study, we demonstrated that neutrophils enhanced production of amphiregulin (AREG), a member of the EGFR ligand family, by IECs, which promoted IEC barrier function and tissue repair. Depletion of neutrophils resulted in more severe colitis in mice because of decreased AREG production by IECs upon dextran sodium sulfate (DSS) insult. Administration of AREG restored epithelial barrier function and ameliorated colitis. Furthermore, neutrophil-derived TGF-β promoted AREG production by IECs. Mechanistically, TGF-β activated MEK1/2 signaling, and inhibition of MEK1/2 abrogated TGF-β-induced AREG production by IECs. Collectively, these findings reveal that neutrophils play an important role in the maintenance of IEC barrier function and homeostasis.

Topics: Amphiregulin; Animals; Cells, Cultured; Colitis; Dextran Sulfate; Disease Models, Animal; Female; Homeostasis; Humans; Inflammatory Bowel Diseases; Intestinal Mucosa; Male; MAP Kinase Kinase 1; Mice; Mice, Inbred C57BL; Neutrophils; Signal Transduction; Transforming Growth Factor beta

Peripherally induced regulatory T (pT reg) cells play indispensable roles in regulating gut inflammation; however, the mechanism underling the differentiation of pT reg cells under inflammatory conditions remains largely unknown. Here, we show that the expression of Sox12, a member of SoxC family, is significantly induced in T reg cells in colitic mice. We also show that TCR-NFAT signaling induces Sox12 expression in CD4

Topics: Animals; Cell Differentiation; Colitis; Disease Models, Animal; Forkhead Transcription Factors; Mice; Mice, Inbred BALB C; Mice, Knockout; Signal Transduction; SOXC Transcription Factors; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Adipose tissue-derived stem cells (ASCs) have been investigated as therapeutic tools for a variety of autoimmune diseases, including inflammatory diseases. However, the mechanisms underlying the immunomodulatory properties of ASCs are not well understood. Here, we investigated the mechanism of regulatory T cell (Treg) induction in ASC therapy in a murine model of inflammatory bowel disease.. Acute colitis was induced in mice using dextran sulfate sodium and ASCs administered intraperitoneally. Tregs and CD103. Systemic administration of ASCs significantly lessened the clinical and histopathological severity of colitis. ASCs were distributed throughout the lymphatic system in the MLNs and spleen. Tregs were increased in the MLNs and CLP, but CD103. These results demonstrate that ASCs induce Tregs by activating latent TGF-β via TSP-1, independent of CD103

Topics: Adipose Tissue; Animals; Antigens, CD; Cell Movement; Colitis; Dendritic Cells; Dextran Sulfate; Disease Models, Animal; Female; Gene Knockdown Techniques; Integrin alpha Chains; Male; Mice; Mice, Inbred C57BL; Stem Cell Transplantation; Stem Cells; T-Lymphocytes, Regulatory; Thrombospondin 1; Transforming Growth Factor beta

Previous studies have indicated that regulatory T cells serve essential roles in maintaining intestinal homeostasis, however, the role of different Treg subsets in modulating inflammatory bowel disease has still not been addressed clearly. In the present study, the authors measured the percentage of Foxp3+ IL‑10+ TGF‑β+ natural Tregs, Foxp3‑ IL‑10+ TGF‑β‑ induced Tregs, CD127‑ induced Tregs and CD8+ Tregs at different time points in DSS‑induced experimental colitis model in murine lamina propria lymphocytes, mesenteric lymph node and peripheral blood. In addition, the authors compared the frequency of four Treg subsets in patients diagnosed of ulcerative colitis at different stages with enrolled healthy controls. The percentage of Foxp3+ IL‑10+ TGF‑β+ natural Tregs decreased in acute stage of both human and mice was observed, but proliferated significantly during remittent stage. Foxp3‑ IL‑10+ TGF‑β‑ inducible (i) Treg and CD127‑ iTreg was observed as being significantly decreased percentage in LPL at 4 and 7 days, the frequency of Foxp3‑ IL‑10+ TGF‑β‑ iTreg cells became decreased and CD127‑ iTreg only slightly increased at the chronic stage following DSS induction. However, the proportion of both Foxp3‑ IL‑10+ TGF‑β‑ iTreg and CD127‑ iTreg was nearly unchanged in human IBD. Although intestinal inflammation decreased the percentage of CD8+ Tregs, it remained lower in the remittent stage of human IBD. Only enhanced proliferation of lamina propria lymphocytes‑derived CD8+ Treg was reported at 7 days in dextran sodium sulfate‑induced murine colitis. The results demonstrated that Foxp3+ IL‑10+ TGF‑β+ natural Tregs may serve an essential role in exhibiting suppressive and protecting from immune‑related mucosal injury during chronic stage in inflammatory bowel disease.

Topics: Animals; Colitis; Dextran Sulfate; Forkhead Transcription Factors; Humans; Inflammatory Bowel Diseases; Interleukin-10; Interleukin-7 Receptor alpha Subunit; Male; Mice; Mice, Inbred C57BL; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

D-mannose, a C-2 epimer of glucose, exists naturally in many plants and fruits, and is found in human blood at concentrations less than one-fiftieth of that of glucose. However, although the roles of glucose in T cell metabolism, diabetes and obesity are well characterized, the function of D-mannose in T cell immune responses remains unknown. Here we show that supraphysiological levels of D-mannose safely achievable by drinking-water supplementation suppressed immunopathology in mouse models of autoimmune diabetes and airway inflammation, and increased the proportion of Foxp3

Topics: Adoptive Transfer; Animals; Colitis; Colon; Diabetes Mellitus, Type 1; Dietary Supplements; Disease Models, Animal; Fatty Acids; Flow Cytometry; Forkhead Transcription Factors; Humans; In Vitro Techniques; Inflammation; Integrins; Lipid Metabolism; Lung; Lung Diseases; Mannose; Mice; Mice, Inbred C57BL; Mice, Inbred NOD; Ovalbumin; Oxidation-Reduction; Pancreas; Reactive Oxygen Species; Real-Time Polymerase Chain Reaction; Respiratory Hypersensitivity; Reverse Transcriptase Polymerase Chain Reaction; Spleen; T-Lymphocytes, Regulatory; Transforming Growth Factor beta; Up-Regulation

CD4

Topics: Amodiaquine; Animals; Cell Proliferation; Colitis; DNA-Binding Proteins; Forkhead Transcription Factors; Interleukin-2; Interleukin-2 Receptor alpha Subunit; Male; Mice, Inbred C57BL; Mice, Knockout; Nerve Tissue Proteins; Nuclear Receptor Subfamily 4, Group A, Member 1; Nuclear Receptor Subfamily 4, Group A, Member 2; Receptors, Interleukin-2; Receptors, Steroid; Receptors, Thyroid Hormone; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Intestinal fibrosis is characterized by abnormal production and deposition of extracellular matrix (ECM) proteins by activated myofibroblasts. The main progenitor cells of activated myofibroblasts are the fibroblasts and the epithelial cells, the latter through the epithelial-mesenchymal transition (EMT).. To evaluate the action of the new PPAR-γ modulator, GED-0507-34 Levo (GED) on the expression of EMT associated and regulatory proteins such as TGF-β, Smad3, E-cadherin, Snail, ZEB1, β-catenin, and GSK-3β, in a mouse model of DSS-induced intestinal fibrosis.. Chronic colitis and fibrosis were induced by oral administration of 2.5% DSS (w/v) for 6 weeks. GW9662 (GW), a selective PPAR-γ inhibitor, was also administered by intraperitoneal injection at the dose of 1 mg/kg/day combined with GED treatment. All drugs were administered at the beginning of the second cycle of DSS (day 12). 65 mice were randomly divided into five groups (H2O as controls n = 10, H2O+GED n = 10, DSS n = 15, DSS+GED n = 15, DSS+GED+GW n = 15). The colon was excised for macroscopic examination and histological and morphometric analyses. The level of expression of molecules involved in EMT and fibrosis, like TGF-β, Smad3, E-cadherin, Snail, ZEB1, β-catenin, GSK-3β and PPAR-γ, was assessed by immunohistochemistry, immunofluorescence, western blot and Real Time PCR.. GED improved the DSS-induced chronic colitis and fibrosis. GED was able to reduce the expression of the main fibrosis markers (α-SMA, collagen I-III and fibronectin) as well as the pivotal pro-fibrotic molecules IL-13, TGF-β and Smad3, while it increased the anti-fibrotic PPAR-γ. All these GED effects were nullified by co-administration of GW with GED. Furthermore, GED was able to normalize the expression levels of E-cadherin and β-catenin and upregulated GSK-3β, that are all known to be involved both in EMT and fibrosis.. The DSS-induced intestinal fibrosis was improved by the new PPAR-γ modulator GED-0507-34 Levo through the modulation of EMT mediators and pro-fibrotic molecules and through GSK-3β induction.

Topics: Animals; Cells, Cultured; Colitis; Dextran Sulfate; Epithelial-Mesenchymal Transition; Fibrosis; Glycogen Synthase Kinase 3 beta; Mice; Mice, Inbred C57BL; PPAR gamma; Rectum; Signal Transduction; Transforming Growth Factor beta

CD4

Topics: Animals; Antigens, Ly; CD4-Positive T-Lymphocytes; Cell Lineage; Colitis; Dextran Sulfate; Fas Ligand Protein; Humans; Interferon-gamma; Interleukin-17; Interleukins; Mice; NK Cell Lectin-Like Receptor Subfamily B; NK Cell Lectin-Like Receptor Subfamily K; T-Lymphocytes, Cytotoxic; Th17 Cells; Th2 Cells; Transforming Growth Factor beta

X-linked inhibitor of apoptosis (xIAP) deficiency is a primary immune deficiency disorder associated with hemophagocytic lymphohistiocytosis. About 17% of xIAP-deficient patients present with very early onset severe colitis with high mortality. We hypothesized that xIAP deficiency leads to defective generation and/or survival of T regulatory cells (Treg) through its involvement in transforming growth factor-β signaling.. We used a T-cell transfer model of chronic colitis and observed a mild increase in colitis severity induced by naïve CD4 T cells from xIAP mice compared with colitis induced by naïve CD4 T cells from WT mice. We did not observe any significant difference in the induction of Treg cells in these studies. We next tested whether xIAP is required for Treg cell function by co-transferring xIAP or WT Treg cells with naïve WT CD4 cells in this model. We demonstrate that XIAP-deficient Treg cells were able to prevent disease similarly to WT Treg cells. In these experiments we, however, found a significantly decreased percentage of IL-17A-producing CD4 T cells in mice receiving Tregs from xIAP mice.. xIAP appears dispensable for the generation of induced Treg cells as well as function of natural Treg cells. There appeared to be a role of xIAP in generation of IL-17-producing cells from either naïve CD4 T cells or Treg cells. Further research is needed to explore the role of xIAP in generation of IL-17-producing cells.

Topics: Animals; Chronic Disease; Colitis; Genetic Diseases, X-Linked; Inhibitor of Apoptosis Proteins; Interleukin-17; Lymphoproliferative Disorders; Mice; Mice, Inbred C57BL; Mice, Transgenic; Signal Transduction; T-Lymphocytes, Regulatory; Th17 Cells; Transforming Growth Factor beta

External radiotherapy is one of the main treatment modalities for a variety of malignancies. However, the lower gastrointestinal tract is sensitive to the ionizing radiation. Hyperbaric oxygen treatment (HOT) has been suggested as a viable treatment for refractory radiation colitis, but the effect of S-Methylisothiourea (SMT) in the radiation colitis have not reported. To investigate the effect of SMT, HOT and the combination of both in an acute radiation-induced enterocolitis model.. Sixty Sprague-Dawley rats were divided randomly into five equal groups. A single dose of gamma irradiation (25 Gy) was administered through the colorectal region to anesthetized rats. In the control group, we applied 2 ml of saline solution intraperitoneally for five days. In the HOT group, 100-per-cent oxygen at 2.5 atm pressure was applied for five days. In the SMT group, 10 mg/kg/day of SMT was applied intraperitoneally for five days. In the HOT+SMT group, HOT and SMT were both applied in the same dosages as in the preceding two groups. At the end of five days, the rats were sacrificed and colon samples were collected for histological grading. Blood samples were collected to test for : tumor necrosis factor-α (TNF-α), interleukin-10 (IL-10), IL-1β, transforming growth factor-β (TGF-β) and intercellular adhesion molecule-1 (ICAM-1) mRNA.. The TNF-α, IL-1β, IL-10 and TGF-β levels were reduced by SMT, HOT and HOT+SMT applications (p < 0.05). However ICAM-1 mRNA levels were not significantly lower (p:0.19). The microscopic scores differed significantly between the SMT, HOT and HOT+SMT groups and the control group. There was significant improvement histologically, especially in the HOT+SMT group. When we compared the weight of the rats before and after the study, weight loss was significantly lower in the SMT, HOT and HOT+SMT groups compared with the control group (p < 0.05).. HOT and SMT together were significantly more effective in preventing weight loss and in reducing inflammation and the severity of colitis histology when compared with HOT and SMT separately.

Topics: Animals; Colitis; Colon; Enzyme Inhibitors; Female; Hyperbaric Oxygenation; Intercellular Adhesion Molecule-1; Interleukin-10; Interleukin-1beta; Isothiuronium; Nitric Oxide Synthase Type II; Radiation Injuries, Experimental; Rats; Rats, Sprague-Dawley; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

FNDC4 is a secreted factor sharing high homology with the exercise-associated myokine irisin (FNDC5). Here we report that Fndc4 is robustly upregulated in several mouse models of inflammation as well as in human inflammatory conditions. Specifically, FNDC4 levels are increased locally at inflamed sites of the intestine of inflammatory bowel disease patients. Interestingly, administration of recombinant FNDC4 in the mouse model of induced colitis markedly reduces disease severity compared with mice injected with a control protein. Conversely, mice lacking Fndc4 develop more severe colitis. Analysis of binding of FNDC4 to different immune cell types reveals strong and specific binding to macrophages and monocytes. FNDC4 treatment of bone marrow-derived macrophages in vitro results in reduced phagocytosis, increased cell survival and reduced proinflammatory chemokine expression. Hence, treatment with FNDC4 results in a state of dampened macrophage activity, while enhancing their survival. Thus, we have characterized FNDC4 as a factor with direct therapeutic potential in inflammatory bowel disease and possibly other inflammatory diseases.

Topics: Amino Acid Sequence; Animals; Anti-Inflammatory Agents; Cells, Cultured; Colitis; Dextran Sulfate; Disease Progression; Gene Expression Regulation; Humans; Intestinal Mucosa; Macrophages; Male; Membrane Proteins; Mice, Inbred C57BL; Mice, Knockout; Molecular Sequence Data; Phagocytosis; Proteins; Signal Transduction; STAT3 Transcription Factor; Transforming Growth Factor beta; Up-Regulation

Carbonic anhydrase I (CA I), a major cecal bacterial antigen, improves inflammatory bowel disease (IBD) symptoms in a murine model. The aim of this study was to identify the responsible epitope region within the CA I protein and evaluate its effect on inflammation using a murine IBD model.. Candidate peptides within the CA I protein sequence that interact with major histocompatibility complex class II were chosen and their immune responses were evaluated using mesentery lymph nodes (MLNs) from a CD4CD25 T-cell transfer murine colitis model. Mice were treated with regulatory dendritic cells (Reg-DCs)-pulsed CA I peptide. We assessed their clinical signs, histopathology, induction of cytokines and transcription factors, and generation of CD103CD11c dendritic cells and regulatory T cells (Tregs).. We identified 4 candidate epitope peptides of CA I. Among these, Reg-DCs pulsed with CA I 58-73 peptide (Reg-DCsCA I 58-73) alone ameliorated colitis. Reg-DCsCA I 58-73-treated mice showed higher mRNA expression levels of forkhead box protein 3, aldehyde dehydrogenase family 1a2, transforming growth factor-β, and interleukin (Il)10, when compared with lower mRNA expression of retinoic acid-related orphan receptor gamma and Il17a in MLNs. Compared with control mice, these mice also showed higher numbers of Foxp3CD4CD25 Tregs and CD103CD11c dendritic cells in MLNs and colon. Administration of Reg-DCsCA I 58-73 induced antigen-specific Tregs in MLNs of colitic mice.. CA I 58-73 peptide induces antigen-specific therapeutic effect in a murine IBD model using Reg-DCs, indicating that CA I 58-73 is a candidate epitope for IBD immunotherapy.

Topics: Aldehyde Dehydrogenase; Aldehyde Dehydrogenase 1 Family; Animals; Antigens, CD; Carbonic Anhydrase I; CD11c Antigen; CD4 Antigens; Colitis; Colon; Dendritic Cells; Epitopes; Female; Forkhead Transcription Factors; Integrin alpha Chains; Interleukin-10; Interleukin-17; Interleukin-2 Receptor alpha Subunit; Lymph Nodes; Mice; Mice, Inbred BALB C; Mice, SCID; Nuclear Receptor Subfamily 1, Group F, Member 1; Peptides; Retinal Dehydrogenase; RNA, Messenger; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Dendritic cells (DCs) mediate host immune responses to gut microbes and play critical roles in inflammatory bowel disease. In this study, we examined the role of TGF-β signaling in DCs in colonic homeostasis. CD11c-cre Tgfbr2(fl/fl) mice developed spontaneous colitis, and CD11c-cre Tgfbr2(fl/+) mice exhibited susceptibility to dextran sulfate sodium-induced colitis. Colitis in these mice was characterized by goblet cell depletion and dysbiosis caused by Enterobacteriaceae enrichment. Wild-type mice gavaged with Enterobacteriaceae from CD11c-cre Tgfbr2(fl/fl) mice feces showed severe colitis after dextran sulfate sodium treatment, whereas those treated with Notch inhibitor exhibited attenuated colonic injury with increased goblet cell numbers, thickened mucus layer, and fewer fecal Enterobacteriaceae Wild-type mice transplanted with CD11c-cre Tgfbr2(fl/fl) bone marrow developed colitis showing increased Jagged1 and Jagged2 in DCs, increased Hes1 levels in epithelium, and goblet cell depletion. These findings suggest that TGF-β signaling in DCs regulates intestinal homeostasis by modulating epithelial cell differentiation and fecal microbiota.

Topics: Animals; Cell Differentiation; Colitis; Colon; Dendritic Cells; Dextran Sulfate; Disease Models, Animal; Epithelial Cells; Gastrointestinal Microbiome; Homeostasis; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Signal Transduction; Transforming Growth Factor beta

Bone-marrow-derived macrophages are divided into two phenotypically and functionally distinct subsets, M1 and M2 macrophages. Recently, it was shown that adoptive transfer of M2-polarized peritoneal macrophages reduced the severity of experimental colitis in mice. However, it is still unclear whether peritoneal macrophages possess the same ability to be polarized to cells with functionally different phenotypes and cytokine production patterns as bone-marrow-derived macrophages. To address this question, we examined the ability of peritoneal macrophages to be polarized to the M1 and M2 phenotypes and determined the specific cytokine profiles of cells with each phenotype. We showed that peritoneal macrophages, as well as bone-marrow-derived macrophages, were differentiated into M1 and M2 phenotypes following stimulation with interferon-γ (IFN-γ) and interleukin-4 (IL-4)/IL-13, respectively. Following in vitro stimulation with lipopolysaccharide, M2-polarized peritoneal macrophages predominantly expressed T helper type 2 (Th2) cytokines and regulatory cytokines, including IL-4, IL-13, transforming growth factor-β and IL-10, whereas M1-polarized peritoneal macrophages expressed negligible amounts of Th1 and pro-inflammatory cytokines. ELISA showed that M2-polarized peritoneal macrophages produced significantly more IL-10 than M1-polarized peritoneal macrophages. Notably, M2-polarized peritoneal macrophages contributed more to the suppression of T-cell proliferation than did M1-polarized peritoneal macrophages. The mRNA expression of Th2 cytokines, including IL-4 and IL-13, increased in T-cells co-cultured with M2-polarized macrophages. Hence, our findings showed that M2 polarization of peritoneal macrophages induced regulatory cytokine production and suppressed T-cell proliferation in vitro, and that resident peritoneal macrophages could be used as a new adoptive transfer therapy for autoimmune/inflammatory diseases after polarization to the regulatory phenotype ex vivo.

Topics: Animals; Cell Differentiation; Cell Proliferation; Cells, Cultured; Coculture Techniques; Colitis; Female; Immunosuppression Therapy; Interleukin-10; Lipopolysaccharides; Macrophages; Macrophages, Peritoneal; Mice; Mice, Inbred C57BL; Phenotype; Th1 Cells; Th2 Cells; Transforming Growth Factor beta

Inflammatory bowel disease (IBD), particularly Crohn's disease, frequently causes intestinal fibrosis that ultimately leads to formation of strictures requiring bowel resection. Currently there is no effective antifibrotic therapy available for this disease. Pirfenidone is a small compound that has a broad spectrum of antifibrogenic effect and has been used for the treatment of fibrotic diseases in various organs. The present study aimed to investigate the antifibrogenic effect of pirfenidone in a dextran sulfate sodium (DSS)-induced murine colitis model. C57BL/6 mice were used and animals were randomly divided into groups receiving pirfenidone or vehicle by oral or transanal routes. Inflammation- and fibrosis-related indexes including body weight, colon length, disease activity, histological change, mRNA expression of pro-inflammatory and pro-fibrogenic cytokines were assessed. Furthermore, we performed in vitro analysis using CCD18-Co fibroblasts to evaluate cell proliferation, transdifferentiation, and viability after the cells were cultured with pirfenidone. It was found that oral administration of pirfenidone reduced deposition of collagen in colitis-associated fibrosis, and significantly suppressed the mRNA expression of col1a2, col3a1, and TGF-β. Moreover, pirfenidone inhibited the activation of TGF-β-related smad and MAPK pathways both in vitro and in vivo. Clinical and histological evaluation demonstrated that pirfenidone had no anti-inflammatory effect. The antifibrogenic effect was reduced when pirfenidone was administered in a delayed manner and was unobserved if given locally. Pirfenidone suppressed fibroblast proliferation and transdifferentiation without observed toxicity. Altogether, our results suggested that oral pirfenidone protects against fibrosis of DSS-induced colitis through inhibiting the proliferation of colonic fibroblasts and TGF-β signaling pathways.

Topics: Administration, Oral; Administration, Rectal; Animals; Anti-Inflammatory Agents, Non-Steroidal; Cell Line; Cell Proliferation; Cell Survival; Colitis; Collagen; Colon; Disease Models, Animal; Female; Fibrosis; Gastrointestinal Agents; Humans; Intestinal Mucosa; Mice, Inbred C57BL; Pyridones; Random Allocation; Signal Transduction; Specific Pathogen-Free Organisms; Transforming Growth Factor beta

Recent studies have revealed various Foxp3(-) regulatory T (Treg) cell subsets effectively protect mice from colitis. In the present study, we demonstrated that B cells induced a particular subset of regulatory T (Treg-of-B) cells, expressing programmed cell death 1 (PD-1), inducible costimulator (ICOS), lymphocyte-activation gene 3 (LAG3), glucocorticoid-induced tumor necrosis factor receptor (GITR), and OX-40, did not express Foxp3. Treg-of-B cells produced abundant levels of IL-10 and low levels of IL-4 and TGF-β. Adoptive transfer of Treg-of-B cells protected mice from CD4(+)CD45RB(hi) T-cell-induced colitis, including infiltration of leukocytes, depletion of goblet cells, epithelial hyperplasia, and inhibition of Th1 and Th17 cytokines. These features were similar to IL-10-producing type 1 regulatory T (Tr1) cells; however, IL-10-deficient Treg-of-B cells maintained their suppressive function in vitro as well as in vivo, while the regulation of Tr1 cells depended on IL-10. In conclusion, Treg-of-B cells protected against experimental colitis through an IL-10-independent mechanism. We reported a novel subpopulation of regulatory T cells was different from conventional Foxp3(+) Treg and IL-10-producing Tr1 cells.

Topics: Adoptive Transfer; Animals; Antigens, CD; B-Lymphocytes; Cell Communication; Colitis; Disease Models, Animal; Forkhead Transcription Factors; Gene Expression Regulation; Glucocorticoid-Induced TNFR-Related Protein; Goblet Cells; Inducible T-Cell Co-Stimulator Protein; Interleukin-10; Interleukin-4; Intestines; Lymphocyte Activation Gene 3 Protein; Mice; Mice, Inbred BALB C; Mice, SCID; Programmed Cell Death 1 Receptor; Receptors, OX40; Severity of Illness Index; Signal Transduction; T-Lymphocytes, Regulatory; Th1 Cells; Th17 Cells; Transforming Growth Factor beta

Myeloid-derived suppressor cells (MDSCs) are well known regulators of regulatory T cells (Treg cells); however, the direct regulation of MDSCs by Treg cells has not been well characterized. We find that colitis caused by functional deficiency of Treg cells leads to altered expansion and reduced function of MDSCs. During differentiation of MDSCs in vitro from bone marrow cells, Treg cells enhanced MDSC function and controlled their differentiation through a mechanism involving transforming growth factor-β (TGF-β). TGF-β-deficient Treg cells were not able to regulate MDSC function in an experimentally induced model of colitis. Finally, we evaluated the therapeutic effect of TGF-β-mediated in-vitro-differentiated MDSCs on colitis. Adoptive transfer of MDSCs that differentiated with TGF-β led to better colitis prevention than the transfer of MDSCs that differentiated without TGF-β. Our results demonstrate an interaction between Treg cells and MDSCs that contributes to the regulation of MDSC proliferation and the acquisition of immunosuppressive functions.

Topics: Adoptive Transfer; Animals; Bone Marrow Cells; Cell Differentiation; Cell Proliferation; Colitis; Dextran Sulfate; Humans; Inflammation; Mice; Myeloid-Derived Suppressor Cells; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Helminth parasites provoke multicellular immune responses in their hosts that can suppress concomitant disease. The gut lumen-dwelling tapeworm Hymenolepis diminuta, unlike other parasites assessed as helminth therapy, causes no host tissue damage while potently suppressing murine colitis. With the goal of harnessing the immunomodulatory capacity of infection with H. diminuta, we assessed the putative generation of anti-colitic regulatory B cells following H. diminuta infection. Splenic CD19(+) B cells isolated from mice infected 7 [HdBc(7(d))] and 14 d (but not 3 d) previously with H. diminuta and transferred to naive mice significantly reduced the severity of dinitrobenzene sulfonic acid (DNBS)-, oxazolone-, and dextran-sodium sulfate-induced colitis. Mechanistic studies with the DNBS model, revealed the anti-colitic HdBc(7(d)) was within the follicular B cell population and its phenotype was not dependent on IL-4 or IL-10. The HdBc(7(d)) were not characterized by increased expression of CD1d, CD5, CD23, or IL-10 production, but did spontaneously, and upon LPS plus anti-CD40 stimulation, produce more TGF-β than CD19(+) B cells from controls. DNBS-induced colitis in RAG1(-/-) mice was inhibited by administration of HdBc(7(d)), indicating a lack of a requirement for T and B cells in the recipient; however, depletion of macrophages in recipient mice abrogated the anti-colitic effect of HdBc(7(d)). Thus, in response to H. diminuta, a putatively unique splenic CD19(+) B cell with a functional immunoregulatory program is generated that promotes the suppression of colitis dominated by TH1, TH2, or TH1-plus-TH2 events, and may do so via the synthesis of TGF-β and the generation of, or cooperation with, a regulatory macrophage.

Topics: Animals; Antigens, CD19; Antigens, CD1d; B-Lymphocytes; Benzenesulfonates; CD40 Antigens; CD5 Antigens; Colitis; Dextran Sulfate; Homeodomain Proteins; Hymenolepiasis; Hymenolepis diminuta; Immunomodulation; Immunotherapy; Interleukin-10; Interleukin-4; Lipopolysaccharides; Macrophages; Male; Mast Cells; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Oxazolone; Receptors, IgE; Th1 Cells; Th2 Cells; Transforming Growth Factor beta

Differentiated CD4(+) T cells preserve plasticity under various conditions. However, the stability of Th1 cells is unclear, as is whether Th1 cells can convert into Th17 cells and thereby contribute to the generation of IFN-γ(+) IL-17(+) CD4(+) T cells, the number of which correlates with severity of colitis. We investigated whether IFN-γ(+) Th1 cells can convert into Th17 cells under intestinal inflammation and the mechanisms involved. IFN-γ(Thy1.1+) Th1 cells were generated by culturing naïve CD4(+) T cells from IFN-γ(Thy1.1) CBir1 TCR-Tg reporter mice, whose TCR is specific for an immunodominant microbiota antigen, CBir1 flagellin, under Th1 polarizing conditions. IFN-γ(Thy1.1+) Th1 cells induced colitis in Rag(-/-) mice after adoptive transfer and converted into IL-17(+) Th17, but not Foxp3(+) Treg cells in the inflamed intestines. TGF-β and IL-6, but not IL-1β and IL-23, regulated Th1 conversion into Th17 cells. TGF-β induction of transcriptional factor Runx1 is crucial for the conversion, since silencing Runx1 by siRNA inhibited Th1 conversion into Th17 cells. Furthermore, TGF-β enhanced histone H3K9 acetylation but inhibited H3K9 trimethylation of Runx1- and ROR-γt-binding sites on il-17 or rorc gene in Th1 cells. We conclude that Th1 cells convert into Th17 cells under inflammatory conditions in intestines, which is possibly mediated by TGF-β induction of Runx1.

Topics: Acetylation; Animals; Binding Sites; Cell Differentiation; Cells, Cultured; Colitis; Core Binding Factor Alpha 2 Subunit; Flagellin; Histones; Homeodomain Proteins; Interferon-gamma; Interleukin-17; Interleukin-1beta; Interleukin-2; Interleukin-23; Interleukin-6; Intestinal Mucosa; Lymphocyte Activation; Lymphocyte Count; Methylation; Mice; Mice, Inbred C57BL; Mice, Knockout; Nuclear Receptor Subfamily 1, Group F, Member 1; Nuclear Receptor Subfamily 1, Group F, Member 3; RNA Interference; RNA, Small Interfering; Th1 Cells; Th17 Cells; Transforming Growth Factor beta

Colorectal cancer is strongly associated with lipid metabolism. NPC1L1, a sterol transporter, plays a key role in modulating lipid homeostasis in vivo. Its inhibitor, ezetimibe, began to be used clinically to lower cholesterol and this caused the great debate on its role in causing carcinogenesis. Here we explored the role of NPC1L1 in colorectal tumorigenesis.. Wild-type mice and NPC1L1(-/-) (NPC1L1 knockout) mice were treated with azoxymethane (AOM)-dextran sodium sulfate (DSS) to induce colitis-associated colorectal tumorigenesis. Mice were sacrificed 10, 15, 18 or 20 weeks after AOM treatment, respectively. Colorectal tumors were counted and analyzed. Plasma lipid concentrations were measured using enzymatic reagent kit. Protein expression level was assayed by western blot.. NPC1L1(-/-) mice significantly had fewer tumors than wild-type. The ratio of malignant/tumor in NPC1L1(-/-) mice was significantly lower than in wild-type 20 weeks after AOM-DSS treatment. NPC1L1 was highly expressed in the small intestine of wild-type mice but its expression was undetectable in colorectal mucous membranes or tumors in either group. NPC1L1 knockout decreased plasma total cholesterol and phospholipid. NPC1L1(-/-) mice had significant lower intestinal inflammation scores and expressed inflammatory markers p-c-Jun, p-ERK and Caspase-1 p20 lower than wild-type. NPC1L1 knockout also reduced lymphadenectasis what may be caused by inflammation. NPC1L1 knockout in mice decreased β-catenin in tumors and regulated TGF-β and p-gp in adjacent colons or tumors. There was not detectable change of p53 by NPC1L1 knockout.. Our results provide the first evidence that NPC1L1 knockout protects against colitis-associated tumorigenesis. NPC1L1 knockout decreasing plasma lipid, especially cholesterol, to reduce inflammation and decreasing β-catenin, p-c-Jun and p-ERK may be involved in the mechanism.

Topics: Animals; ATP Binding Cassette Transporter, Subfamily B, Member 1; Azoxymethane; beta Catenin; Cell Transformation, Neoplastic; Colitis; Colonic Neoplasms; Dextran Sulfate; Disease Models, Animal; Homozygote; Intestinal Mucosa; Lipids; Membrane Transport Proteins; Mice; Mice, Knockout; Transforming Growth Factor beta; Tumor Suppressor Protein p53

Different studies have described the successful use of recombinant lactic acid bacteria (recLAB) to deliver anti-inflammatory molecules at the mucosal level to treat Inflammatory Bowel Disease (IBD).. In order to identify the best strategy to treat IBD using recLAB, we compared the efficacy of different recombinant strains of Lactococcus lactis (the model LAB) secreting two types of anti-inflammatory molecules: cytokines (IL-10 and TGF-β1) and serine protease inhibitors (Elafin and Secretory Leukocyte Protease Inhibitor: SLPI), using a dextran sulfate sodium (DSS)-induced mouse model of colitis.. Our results show that oral administration of recombinant L. lactis strains expressing either IL-10 or TGF-β1 display moderate anti-inflammatory effects in inflamed mice and only for some clinical parameters. In contrast, delivery of either serine protease inhibitors Elafin or SLPI by recLAB led to a significant reduction of intestinal inflammation for all clinical parameters tested. Since the best results were obtained with Elafin-producing L. lactis strain, we then tried to enhance Elafin expression and hence its delivery rate by producing it in a L. lactis mutant strain inactivated in its major housekeeping protease, HtrA. Strikingly, a higher reduction of intestinal inflammation in DSS-treated mice was observed with the Elafin-overproducing htrA strain suggesting a dose-dependent Elafin effect.. Altogether, these results strongly suggest that serine protease inhibitors are the most efficient anti-inflammatory molecules to be delivered by recLAB at the mucosal level for IBD treatment.

Topics: Administration, Oral; Animals; Colitis; Disease Models, Animal; Elafin; Gene Expression; Interleukin-10; Lactococcus lactis; Mice; Mice, Inbred C57BL; Nisin; Secretory Leukocyte Peptidase Inhibitor; Serine Proteinase Inhibitors; Transforming Growth Factor beta

Regulatory T (Treg) cells play a pivotal role in suppressing self-harmful T cell responses, but how Treg cells mediate suppression to maintain immune homeostasis and limit responses during inflammation is unclear. Here we show that effector Treg cells express high amounts of the integrin αvβ8, which enables them to activate latent transforming growth factor-β (TGF-β). Treg-cell-specific deletion of integrin αvβ8 did not result in a spontaneous inflammatory phenotype, suggesting that this pathway is not important in Treg-cell-mediated maintenance of immune homeostasis. However, Treg cells lacking expression of integrin αvβ8 were unable to suppress pathogenic T cell responses during active inflammation. Thus, our results identify a mechanism by which Treg cells suppress exuberant immune responses, highlighting a key role for effector Treg-cell-mediated activation of latent TGF-β in suppression of self-harmful T cell responses during active inflammation.

Topics: Animals; Cell Proliferation; Colitis; Disease Models, Animal; Gene Expression Regulation; Humans; Inflammation; Inflammation Mediators; Integrins; Mice; Models, Immunological; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Monocytes (Mos) play an important role in the pathogenesis of intestinal mucosal inflammation. This study aims to investigate the mechanism by which the intestinal epithelial cell-derived thrombospondin 1 (TSP1) modulates Mo properties and regulates intestinal inflammatory responses. In this study, the production of TSP1 by intestinal epithelial cells was evaluated by quantitative real-time PCR and Western blotting. The properties of Mos were analyzed by flow cytometry. A mouse model of colitis was created to assess the role of epithelium-derived TSP1 in the suppression of intestinal inflammation. The results demonstrated that mouse intestinal epithelial cells (IECs) expressed TSP1, which was markedly upregulated by butyrate or feeding with Clostridium butyricum. Coculture of the butyrate-primed IECs and Mos or exposure of Mos to TSP1 in the culture induced the expression of transforming growth factor (TGF)-β in Mos. These TGF-β+ Mos had tolerogenic properties that could promote generation of inducible regulatory T cells. Adoptive transfer with TSP1-primed Mos, or feeding C. butyricum could prevent experimental colitis in mice. In summary, C. butyricum induces intestinal epithelial cells to produce TSP1 and induces TGF-β+ Mos, which further suppress experimental colitis in mice. The results implicate that the administration of C. butyricum or butyrate may have the potential to ameliorate chronic intestinal inflammation through inducing immunosuppressive Mos.

Topics: Animals; Butyrates; Clostridium butyricum; Colitis; Disease Models, Animal; Inflammation; Intestinal Mucosa; Male; Mice; Mice, Knockout; Monocytes; Thrombospondin 1; Transforming Growth Factor beta

Regulatory T (Treg) cells are essential for maintenance of immune homeostasis. Here we found that hydrogen sulfide (H2S) was required for Foxp3(+) Treg cell differentiation and function and that H2S deficiency led to systemic autoimmune disease. H2S maintained expression of methylcytosine dioxygenases Tet1 and Tet2 by sulfhydrating nuclear transcription factor Y subunit beta (NFYB) to facilitate its binding to Tet1 and Tet2 promoters. Transforming growth factor-β (TGF-β)-activated Smad3 and interleukin-2 (IL-2)-activated Stat5 facilitated Tet1 and Tet2 binding to Foxp3. Tet1 and Tet2 catalyzed conversion of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC) in Foxp3 to establish a Treg-cell-specific hypomethylation pattern and stable Foxp3 expression. Consequently, Tet1 and Tet2 deletion led to Foxp3 hypermethylation, impaired Treg cell differentiation and function, and autoimmune disease. Thus, H2S promotes Tet1 and Tet2 expression, which are recruited to Foxp3 by TGF-β and IL-2 signaling to maintain Foxp3 demethylation and Treg-cell-associated immune homeostasis.

Topics: Adoptive Transfer; Animals; CCAAT-Binding Factor; Cell Differentiation; Colitis; Dioxygenases; DNA Methylation; DNA-Binding Proteins; Forkhead Transcription Factors; Homeostasis; Humans; Hydrogen Sulfide; Interleukin-2; Mice; Mice, Inbred C57BL; Mice, Knockout; Proto-Oncogene Proteins; Smad3 Protein; STAT5 Transcription Factor; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

The intestinal epithelial barrier plays a critical role in the mucosal immunity. However, it remains largely unknown how the epithelial barrier is maintained after damage. Here we show that growth factor FGF2 synergized with interleukin-17 (IL-17) to induce genes for repairing of damaged epithelium. FGF2 or IL-17 deficiency resulted in impaired epithelial proliferation, increased pro-inflammatory microbiota outgrowth, and consequently worse pathology in a DSS-induced colitis model. The dysregulated microbiota in the model induced transforming growth factor beta 1 (TGFβ1) expression, which in turn induced FGF2 expression mainly in regulatory T cells. Act1, an essential adaptor in IL-17 signaling, suppressed FGF2-induced ERK activation through binding to adaptor molecule GRB2 to interfere with its association with guanine nucleotide exchange factor SOS1. Act1 preferentially bound to IL-17 receptor complex, releasing its suppressive effect on FGF2 signaling. Thus, microbiota-driven FGF2 and IL-17 cooperate to repair the damaged intestinal epithelium through Act1-mediated direct signaling cross-talk.

Topics: Adaptor Proteins, Signal Transducing; Animals; Blotting, Western; Cell Line, Tumor; Cells, Cultured; Colitis; Fibroblast Growth Factor 2; Gene Expression Profiling; HEK293 Cells; HeLa Cells; HT29 Cells; Humans; Interleukin-17; Intestinal Mucosa; Intestines; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Microbiota; Oligonucleotide Array Sequence Analysis; Reverse Transcriptase Polymerase Chain Reaction; T-Lymphocyte Subsets; T-Lymphocytes, Regulatory; Transforming Growth Factor beta; Wound Healing

Thymic stromal lymphopoietin (TSLP) is a cytokine known to mature dendritics cells, lower pro-inflammatory IL-12 secretion, induce differentiation of anti-inflammatory FoxP3+ regulatory T cells (Treg). Moreover, Crohn's disease patients have shown a reduction of intestinal TSLP expression. To understand the role of TSLP in inflammation, we constructed Lactococcus lactis strain producing TSLP (LL-TSLP) and investigated the effect of its administration on dextran sulfate sodium (DSS)-induced colitis model in mice.. LL-TSLP secrete an active molecule which lowers secretion of IL-12 by dendritic cells. Treatment with LL-TSLP, increases the amount of TGF-β secreted by T cells in Mesenteric Lymph Node in healthy mice. In acute DSS-induced colitis, LL-TSLP delayed the Disease Activity Index and lowered histological score and colonic INF-γ production. In a DSS-recovery model, LL-TSLP induced a better protective effect if the strain was administered at the beginning of the colitis. At Day 4 of colitis we observed an induction of Treg by LL-TSLP.. TSLP showed an anti-inflammatory protective role in DSS-induced colitis. We have demonstrated that a short and early administration of LL-TSLP is more efficient than a long lasting treatment.

Topics: Administration, Oral; Animals; Colitis; Colon; Cytokines; Dendritic Cells; Dextran Sulfate; Disease Models, Animal; Inflammation; Interleukin-12; Intestinal Mucosa; Lactococcus lactis; Lymph Nodes; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; T-Lymphocytes, Regulatory; Thymic Stromal Lymphopoietin; Transforming Growth Factor beta

The adoptive transfer of the natural regulatory B cells and macrophages should be a useful treatment for inflammation and autoimmune disease. However, it is usually difficult to isolate these cells from the tissues and expand them. Here, we investigated the feasibility of adoptively transferring peritoneal cells (PCs) as a treatment for DSS-induced colitis. We found that peritoneal cavity can provide an easily accessible site for harvesting enough number of PCs, namely, two-dose PCs for the treatment from a mouse in one operation. Adoptive therapy of these cells from healthy mice or those with disease is effectively in reducing the disease activity score. The natural B cells and macrophages of the infused PCs can selectively migrate to lesion sites and regulate the expression of Stat3, NF-κB, Smad3 and Smad7. Additionally, PCs exert dual activity of IL-10 and TGF-β secreted spontaneously by both peritoneal B cells and macrophages, which in turn enhance the induction of regulatory B cells and Macrophages in microenvironment of inflammation. Moreover, PCs can re-establish immunological tolerance in the OVA-immunized mice. Thus, our findings provide a new strategy for colitis therapy and could be of importance in additional exploration of other inflammation and autoimmune diseases therapy.

Topics: Adoptive Transfer; Animals; B-Lymphocytes; Colitis; Dextran Sulfate; Disease Models, Animal; Female; Inflammation; Interleukin-10; Intestinal Mucosa; Macrophages; Mice; Mice, Inbred C57BL; NF-kappa B; Signal Transduction; Smad Proteins, Receptor-Regulated; Transforming Growth Factor beta

Food protein-induced allergic proctocolitis (FPIAP) is mostly a non-immunoglobulin E-mediated disease where a T-cell-mediated reaction to cow's milk protein has been suggested. We determined the expression of transforming growth factor (TGF)-β, TGF-β receptor-1, tumor necrosis factor (TNF)-α, CD86, and CD23 on the colon mucosa to investigate their roles in the pathogenesis of the two subtypes of FPIAP, i.e. infantile FPIAP and FPIAP in older children.. Group 1 comprised children with infantile FPIAP (age <6 months, n = 21), group 2 referred to FPIAP in older children (age >1.5 years, n = 7), and group 3 included children with juvenile hyperplastic polyps (n = 22). Immunohistochemical staining of colonic biopsy specimens was performed.. The expression of TNF-α was significantly higher in groups 1 and 2 compared to group 3. Group 2 patients had a significantly lower TGF-β expression compared to the other groups. The expression of CD86 was higher in group 1 than in group 3 (p = 0.012). Eosinophil counts per high-power field in the lamina propria were significantly correlated with CD86 expression (p = 0.026, r = 0.388).. Our results suggest that TNF-α is implicated in the pathogenesis of both types of FPIAP. The decreased activity of TGF-β receptor-1 accompanied by the increased expression of CD86 in infants and the decreased activity of TGF-β in older children appear to play a role in the development of FPIAP.

Topics: B7-2 Antigen; Biopsy; Child, Preschool; Colitis; Cytokines; Enterocolitis; Female; Food Hypersensitivity; Humans; Immunoglobulin E; Infant; Intestinal Mucosa; Male; Milk Hypersensitivity; Protein Serine-Threonine Kinases; Receptor, Transforming Growth Factor-beta Type I; Receptors, IgE; Receptors, Transforming Growth Factor beta; Transforming Growth Factor beta

Transforming growth factor-βs (TGF-βs) are secreted from cells as latent complexes and the activity of TGF-βs is controlled predominantly through activation of these complexes. Tolerance to the fetal allograft is essential for pregnancy success; TGF-β1 and TGF-β2 play important roles in regulating these processes. Pregnancy-specific β-glycoproteins (PSGs) are present in the maternal circulation at a high concentration throughout pregnancy and have been proposed to have anti-inflammatory functions. We found that recombinant and native PSG1 activate TGF-β1 and TGF-β2 in vitro. Consistent with these findings, administration of PSG1 protected mice from dextran sodium sulfate (DSS)-induced colitis, reduced the secretion of pro-inflammatory cytokines, and increased the number of T regulatory cells. The PSG1-mediated protection was greatly inhibited by the coadministration of neutralizing anti-TGF-β antibody. Our results indicate that proteins secreted by the placenta directly contribute to the generation of active TGF-β and identify PSG1 as one of the few known biological activators of TGF-β2.

Topics: Animals; CD4-Positive T-Lymphocytes; Colitis; Cytokines; Dextran Sulfate; Disease Models, Animal; Forkhead Transcription Factors; Intestinal Mucosa; Mice; Pregnancy-Specific beta 1-Glycoproteins; Protein Binding; Transforming Growth Factor beta; Transforming Growth Factor beta1; Transforming Growth Factor beta2

Tenascin-C (TnC) is a multi-domain extracellular matrix glycoprotein that is expressed at a high level during embryogenesis but is almost absent during normal postnatal life. This multi-domain complex molecule is reported to associate with both pro-inflammatory and anti-inflammatory signalling cascades. In this study, we examined how TnC modulated intestinal inflammation.. TnC pathophysiology was evaluated in cultures of rat intestinal subepithelial myofibroblasts (ISEMF) and intestinal epithelial cells. Wild-type and TnC(-/-) mice were treated with dextran sodium sulfate (DSS) to induce colitis.. DSS-induced colitis in mice markedly increased TnC in the damaged mucosal areas and up-regulated mRNA for TnC, pro-inflammatory cytokines and growth factors (PDGF-B and TGF-β1). In addition, 2,4,6-trinitrobenzene sulfonic acid-induced colitis and SAMP1/Yit mice, a model of spontaneous Crohn's disease, also exhibited increased mucosal TnC in colon and ilea respectively. PDGF receptor-α (PDGFRα) positive ISEMF were the primary TnC-producing cells in colon tissues. Accordingly, ISEMF collected from the rat colon constitutively expressed both TnC and PDGFRα. PDGF-BB and TGF-β1 up-regulated both TnC mRNA and protein levels in ISEMF. Knock-down of TnC gene increased susceptibility to DSS-induced colitis, compared with TnC(+/+) littermates. TnC(-/-) mice showed marked abrasion of intestinal mucosal barrier and increased inflammatory scores. Moreover, TnC accelerated both trans-well migration and wound healing in epithelial cells.. The pharmacological profiles of PDGF-BB and TGF-β in colitis tissues and ISEMF suggest that increased TnC production during inflammation contributed to epithelial cell migration, remodelling and protection of intestinal barriers.

Topics: Actins; Animals; Anti-Ulcer Agents; Blotting, Western; Cell Movement; Colitis; Cytokines; Epithelial Cells; Fluorescent Antibody Technique; Intestinal Mucosa; Lac Operon; Male; Mice; Microscopy, Immunoelectron; Molecular Sequence Data; Myofibroblasts; Platelet-Derived Growth Factor; Rats; Rats, Sprague-Dawley; Receptor, Platelet-Derived Growth Factor alpha; RNA; Tenascin; Transforming Growth Factor beta; Wound Healing

Disturbance of T-cell homeostasis could lead to intestinal inflammation. Naive CD4 T cells undergoing spontaneous proliferation, a robust proliferative response that occurs under severe lymphopenic conditions, differentiate into effector cells producing Th1- and/or Th17-type cytokines and induce a chronic inflammation in the intestine that resembles human inflammatory bowel disease. In this study, we investigated the key properties of CD4 T cells necessary to induce experimental colitis. α4β7 upregulation was primarily induced by mesenteric lymph node (mLN) resident CD11b(+) dendritic cell subsets via transforming growth factor beta (TGFβ)/retinoic acid-dependent mechanism. Interestingly, α4β7 expression was essential but not sufficient to induce inflammation. In addition to gut-homing specificity, expression of gut Ag specificity was also crucial. T-cell acquisition of the specificity was dramatically enhanced by the presence of γδ T cells, a population previously shown to exacerbate T-cell-mediated colitis. Importantly, interleukin (IL)-23-mediated γδ T cell stimulation was necessary to enhance colitogenicity but not gut antigen reactivity of proliferating CD4 T cells. These findings demonstrate that T-cell colitogenicity is achieved through multiple processes, offering a therapeutic rationale by intervening these pathways.

Topics: Animals; Antineoplastic Agents; CD4-Positive T-Lymphocytes; Colitis; Dendritic Cells; Gastrointestinal Tract; Genes, T-Cell Receptor beta; Homeodomain Proteins; Humans; Inflammation; Integrins; Interleukin-16; Interleukin-23 Subunit p19; Lymph Nodes; Mesenteric Veins; Mice; Mice, Inbred C57BL; Mice, Knockout; Real-Time Polymerase Chain Reaction; Receptors, Antigen, T-Cell, gamma-delta; RNA, Messenger; T-Lymphocytes, Regulatory; Th17 Cells; Transforming Growth Factor beta; Tretinoin

Chronic intestinal inflammation is associated with pathological angiogenesis that further amplifies the inflammatory response. Vascular endothelial growth factor (VEGF), is a major angiogenic cytokine that has been implicated in chronic colitis and inflammatory bowel diseases. Endoglin (CD105), a transforming growth factor-β superfamily co-receptor expressed on endothelial and some myeloid cells, is a modulator of angiogenesis involved in wound healing and potentially in resolution of inflammation. We showed previously that Endoglin heterozygous (Eng (+/-)) mice subjected to dextran sodium sulfate developed severe colitis, abnormal colonic vessels and high tissue VEGF. We therefore tested in the current study if treatment with a monoclonal antibody to VEGF could ameliorate chronic colitis in Eng (+/-) mice. Tissue inflammation and microvessel density (MVD) were quantified on histological slides. Colonic wall thickness, microvascular hemodynamics and targeted MAdCAM-1(+) inflamed vessels were assessed in vivo by ultrasound. Mediators of angiogenesis and inflammation were measured by Milliplex and ELISA assays. Colitic Eng (+/-) mice showed an increase in intestinal inflammation, MVD, colonic wall thickness, microvascular hemodynamics and the number of MAdCAM-1(+) microvessels relative to colitic Eng (+/+) mice; these parameters were all attenuated by anti-VEGF treatment. Of all factors up-regulated in the inflamed gut, granulocyte colony-stimulating factor (G-CSF) and amphiregulin were further increased in colitic Eng (+/-) versus Eng (+/+) mice. Anti-VEGF therapy decreased tissue VEGF and inflammation-induced endoglin, IL-1β and G-CSF in colitic Eng (+/-) mice. Our results suggest that endoglin modulates intestinal angiogenic and inflammatory responses in colitis. Furthermore, contrast-enhanced ultrasound provides an excellent non-invasive imaging modality to monitor gut angiogenesis, inflammation and responses to anti-angiogenic treatment.

Topics: Animals; Antibodies, Monoclonal; Colitis; Colon; Endoglin; Female; Granulocyte Colony-Stimulating Factor; Hemodynamics; Heterozygote; Inflammation; Inflammation Mediators; Interleukin-1beta; Intestines; Intracellular Signaling Peptides and Proteins; Male; Mice, Inbred C57BL; Microvessels; Platelet Endothelial Cell Adhesion Molecule-1; Smad Proteins; Transforming Growth Factor beta; Vascular Endothelial Growth Factor A

Astilbin, a major bioactive compound from Rhizoma smilacis glabrae, has been reported to possess anti-inflammatory properties. Our study first evaluated astilbin on dextran sulfate sodium (DSS)-induced acute colitis in mice. By intraperitoneal injection of astilbin, the severity of colitis was attenuated, and the serum levels of IL-10 and TGF-β were increased. Using flow cytometry, a higher number of IL-10(+) dendritic cells (DCs) and TGF-β(+) DCs and a lower number of CD86(+) DCs, IL-12 p40(+) DCs, and IL-1β(+) DCs were detected in the spleen of mice with colitis after astilbin treatment. The administration of astilbin also resulted in the upregulation of CD103(+) expression in colonic DCs. In a coculture system, murine bone marrow-derived DCs pretreated with astilbin resulted in an enhanced production of CD4(+)CD25(+)Foxp3(+) T cells. The results of this study show that astilbin could be a candidate drug for inflammatory bowel disease by mediating the regulatory functions of DCs.

Topics: Animals; Anti-Inflammatory Agents; Colitis; Dendritic Cells; Dextran Sulfate; Flavonols; Interleukin-10; Mice; Mice, Inbred C57BL; Transforming Growth Factor beta; Treatment Outcome

The binding of NKG2D to its ligands strengthens the cross-talk between natural killer (NK) cells and dendritic cells, particularly at early stages, before the initiation of the adaptive immune response. We found that retinoic acid early transcript-1ε (RAE-1ε), one of the ligands of NKG2D, was persistently expressed on antigen-presenting cells in a transgenic mouse model (pCD86-RAE-1ε). By contrast, NKG2D expression on NK cells, NKG2D-dependent cytotoxicity and tumour rejection, and dextran sodium sulphate-induced colitis were all down-regulated in this mouse model. The down-regulation of NKG2D on NK cells was reversed by stimulation with poly (I:C). The ectopic expression of RAE-1ε on dendritic cells maintained NKG2D expression levels and stimulated the activity of NK cells ex vivo, but the higher frequency of CD4(+) NKG2D(+) T cells in transgenic mice led to the down-regulation of NKG2D on NK cells in vivo. Hence, high levels of RAE-1ε expression on antigen-presenting cells would be expected to induce the down-regulation of NK cell activation by a regulatory T-cell subset.

Topics: Animals; B7-2 Antigen; CD4-Positive T-Lymphocytes; Cells, Cultured; Colitis; Dendritic Cells; Dextran Sulfate; Disease Models, Animal; Down-Regulation; Killer Cells, Natural; Ligands; Melanoma, Experimental; Membrane Proteins; Mice; Mice, Inbred C57BL; Mice, Transgenic; NK Cell Lectin-Like Receptor Subfamily K; Poly I-C; Promoter Regions, Genetic; T-Lymphocytes, Regulatory; Time Factors; Transforming Growth Factor beta

Alleviation of dextran sulfate sodium (DSS)-induced colitis was shown after by transplantation of bone marrow derived cells in mice. Nevertheless, the underlying mechanism remains elusive. In the present study, we transplanted primary mouse mesenchymal stem cells (MSC) into isogeneic mice with DSS-induced colitis. We found that MSC transplantation significantly alleviated the DSS-induced colitis. Inhibition of transforming growth factor beta (TGF-beta) signaling abrogated the therapeutic effect of MSC transplantation on DSS-colitis, suggesting a TGF-beta signaling-dependent manner. Moreover, MSC transplantation seemed to induce M2 macrophage polarization, which appeared to be the major source of TGF-beta in this model. Our data thus demonstrate that MSC transplantation may activate TGF-beta signaling pathways to promote the recovery of DSS-colitis.

Topics: Animals; Blotting, Western; Colitis; Dextran Sulfate; Flow Cytometry; Mesenchymal Stem Cell Transplantation; Mice; Reverse Transcriptase Polymerase Chain Reaction; Signal Transduction; Transforming Growth Factor beta

Ginsenoside Rh2 (GRh2) has been reported to have therapeutic effects on various diseases. However, whether it may also affect the recovery from ulcerative colitis remains unknown. Here we induced colitis in mice by dextran sulfate sodium (DSS) administration, and then treated the mice with GRh2. We found that GRh2-treated mice showed significant alleviation of the DSS-induced colitis. Moreover, significant increase in the activity of TGFβ signaling was detected in the GRh2-treated colon that had received DSS. To investigate whether there is a causative link among GRh2 treatment, TGFβ signaling augment and the cure of colitis, we gave the DSS-treated mice a combination of GRh2 and a specific TGFβ receptor I inhibitor, SB431542. SB431542 significantly decreased the activation of TGFβ signaling in the colon from the GRh2-administrated mice, and consequently attenuated the therapeutic effect of GRh2. Our data thus demonstrate that GRh2 may alleviate DSS-induced colitis via augmenting TGFβ signaling.

Topics: Animals; Colitis; Colon; Dextran Sulfate; Ginsenosides; Male; Mice; Mice, Inbred C57BL; Signal Transduction; Transforming Growth Factor beta

FOXP3(+) regulatory T cells (Treg cells) are abundant in the intestine, where they prevent dysregulated inflammatory responses to self and environmental stimuli. It is now appreciated that Treg cells acquire tissue-specific adaptations that facilitate their survival and function; however, key host factors controlling the Treg response in the intestine are poorly understood. The interleukin (IL)-1 family member IL-33 is constitutively expressed in epithelial cells at barrier sites, where it functions as an endogenous danger signal, or alarmin, in response to tissue damage. Recent studies in humans have described high levels of IL-33 in inflamed lesions of inflammatory bowel disease patients, suggesting a role for this cytokine in disease pathogenesis. In the intestine, both protective and pathological roles for IL-33 have been described in murine models of acute colitis, but its contribution to chronic inflammation remains ill defined. Here we show in mice that the IL-33 receptor ST2 is preferentially expressed on colonic Treg cells, where it promotes Treg function and adaptation to the inflammatory environment. IL-33 signalling in T cells stimulates Treg responses in several ways. First, it enhances transforming growth factor (TGF)-β1-mediated differentiation of Treg cells and, second, it provides a necessary signal for Treg-cell accumulation and maintenance in inflamed tissues. Strikingly, IL-23, a key pro-inflammatory cytokine in the pathogenesis of inflammatory bowel disease, restrained Treg responses through inhibition of IL-33 responsiveness. These results demonstrate a hitherto unrecognized link between an endogenous mediator of tissue damage and a major anti-inflammatory pathway, and suggest that the balance between IL-33 and IL-23 may be a key controller of intestinal immune responses.

Topics: Animals; Colitis; Colon; Disease Models, Animal; Female; Immunity, Mucosal; Inflammation; Interleukin-23; Interleukin-33; Interleukins; Intestines; Male; Mice; Mice, Inbred C57BL; Receptors, Interleukin; Signal Transduction; T-Lymphocytes, Regulatory; Thymus Gland; Transforming Growth Factor beta

The β-galactoside-binding protein galectin-9 is critical in regulating the immune response, but the mechanism by which it functions remains unclear. We have demonstrated that galectin-9 is highly expressed by induced regulatory T cells (iTreg) and was crucial for the generation and function of iTreg cells, but not natural regulatory T (nTreg) cells. Galectin-9 expression within iTreg cells was driven by the transcription factor Smad3, forming a feed-forward loop, which further promoted Foxp3 expression. Galectin-9 increased iTreg cell stability and function by directly binding to its receptor CD44, which formed a complex with transforming growth factor-β (TGF-β) receptor I (TGF-βRI), and activated Smad3. Galectin-9 signaling was further found to regulate iTreg cell induction by dominantly acting through the CNS1 region of the Foxp3 locus. Our data suggest that exogenous galectin-9, in addition to being an effector molecule for Treg cells, acts synergistically with TGF-β to enforce iTreg cell differentiation and maintenance.

Topics: Animals; Cell Differentiation; Colitis; Forkhead Transcription Factors; Galectins; Hepatitis A Virus Cellular Receptor 2; Hyaluronan Receptors; Lymphocyte Activation; Mice; Mice, Inbred C57BL; Mice, Knockout; Protein Serine-Threonine Kinases; Receptor, Transforming Growth Factor-beta Type I; Receptors, Transforming Growth Factor beta; Receptors, Virus; Signal Transduction; Smad3 Protein; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

The commensal yeast Candida albicans is part of the human intestinal microflora and is considered a "pathobiont", a resident microbe with pathogenic potential yet harmless under normal conditions. The aim of this study was to investigate the effect of C. albicans on inflammation of the intestinal tract and the role of Bruton's tyrosine kinase (Btk). Btk is an enzyme that modulates downstream signaling of multiple receptors involved in innate and adaptive immunity, including the major anti-fungal receptor Dectin-1. Colitis was induced in wild type and Btk-/- mice by treatment with dextran sodium sulfate (DSS) and the gastrointestinal tract of selected treatment groups were then colonized with C. albicans. Colonization by C. albicans neither dampened nor exacerbated inflammation in wild type mice, but colon length and spleen weight were improved in Btk-deficient mice colonized with C. albicans. Neutrophil infiltration was comparable between wild type and Btk-/- mice, but the knockout mice displayed severely reduced numbers of macrophages in the colon during both DSS and DSS/Candida treatment. Smaller numbers and reduced responsiveness of Btk-/- macrophages might partially explain the improved colon length of Btk-/- mice as a result of Candida colonization. Surprisingly, DSS/Candida-treated Btk-/- animals had higher levels of certain pro-inflammatory cytokines and levels of the anti-inflammatory cytokine TGF-β were reduced compared to wild type. A clustering and correlation analysis showed that for wild type animals, spleen TGF-β and colon IL-10 and for Btk-/- spleen and colon levels of IL-17A best correlated with the inflammatory parameters. We conclude that in Btk-/- immunocompromised animals, colonization of the gastrointestinal tract by the commensal yeast C. albicans alters inflammatory symptoms associated with colitis.

Topics: Agammaglobulinaemia Tyrosine Kinase; Animals; Candida albicans; Candidiasis; Colitis; Colon; Cytokines; Dextran Sulfate; Host-Pathogen Interactions; Humans; Immunohistochemistry; Inflammation Mediators; Interleukin-10; Interleukin-17; Intestinal Mucosa; Intestines; Macrophages; Mice, Knockout; Organ Size; Protein-Tyrosine Kinases; Spleen; Transforming Growth Factor beta

The molecular mechanism of the extrathymic generation of adaptive, or inducible, CD4(+)Foxp3(+) regulatory T cells (iTregs) remains incompletely defined. We show that exposure of splenic CD4(+)CD25(+)Foxp3(-) cells to IL-2, but not other common γ-chain cytokines, resulted in Stat5 phosphorylation and induced Foxp3 expression in ∼10% of the cells. Thus, IL-2/Stat5 signaling may be critical for Foxp3 induction in peripheral CD4(+)CD25(+)Foxp3(-) iTreg precursors. In this study, to further define the role of IL-2 in the formation of iTreg precursors as well as their subsequent Foxp3 expression, we designed a two-step iTreg differentiation model. During the initial "conditioning" step, CD4(+)CD25(-)Foxp3(-) naive T cells were activated by TCR stimulation. Inhibition of IL-2 signaling via Jak3-Stat5 was required during this step to generate CD4(+)CD25(+)Foxp3(-) cells containing iTreg precursors. During the subsequent Foxp3-induction step driven by cytokines, IL-2 was the most potent cytokine to induce Foxp3 expression in these iTreg precursors. This two-step method generated a large number of iTregs with relatively stable expression of Foxp3, which were able to prevent CD4(+)CD45RB(high) cell-mediated colitis in Rag1(-/-) mice. In consideration of this information, whereas initial inhibition of IL-2 signaling upon T cell priming generates iTreg precursors, subsequent activation of IL-2 signaling in these precursors induces the expression of Foxp3. These findings advance the understanding of iTreg differentiation and may facilitate the therapeutic use of iTregs in immune disorders.

Topics: Animals; CD4 Antigens; CD4-Positive T-Lymphocytes; Cell Differentiation; Colitis; Forkhead Transcription Factors; Interleukin-2; Interleukin-2 Receptor alpha Subunit; Mice; Mice, Knockout; Phosphorylation; Precursor Cells, T-Lymphoid; Receptors, Antigen, T-Cell; Signal Transduction; STAT5 Transcription Factor; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Inflammatory bowel disease is characterized by dysregulated immune responses in inflamed intestine, with dominance of interleukin-17 (IL-17)--producing cells and deficiency of regulatory T (Treg) cells. The aim of this study was to investigate the effect and mechanisms of sirolimus, an inhibitor of the mammalian target of rapamycin, on immune responses in a murine model of Crohn's disease. Murine colitis was induced by intrarectal administration of 2,4,6-trinitrobenzene sulphonic acid at day 0. Mice were then treated intraperitoneally with sirolimus daily for 3 days. The gross and histological appearances of the colon and the numbers, phenotype and cytokine production of lymphocytes were compared with these characteristics in a control group. Sirolimus treatment significantly decreased all macroscopic, microscopic and histopathological parameters of colitis that were analysed. The therapeutic effects of sirolimus were associated with a down-regulation of pro-inflammatory cytokines tumour necrosis factor-α, IL-6 and IL-17A. Intriguingly, sirolimus administration resulted in a prominent up-regulation of the regulatory cytokine transforming growth factor-β. Supporting the hypothesis that sirolimus directly affects the functional activity of CD4+ CD25+ Treg cells, we observed a remarkable enhancement of FoxP3 expression in colon tissues and isolated CD4+ T cells of sirolimus-treated mice. Simultaneously, sirolimus treatment led to a significant reduction in the number of CD4+ IL-17A+ T cells in the mesenteric lymph node cells as well as IL-17A production in mesenteric lymph node cells. Therefore, sirolimus may offer a promising new therapeutic strategy for the treatment of inflammatory bowel disease.

Topics: Animals; Anti-Inflammatory Agents; CD4 Lymphocyte Count; Cells, Cultured; Colitis; Colon; Disease Models, Animal; Forkhead Transcription Factors; Immunosuppressive Agents; Inflammation Mediators; Injections, Intraperitoneal; Interleukin-17; Interleukin-6; Lymph Nodes; Male; Mice; Mice, Inbred BALB C; Phenotype; Sirolimus; Th17 Cells; Time Factors; Transforming Growth Factor beta; Trinitrobenzenesulfonic Acid; Tumor Necrosis Factor-alpha

Fibrosis represents a major complication of several chronic diseases, including inflammatory bowel disease (IBD). Treatment of IBD remains a clinical challenge despite several recent therapeutic advances. Melanin-concentrating hormone (MCH) is a hypothalamic neuropeptide shown to regulate appetite and energy balance. However, accumulating evidence suggests that MCH has additional biological effects, including modulation of inflammation. In the present study, we examined the efficacy of an MCH-blocking antibody in treating established, dextran sodium sulfate-induced experimental colitis. Histological and molecular analysis of mouse tissues revealed that mice receiving anti-MCH had accelerated mucosal restitution and lower colonic expression of several proinflammatory cytokines, as well as fibrogenic genes, including COL1A1. In parallel, they spared collagen deposits seen in the untreated mice, suggesting attenuated fibrosis. These findings raised the possibility of perhaps direct effects of MCH on myofibroblasts. Indeed, in biopsies from patients with IBD, we demonstrate expression of the MCH receptor MCHR1 in α-smooth muscle actin(+) subepithelial cells. CCD-18Co cells, a primary human colonic myofibroblast cell line, were also positive for MCHR1. In these cells, MCH acted as a profibrotic modulator by potentiating the effects of IGF-1 and TGF-β on proliferation and collagen production. Thus, by virtue of combined anti-inflammatory and anti-fibrotic effects, blocking MCH might represent a compelling approach for treating IBD.

Topics: Actins; Animals; Biomarkers; Cell Line; Cell Proliferation; Colitis; Collagen; Colonic Diseases; Fibrosis; Hypothalamic Hormones; Insulin-Like Growth Factor I; Male; Melanins; Mice; Myofibroblasts; Pituitary Hormones; Real-Time Polymerase Chain Reaction; Receptors, Somatostatin; Transforming Growth Factor beta; Up-Regulation; Wound Healing

Growth factors such as keratinocyte growth factor-2 (KGF-2) and transforming growth factor-beta (TGF-β) are important immunoregulatory and epithelial growth factors. They are also potential therapeutic proteins for inflammatory bowel disease. However, owing to protein instability in the upper gastrointestinal tract, it is difficult to achieve therapeutic levels of these proteins in the injured colon when given orally. Furthermore, the short half-life necessitates repeated dosage with large amounts of the growth factor, which may have dangerous side effects, hence the importance of temporal and spatial control of growth factor delivery.. The human commensal gut bacterium, Bacteroides ovatus, was genetically engineered to produce human KGF-2 or TGF-β1 (BO-KGF or BO-TGF) in a regulated manner in response to the dietary polysaccharide, xylan. The successful application of BO-KGF or BO-TGF in the prevention of dextran sodium sulphate induced murine colitis is presented here.. This novel drug delivery system had a significant prophylactic effect, limiting the development of intestinal inflammation both clinically and histopathologically. The ability to regulate heterologous protein production by B ovatus using xylan is both unique and an important safety feature of this drug delivery system.. The use of genetically engineered B ovatus for the controlled and localised delivery of epithelial growth promoting and immunomodulatory proteins has potential clinical applications for the treatment of various diseases targeting the colon.

Topics: Animals; Anti-Inflammatory Agents; Bacteroides; Bacteroides Infections; Colitis; Dextran Sulfate; Drug Delivery Systems; Fibroblast Growth Factor 10; Genetic Engineering; Irritants; Male; Mice; Mice, Inbred C57BL; Probiotics; Transforming Growth Factor beta; Xylans

Although the etiology of two major forms of inflammatory bowel disease (IBD), Crohn's disease (CD) and ulcerative colitis (UC) are unknown and evidence suggests that chronic intestinal inflammation is caused by an excessive immune response to mucosal antigens. Previous studies support the role for TGF-β1 through 3 in the initiation and maintenance of tolerance via the induction of regulatory T cells (Tregs) to control intestinal inflammation. Leptin, a satiety hormone produced primarily by adipose tissue, has been shown to increase during colitis progression and is believed to contribute to disease genesis and/or progression.. We investigated the ability of a pegylated leptin antagonist (PG-MLA) to ameliorate the development of chronic experimental colitis.. Compared to vehicle control animals, PG-MLA treatment of mice resulted in an (1) attenuated clinical score; (2) reversed colitis-associated pathogenesis including a decrease in body weight; (3) reduced systemic and mucosal inflammatory cytokine expression; (4) increased insulin levels and (5) enhanced systemic and mucosal Tregs and CD39⁺ Tregs in mice with chronic colitis. The percentage of systemic and mucosal TGF-β1, -β2 and -β3 expressing CD4⁺ T cells were augmented after PG-MLA treatment. The activation of STAT1 and STAT3 and the expression of Smad7 were also reduced after PG-MLA treatment in the colitic mice. These findings clearly suggest that PG-MLA treatment reduces intestinal Smad7 expression, restores TGF-β1-3 signaling and reduces STAT1/STAT3 activation that may increase the number of Tregs to ameliorate chronic colitis.. This study clearly links inflammation with the metabolic hormone leptin suggesting that nutritional status influences immune tolerance through the induction of functional Tregs. Inhibiting leptin activity through PG-MLA might provide a new and novel therapeutic strategy for the treatment of IBD.

Topics: Animals; Antigens, CD; Apyrase; Body Weight; Chronic Disease; Colitis; Down-Regulation; Female; Insulin; Interleukin-10; Intestinal Mucosa; Leptin; Mice; Mice, Inbred C57BL; Mice, Knockout; Polyethylene Glycols; Recombinant Proteins; Signal Transduction; Smad7 Protein; STAT1 Transcription Factor; STAT3 Transcription Factor; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Selenium (Se) is an essential micronutrient that exerts its functions via selenoproteins. Little is known about the role of Se in inflammatory bowel disease (IBD). Epidemiological studies have inversely correlated nutritional Se status with IBD severity and colon cancer risk. Moreover, molecular studies have revealed that Se deficiency activates WNT signaling, a pathway essential to intestinal stem cell programs and pivotal to injury recovery processes in IBD that is also activated in inflammatory neoplastic transformation. In order to better understand the role of Se in epithelial injury and tumorigenesis resulting from inflammatory stimuli, we examined colonic phenotypes in Se-deficient or -sufficient mice in response to dextran sodium sulfate (DSS)-induced colitis, and azoxymethane (AOM) followed by cyclical administration of DSS, respectively. In response to DSS alone, Se-deficient mice demonstrated increased morbidity, weight loss, stool scores, and colonic injury with a concomitant increase in DNA damage and increases in inflammation-related cytokines. As there was an increase in DNA damage as well as expression of several EGF and TGF-β pathway genes in response to inflammatory injury, we sought to determine if tumorigenesis was altered in the setting of inflammatory carcinogenesis. Se-deficient mice subjected to AOM/DSS treatment to model colitis-associated cancer (CAC) had increased tumor number, though not size, as well as increased incidence of high grade dysplasia. This increase in tumor initiation was likely due to a general increase in colonic DNA damage, as increased 8-OHdG staining was seen in Se-deficient tumors and adjacent, non-tumor mucosa. Taken together, our results indicate that Se deficiency worsens experimental colitis and promotes tumor development and progression in inflammatory carcinogenesis.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Azoxymethane; Carcinogenesis; Colitis; Colonic Neoplasms; Deoxyguanosine; Dextran Sulfate; Diet; DNA Damage; Epidermal Growth Factor; Gene Expression Regulation; Inflammation; Mice; Mice, Inbred C57BL; Selenium; Signal Transduction; Transforming Growth Factor beta; Weight Loss

TGF-β is widely held to be critical for the maintenance and function of regulatory T (T(reg)) cells and thus peripheral tolerance. This is highlighted by constitutive ablation of TGF-β receptor (TR) during thymic development in mice, which leads to a lethal autoimmune syndrome. Here we describe that TGF-β-driven peripheral tolerance is not regulated by TGF-β signalling on mature CD4⁺ T cells. Inducible TR2 ablation specifically on CD4⁺ T cells did not result in a lethal autoinflammation. Transfer of these TR2-deficient CD4⁺ T cells to lymphopenic recipients resulted in colitis, but not overt autoimmunity. In contrast, thymic ablation of TR2 in combination with lymphopenia led to lethal multi-organ inflammation. Interestingly, deletion of TR2 on mature CD4⁺ T cells does not result in the collapse of the T(reg) cell population as observed in constitutive models. Instead, a pronounced enlargement of both regulatory and effector memory T cell pools was observed. This expansion is cell-intrinsic and seems to be caused by increased T cell receptor sensitivity independently of common gamma chain-dependent cytokine signals. The expression of Foxp3 and other regulatory T cells markers was not dependent on TGF-β signalling and the TR2-deficient T(reg) cells retained their suppressive function both in vitro and in vivo. In summary, absence of TGF-β signalling on mature CD4⁺ T cells is not responsible for breakdown of peripheral tolerance, but rather controls homeostasis of mature T cells in adult mice.

Topics: Animals; Autoimmunity; Cell Proliferation; Colitis; Gene Deletion; Homeostasis; Inflammation; Integrases; Lymphopenia; Mice; Mice, Inbred C57BL; NIH 3T3 Cells; Receptors, Antigen, T-Cell; Reproducibility of Results; Signal Transduction; T-Lymphocytes, Regulatory; Tamoxifen; Thymus Gland; Transforming Growth Factor beta

There are few mouse models that adequately mimic large bowel cancer in humans or the gastrointestinal inflammation which frequently precedes it. Dextran sodium sulphate (DSS)-induces colitis in many animal models and has been used in combination with the carcinogen azoxymethane (AOM) to induce cancer in mice. Smad3(-/-) mice are deficient in the transforming growth factor beta (TGFβ) signaling molecule, SMAD3, resulting in dysregulation of the cellular pathway most commonly affected in human colorectal cancer, and develop inflammation-associated colon cancer. Previous studies have shown a requirement for a bacterial trigger for the colitis and colon cancer phenotype in Smad3(-/-) mice. Studies presented here in Smad3(-/-) mice detail disease induction with DSS, without the use of AOM, and show a) Smad3(-/-) mice develop a spectrum of lesions ranging from acute and chronic colitis, crypt herniation, repair, dysplasia, adenomatous polyps, disseminated peritoneal adenomucinosis, adenocarcinoma, mucinous adenocarcinoma (MAC) and squamous metaplasia; b) the colon lesions have variable galactin-3 (Mac2) staining c) increased DSS concentration and duration of exposure leads to increased severity of colonic lesions; d) heterozygosity of SMAD3 does not confer increased susceptibility to DSS-induced disease and e) disease is partially controlled by the presence of T and B cells as Smad3(-/-) Rag2(-/-) double knock out (DKO) mice develop a more severe disease phenotype. DSS-induced disease in Smad3(-/-) mice may be a useful animal model to study not only inflammation-driven MAC but other human diseases such as colitis cystica profunda (CCP) and pseudomyxomatous peritonei (PMP).

Topics: Animals; Colitis; Colorectal Neoplasms; Dextran Sulfate; DNA-Binding Proteins; Humans; Inflammation; Mice; Mice, Knockout; Smad3 Protein; Transforming Growth Factor beta

To determine the effects of heat-killed VSL#3 (B. breve, B. longum and B. infantis; L. plantarum, L. bulgaricus, L. casei and L. acidophilus; S. salivarius subsp. thermophilus) therapy in the dextran sulfate sodium (DSS)-induced acute experimental colitis in rats. Acute experimental colitis was induced in rats by 5% DSS and freely drink for seven days. Beginning on Day 8, rats underwent gavage once daily for seven days with heat-killed probiotic VSL#3 (0.6 g/kg/day), colonic damage was evaluated histologically and biochemically seven days after gavage. Expression of inflammatory related mediators (STAT3, P-STAT3) and cytokines (IL-6, IL-23, TGFβ) in colonic tissue were detected. The results revealed that heat-killed and live VSL#3 have identical anti-inflammatory properties by the assessed DAI (disease activity index), colon length, histological tissue and MPO activity. Heat-killed and live VSL#3 results in reduced IL-6, IL-23, TGFβ, STAT3 and P-STAT3 expression in colonic tissue. Heat-killed and live VSL#3 have showed the similar anti-inflammatory activity by inhibiting IL-6/STAT3 pathway in the DSS-induced acute experimental colitis in rats.

Topics: Acute Disease; Animals; Colitis; Dextran Sulfate; Disease Models, Animal; Hot Temperature; Interleukin-23; Interleukin-6; Male; Mesalamine; Phosphorylation; Probiotics; Rats; Rats, Sprague-Dawley; RNA, Messenger; Severity of Illness Index; STAT3 Transcription Factor; Transforming Growth Factor beta

The FVB.mdr1a(-/-) mouse, lacking the small molecule pump P-glycoprotein (P-gp), is a commonly used model for the study of spontaneous T cell-mediated colitis. In addition, MDR1 polymorphisms and P-gp deficiency in humans have been linked to the development of ulcerative colitis. We now demonstrate that mice with P-gp deficiency have decreased levels of Foxp3(+) regulatory T cells (Tregs) in the intestinal lamina propria. This decrease is not due to either increased Treg apoptosis, altered Treg trafficking, or enhanced Treg plasticity to become Foxp3(+)IL-17(+) cells. Instead, P-gp deficiency appears to restrict the development of induced Treg cells (iTregs), as fewer Foxp3(+) iTregs developed from naive FVB.mdr1a(-/-) T cells both upon transforming growth factor-β (TGF-β) treatment in vitro and after adoptive transfer into FVB.rag2(-/-) recipients. Rather, in vitro TGF-β treatment results in a IL-17(+)CD4(+) T cell. This failure of iTregs to develop explains the decrease in Foxp3(+) Tregs in the FVB.mdr1a(-/-) intestine, representing a need to investigate this novel disease mechanism in human inflammatory bowel disease patients with MDR1 polymorphisms.

Topics: Animals; Apoptosis; ATP Binding Cassette Transporter, Subfamily B; CD4 Antigens; Cell Movement; Colitis; Disease Models, Animal; Forkhead Transcription Factors; Interleukin-2 Receptor alpha Subunit; Intestinal Mucosa; Intestines; Male; Mice; Mice, Knockout; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Although the T-helper type 9 (Th9) subset has recently been revisited, interleukin (IL)-9-producing invariant natural killer T (iNKT) cells remain poorly characterized. Moreover, whether IL-9-producing iNKT cells regulate colitis is unknown. Here, we investigated functions of IL-9-producing iNKT cells in dextran sulfate sodium (DSS)-induced colitis. Wild-type (WT) mice attenuated colitis compared to Jα18(-/-) mice, which were restored by the adoptive transfer of WT, but not IL-4-deficient iNKT cells. IL-4-deficient iNKT cells failed to produce IL-9, which was reversed by recombinant IL-4. Furthermore, iNKT cells, pre-incubated with anti-CD3+CD28 monoclonal antibodies and IL-4+tumor growth factor (TGF)-β (IL-9(+) iNKT), suppressed colitis in Jα18(-/-) mice, whereas pre-incubated IL-4-deficient iNKT cells did not. IL-9 blockade reversed IL-9(+) iNKT cell-mediated colitis by increasing colonic IL-17A and interferon (IFN)-γ transcripts, but decreasing IL-9, IL-10, TGF-β, PU.1, IFN regulatory factor 4, and signal transducer and activator of transcription 5 in Jα18(-/-) mice. In conclusion, IL-9-producing iNKT cells protect against DSS-induced colitis through IFN-γ and IL-17A suppression, but IL-10 and TGF-β enhancement, depending on the IL-4 production by iNKT cells.

Topics: Animals; Colitis; Dextran Sulfate; Interferon-gamma; Interleukin-10; Interleukin-17; Interleukin-4; Interleukin-9; Mice; Mice, Knockout; Natural Killer T-Cells; Receptors, Antigen, T-Cell; Transforming Growth Factor beta

Tributyrin (TBT) is a TAG composed of three butyric acids that has beneficial effects on ulcerative colitis due to its trophic, anti-inflammatory, pro-apoptotic and anti-carcinogenic properties. The goal of the present study was to evaluate the efficacy and mechanisms of action of TBT supplementation in the prevention of mucosal damage in experimental colitis. Mice received either a control diet or a TBT-supplemented diet for 15 d. Colitis was induced by dextran sodium sulphate administration during the last 7 d. Mucosal damage and the activation of immune cells and cytokines were determined by histological score, flow cytometry and ELISA. Leucocyte rolling and adhesion were assessed by intravital microscopy. Oxidative stress was determined by monitoring hydroperoxide concentration and evaluating superoxide dismutase (SOD) and catalase activities. Intestinal permeability was analysed using diethylenetriaminepentaacetate acid (99mTcDTPA). Compared with the colitis group, the animals in the colitis+TBT group had reduced mucosal damage and neutrophil and eosinophil mucosal infiltration, which were associated with a higher percentage of regulatory T cells (Treg) and higher levels of transforming growth factor β and IL-10 in the lamina propria. The level of in vivo leucocyte adhesion in the colon microvasculature was reduced after TBT supplementation. A lower level of hydroperoxide and higher levels of SOD and catalase activities were associated with TBT supplementation. TBT-supplemented mice showed reduced intestinal permeability to the levels intermediate between the control and colitis groups. In conclusion, the present results show that TBT has positive effects on colonic restructuring in experimental colitis. Additionally, TBT supplementation changes the immune response by controlling inflammation and regulating the expression of anti-inflammatory cytokines and Treg.

Topics: Animals; Anti-Inflammatory Agents; Antioxidants; Colitis; Colon; Dietary Supplements; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Interleukin-10; Intestinal Mucosa; Male; Mice; Mice, Inbred C57BL; Oxidative Stress; Superoxide Dismutase; T-Lymphocytes, Regulatory; Transforming Growth Factor beta; Triglycerides

To study the effect of recombinant Lactobacillus casei (L.casei) expressing interleukin (IL)-10 combined with 5-aminosalicylic acid (5-ASA) in dextran sulfate sodium (DSS)-induced colitis mice.. Recombinant L. casei CECT 5276, which can secrete IL-10, was constructed. The length of colon tissue, disease activity index (DAI) and histological score (HS) of the mice were determined to evaluate the modeling and the effectiveness of L. casei. Real-time polymerase chain reaction (PCR), Western blot and ELISA were used to determine the levels of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), peroxisome proliferator-activated receptor (PPAR)-γ, interferon (IFN)-γ, transforming growth factor (TGF)-β and IL-10.. Recombinant L. casei expressing IL-10 combined with 5-ASA was more effective than L. casei with 5-ASA. Among the three different concentrations of the recombinant L. casei, the highest concentration group (2 × 10(9) colony-forming units/mL) had the best effectiveness.. Recombinant L. casei combined with 5-ASA is effective in the treatment of DSS-induced colitis. The possible mechanism might be the blocking of the excessive activation of NF-κB pathway, thus suppressing the release of inflammation-related factors.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Colitis; Colon; Dextran Sulfate; Female; Interferon-gamma; Interleukin-10; Lacticaseibacillus casei; Mesalamine; Mice; Mice, Inbred BALB C; NF-kappa B; PPAR gamma; Recombinant Proteins; Severity of Illness Index; Transforming Growth Factor beta

Bone marrow mesenchymal stem cells (MSCs) comprise a heterogeneous population of postnatal progenitor cells with profound immunomodulatory properties, such as upregulation of Foxp3(+) regulatory T cells (Tregs) and downregulation of Th17 cells. However, it is unknown whether different MSC subpopulations possess the same range of immunomodulatory function. Here, we show that a subset of single colony-derived MSCs producing IL-17 is different from bulk MSC population in that it cannot upregulate Tregs, downregulate Th17 cells, or ameliorate disease phenotypes in a colitis mouse model. Mechanistically, we reveal that IL-17, produced by these MSCs, activates the NFκB pathway to downregulate TGF-β production in MSCs, resulting in abolishment of MSC-based immunomodulation. Furthermore, we show that NFκB is able to directly bind to TGF-β promoter region to regulate TGF-β expression in MSCs. Moreover, these IL-17(+) MSCs possess anti-Candida albicans growth effects in vitro and therapeutic effect in C. albicans-infected mice. In summary, this study shows that MSCs contain an IL-17(+) subset capable of inhibiting C. albicans growth, but attenuating MSC-based immunosuppression via NFκB-mediated downregulation of TGF-β.

Topics: Adult; Animals; Bone Marrow Cells; Candida albicans; Candidiasis; Cells, Cultured; Colitis; Disease Models, Animal; Female; Humans; Immunosuppression Therapy; Interleukin-17; Mesenchymal Stem Cell Transplantation; Mesenchymal Stem Cells; Mice; Mice, Inbred C57BL; NF-kappa B; Promoter Regions, Genetic; Recombinant Proteins; RNA Interference; RNA, Small Interfering; T-Lymphocytes, Regulatory; Th17 Cells; Transforming Growth Factor beta

The gut mucosa hosts large numbers of activated lymphocytes that are exposed to stimuli from the diet, microbiota and pathogens. Although CD4(+) T cells are crucial for defense, intestinal homeostasis precludes exaggerated responses to luminal contents, whether they are harmful or not. We investigated mechanisms used by CD4(+) T cells to avoid excessive activation in the intestine. Using genetic tools to label and interfere with T cell-development transcription factors, we found that CD4(+) T cells acquired the CD8-lineage transcription factor Runx3 and lost the CD4-lineage transcription factor ThPOK and their differentiation into the T(H)17 subset of helper T cells and colitogenic potential, in a manner dependent on transforming growth factor-β (TGF-β) and retinoic acid. Our results demonstrate considerable plasticity in the CD4(+) T cell lineage that allows chronic exposure to luminal antigens without pathological inflammation.

Topics: Animals; CD4-Positive T-Lymphocytes; CD8 Antigens; Cell Differentiation; Cells, Cultured; Citrobacter rodentium; Colitis; Core Binding Factor Alpha 3 Subunit; Enterobacteriaceae Infections; Homeodomain Proteins; Inflammation; Intestinal Mucosa; Intestines; Lymphocyte Activation; Mice; Mice, Inbred C57BL; Mice, Knockout; Signal Transduction; Tamoxifen; Transcription Factors; Transforming Growth Factor beta; Tretinoin

Interleukin (IL)-25, a Th2-related factor, inhibits the synthesis of inflammatory cytokines by macrophages and attenuates experimental colitis in mice. The mechanism underlying the counterregulatory effect of IL-25, however, remains unknown. Since Th2-cytokines can abrogate inflammatory pathways by inducing alternatively activated macrophages (AAMs), we evaluated whether AAMs are involved in the IL-25-mediated anticolitic effect.. AAM-related markers were evaluated in peritoneal and lamina propria mononuclear cells of mice with or without 2,4,6-trinitrobenzenesulphonic acid (TNBS)-induced colitis treated with IL-25 and/or neutralizing IL-4, IL-13, and transforming growth factor beta 1 (TGF-β1) antibodies. Peritoneal AAMs induced in vivo by injecting mice with IL-25 were transferred to mice with TNBS colitis. Finally, we assessed the in vitro effect of IL-25 on the alternative activation of peritoneal F4/80+ cells.. IL-25 enhanced the expression of AAM-related markers in F4/80(+) cells infiltrating the peritoneum and colon of naïve and colitic mice. Peritoneal F4/80(+) cells isolated from IL-25-treated mice reduced the severity of TNBS colitis when injected intraperitoneally to recipient mice. Since IL-25 did not directly induce AAM in vitro and in vivo in mice, IL-25 administration enhanced the expression of IL-4, IL-13, and TGF-β1, which are known to promote AAM differentiation, we finally assessed whether such cytokines were involved in the IL-25-driven AAM induction. Blockade of IL-4, IL-13, and TGF-β1 with neutralizing antibodies in mice did not inhibit the stimulatory effect of IL-25 on AAM gene expression.. The IL-25-mediated anticolitic effect is associated with induction of AAMs, a subset of macrophages with antiinflammatory properties.

Topics: Analysis of Variance; Animals; Antibodies, Neutralizing; Antigens, Differentiation; Arginase; beta-N-Acetylhexosaminidases; Cell Count; Colitis; Gene Expression; Intercellular Signaling Peptides and Proteins; Interleukin-13; Interleukin-17; Interleukin-4; Intestinal Mucosa; Lectins; Macrophage Activation; Macrophages, Peritoneal; Mice; Statistics, Nonparametric; Transforming Growth Factor beta; Trinitrobenzenesulfonic Acid

Crohn's disease causes intestinal inflammation leading to intestinal fibrosis. Spironolactone is an antifibrotic medication commonly used in heart failure to reduce mortality. We examined whether spironolactone is antifibrotic in the context of intestinal inflammation.. In vitro, spironolactone repressed fibrogenesis in transforming growth factor beta (TGF-β)-stimulated human colonic myofibroblasts. However, spironolactone therapy significantly increased mortality in two rodent models of inflammation-induced intestinal fibrosis, suggesting spironolactone could be harmful during intestinal inflammation. Since inflammatory bowel disease (IBD) patients rarely receive spironolactone therapy, we examined whether spironolactone use was associated with mortality in a common cause of inflammatory colitis, Clostridium difficile infection (CDI).. Spironolactone use during CDI infection was associated with increased mortality in a retrospective cohort of 4008 inpatients (15.9% vs. 9.1%, n = 390 deaths, P < 0.0001). In patients without liver disease, the adjusted odds ratio (OR) for inpatient mortality associated with 80 mg spironolactone was 1.99 (95% confidence interval [CI]: 1.51-2.63) In contrast to the main effect of spironolactone mortality, multivariate modeling revealed a protective interaction between liver disease and spironolactone dose. The adjusted OR for mortality after CDI was 1.96 (95% CI: 1.50-2.55) for patients without liver disease on spironolactone vs. 1.28 (95% CI: 0.82-2.00) for patients with liver disease on spironolactone when compared to a reference group without liver disease or spironolactone use.. We propose that discontinuation of spironolactone in patients without liver disease during CDI could reduce hospital mortality by 2-fold, potentially reducing mortality from CDI by 35,000 patients annually across Europe and the U.S.

Topics: Animals; Clostridioides difficile; Clostridium Infections; Colitis; Crohn Disease; Female; Fibrosis; Hospitalization; Humans; Inflammation; Intestinal Diseases; Male; Middle Aged; Mineralocorticoid Receptor Antagonists; Myofibroblasts; Rats; Retrospective Studies; Spironolactone; Survival Rate; Transforming Growth Factor beta; Trinitrobenzenesulfonic Acid

Transforming growth factor beta (TGF-β)- and Interleukin-2 (IL-2)-mediated signaling enables the generation and expansion of induced regulatory T (iTreg) cells that carry high hopes for the treatment of chronic inflammatory and autoimmune diseases. Knowledge about factors stabilizing their lineage commitment and lifespan, however, is limited. Here, we investigated the behavior of iTreg cells, derived from apoptosis-defective mouse mutants, during activated cell autonomous cell death, triggered by cytokine-deprivation, or activation-induced cell death (AICD) after restimulation of the T-cell receptor, and compared these responses with those of effector T cells. We observed that iTreg cells were much more sensitive to IL-2-deprivation but poorly susceptible to AICD. In fact, when apoptosis was compromised, T-cell receptor (TCR)-religation resulted in methylation-independent, ERK- and PI3K/mTOR-mediated loss of Foxp3 expression, impaired suppressive capacity and effector cytokine production. Although iTreg cells prevented colitis induction they rapidly lost Foxp3-GFP expression and gained ability to produce effector cytokines thereby imposing Th1 cell fate on resident effector cells. Surprisingly, iTreg cell conversion itself was limited by TGF-β-mediated Bim/Bcl2L11-dependent apoptosis. Hence, the very same cytokine that drives the generation of iTreg cells can trigger their demise. Our results provide novel insights in iTreg cell biology that will assist optimization of iTreg-based therapy.

Topics: Animals; Apoptosis; Cell Survival; Colitis; Fas Ligand Protein; fas Receptor; Forkhead Transcription Factors; Interleukin-2; Mice; Mice, Inbred C57BL; Mice, Transgenic; Peptide Fragments; Proto-Oncogene Proteins; Receptors, Antigen, T-Cell; Signal Transduction; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Transforming growth factor (TGF)-β signaling, which is down-regulated by the E3 ubiquitin ligase Smad ubiquitin regulating factor 2 (Smurf2), promotes development of cancer. We identified a splice variant of Smurf2 (ΔE2Smurf2) and investigated its role in colon carcinogenesis in mice.. Colitis-associated colon cancer was induced in mice by administration of azoxymethane, followed by 3 cycles of oral administration of dextran sodium sulfate. Messenger RNA levels of Smurf2 in colon tumors and control tissue were measured by quantitative polymerase chain reaction; lymphocyte and cytokine levels were measured in tumor and tissue samples.. Tumor-infiltrating CD4(+) cells expressed higher levels of ΔE2Smurf2 than CD4(+) cells from nontumor tissues of wild-type mice. T cell-specific overexpression of ΔE2Smurf2 increased TGF-β signaling by suppressing protein levels of Smurf2, accompanied by an increase in levels of TGF-β receptor type II. Transgenic mice that overexpress ΔE2Smurf2 were protected against development of colitis-associated tumors and down-regulated proinflammatory cytokines such as interleukin-6. Patients with chronic inflammatory bowel disease had a significantly lower ratio of Smurf2/ΔE2Smurf2 than control individuals.. T cell-specific ΔE2Smurf2 degrades wild-type Smurf2 and controls intestinal tumor growth in mice by up-regulating TGF-β receptor type II, reducing proliferation and production of proinflammatory cytokines.

Topics: Animals; Cells, Cultured; Colitis; Colonic Neoplasms; Gene Expression Profiling; Hyaluronan Receptors; Mice; Mice, Inbred C57BL; Proto-Oncogene Proteins c-kit; Receptors, G-Protein-Coupled; Signal Transduction; Transforming Growth Factor beta; Ubiquitin-Protein Ligases

Systemic infusion of bone marrow mesenchymal stem cells (BMMSCs) yields therapeutic benefit for a variety of autoimmune diseases, but the underlying mechanisms are poorly understood. Here we show that in mice systemic infusion of BMMSCs induced transient T cell apoptosis via the FAS ligand (FASL)-dependent FAS pathway and could ameliorate disease phenotypes in fibrillin-1 mutated systemic sclerosis (SS) and dextran-sulfate-sodium-induced experimental colitis. FASL(-/-) BMMSCs did not induce T cell apoptosis in recipients, and could not ameliorate SS and colitis. Mechanistic analysis revealed that FAS-regulated monocyte chemotactic protein 1 (MCP-1) secretion by BMMSCs recruited T cells for FASL-mediated apoptosis. The apoptotic T cells subsequently triggered macrophages to produce high levels of TGFβ, which in turn led to the upregulation of CD4(+)CD25(+)Foxp3(+) regulatory T cells and, ultimately, immune tolerance. These data therefore demonstrate a previously unrecognized mechanism underlying BMMSC-based immunotherapy involving coupling via FAS/FASL to induce T cell apoptosis.

Topics: Animals; Apoptosis; Bone Marrow Cells; Chemokine CCL2; Colitis; Dextran Sulfate; Fas Ligand Protein; fas Receptor; Fibrillin-1; Fibrillins; Humans; Immune Tolerance; Immunotherapy; Mesenchymal Stem Cell Transplantation; Mesenchymal Stem Cells; Mice; Mice, Knockout; Microfilament Proteins; Scleroderma, Systemic; T-Lymphocyte Subsets; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

New therapeutic approaches are needed for inflammatory bowel diseases. A monoclonal antibody against CD3 (anti-CD3) suppresses T-cell-mediated autoimmune diseases such as experimental allergic encephalomyelitis. We explored the effects of anti-CD3 in mice with colitis.. Severe combined immunodeficient mice were given injections of CD4(+)CD45RB(high) T cells to induce colitis. Four weeks later, the mice were given 2 or 5 μg/day of anti-CD3 or hamster immunoglobulin (Ig)G (control), via gavage, for 5 or 10 days. The effect of oral anti-CD3 on cytokine responses was studied by activating T cells using intraperitoneal injections of anti-CD3 monoclonal antibody 2 days after oral administration of the antibody. We collected intestine samples for histology analysis and cells were analyzed by flow cytometry. Cytokines in sera were analyzed by cytometric bead array.. Oral administration of anti-CD3 protected the mice from wasting disease and intestinal inflammation. Analyses of spleen and mesenteric lymph node cells showed no differences in total cell counts, or percentages of CD4(+) and forkhead box P3(+) regulatory T cells, between mice given anti-CD3 or the control immunoglobulin. Colitis therefore was not suppressed by induction of forkhead box P3(+) regulatory T cells, or depletion or limited expansion of T cells. Oral administration of anti-CD3 ameliorated the enteropathy induced by intraperitoneal injection of the antibody. In mice with enteropathy, oral anti-CD3 reduced levels of inflammatory cytokines such as interferon-γ, tumor necrosis factor-α, and interleukin (IL)-6; it also increased levels of the anti-inflammatory cytokines IL-10 and transforming growth factor-β. The effects of oral anti-CD3 required IL-10.. Oral administration of anti-CD3 to mice induces changes in the mucosal immune response that prevent colitis, independent of specific antigen, and reduce T-cell activation in an IL-10-dependent manner. Oral anti-CD3 therefore might be developed for the treatment of patients with inflammatory bowel disease.

Topics: Animals; Antibodies, Monoclonal; CD3 Complex; CD4 Lymphocyte Count; Colitis; Cytokines; Forkhead Transcription Factors; Immunologic Factors; Interferon-gamma; Interleukin-10; Interleukin-17; Interleukin-6; Intestinal Mucosa; Lymph Nodes; Lymphocyte Activation; Mice; Mice, SCID; Spleen; T-Lymphocytes, Regulatory; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha; Wasting Syndrome

Tissue inhibitor of metalloproteinases (TIMP)-3 is an inhibitor of matrix metalloproteinases, which regulates tissue inflammation, damage, and repair. We investigated the role of TIMP-3 in intestinal inflammation in human beings and mice.. We used real-time polymerase chain reaction and flow cytometry to measure levels of TIMP-3 in intestine samples from patients with Crohn's disease (CD) and those without (controls). We also analyzed TIMP-3 levels in lamina propria mononuclear cells (LPMCs) collected from biopsy samples of individuals with or without CD (controls) and then stimulated with transforming growth factor (TGF)-β1, as well as in biopsy samples collected from patients with CD and then incubated with a Smad7 anti-sense oligonucleotide (knock down). LPMCs and biopsy samples from patients with CD were cultured with exogenous TIMP-3 and levels of inflammatory cytokines were measured. We evaluated the susceptibility of wild-type, TIMP-3-knockout (TIMP-3-KO), and transgenic (TIMP-3-Tg) mice to induction of colitis with 2, 4, 6-trinitrobenzene-sulfonic-acid (TNBS), and the course of colitis in recombinase-activating gene-1-null mice after transfer of wild-type or TIMP-3-KO T cells.. Levels of TIMP-3 were reduced in intestine samples from patients with CD compared with controls. Incubation of control LPMCs with TGF-β1 up-regulated TIMP-3; knockdown of Smad7, an inhibitor of TGF-β1, in biopsy samples from patients with CD increased levels of TIMP-3. Exogenous TIMP-3 reduced levels of inflammatory cytokines in CD LPMCs and biopsy samples. TIMP-3-KO mice developed severe colitis after administration of TNBS, whereas TIMP-3-Tg mice were resistant to TNBS-induced colitis. Reconstitution of recombinase-activating gene-1-null mice with T cells from TIMP-3-KO mice increased the severity of colitis, compared with reconstitution with wild-type T cells.. TIMP-3 is down-regulated in inflamed intestine of patients with CD. Its expression is regulated by TGF-β1, and knock-down of Smad7 in intestinal tissues from patient with CD up-regulates TIMP-3. Loss or reduction of TIMP-3 in mice promotes development of colitis.

Topics: Adult; Aged; Amyloid Precursor Protein Secretases; Animals; Cells, Cultured; Colitis; Colitis, Ulcerative; Crohn Disease; Cytokines; Down-Regulation; Gene Knockdown Techniques; Gene Knockout Techniques; Humans; Intestinal Mucosa; Leukocytes, Mononuclear; Mice; Mice, Knockout; Mice, Transgenic; Middle Aged; Oligonucleotides, Antisense; RNA, Messenger; Smad7 Protein; Tissue Inhibitor of Metalloproteinase-3; Transforming Growth Factor beta; Trinitrobenzenesulfonic Acid

Regulatory T cells (T reg cells) are essential for the prevention of autoimmunity throughout life. T reg cell development occurs intrathymically but a subset of T reg cells can also differentiate from naive T cells in the periphery. In vitro, Smad signaling facilitates conversion of naive T cells into T reg cells but results in unstable Foxp3 expression. The TGF-β-Smad response element in the foxp3 locus is located in the CNS1 region in close proximity to binding sites for transcription factors implicated in TCR and retinoic acid signaling. From in vitro experiments it was previously postulated that foxp3 transcription represents a hierarchical process of transcription factor binding in which Smad3 would play a central role in transcription initiation. However, in vitro conditions generate T reg cells that differ from T reg cells encountered in vivo. To address the relevance of Smad3 binding to the CNS1 enhancer in vivo, we generated mice that exclusively lack the Smad binding site (foxp3(CNS1mut)). We show that binding of Smad3 to the foxp3 enhancer is dispensable for T reg cell development in newborn and adult mice with the exception of the gut.

Topics: Animals; Animals, Newborn; Binding Sites; Colitis; Enhancer Elements, Genetic; Forkhead Transcription Factors; Gastrointestinal Tract; Mice; Mice, Mutant Strains; Signal Transduction; Smad3 Protein; T-Lymphocytes, Regulatory; Thymus Gland; Transforming Growth Factor beta

Mice lacking junctional adhesion molecule A (JAM-A, encoded by F11r) exhibit enhanced intestinal epithelial permeability, bacterial translocation, and elevated colonic lymphocyte numbers, yet do not develop colitis. To investigate the contribution of adaptive immune compensation in response to increased intestinal epithelial permeability, we examined the susceptibility of F11r(-/-)Rag1(-/-) mice to acute colitis. Although negligible contributions of adaptive immunity in F11r(+/+)Rag1(-/-) mice were observed, F11r(-/-)Rag1(-/-) mice exhibited increased microflora-dependent colitis. Elimination of T cell subsets and cytokine analyses revealed a protective role for TGF-β-producing CD4(+) T cells in F11r(-/-) mice. Additionally, loss of JAM-A resulted in elevated mucosal and serum IgA that was dependent upon CD4(+) T cells and TGF-β. Absence of IgA in F11r(+/+)Igha(-/-) mice did not affect disease, whereas F11r(-/-)Igha(-/-) mice displayed markedly increased susceptibility to acute injury-induced colitis. These data establish a role for adaptive immune-mediated protection from acute colitis under conditions of intestinal epithelial barrier compromise.

Topics: Adaptive Immunity; Animals; Bacterial Translocation; CD4-Positive T-Lymphocytes; Cell Adhesion Molecules; Colitis; Dextran Sulfate; Epithelium; Female; Flow Cytometry; Gene Expression; Homeodomain Proteins; Immunoglobulin A; Intestinal Mucosa; Intestines; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Permeability; Receptors, Cell Surface; Reverse Transcriptase Polymerase Chain Reaction; Transforming Growth Factor beta

Recent studies have demonstrated dramatic shifts in metabolic supply-and-demand ratios during inflammation, a process resulting in localized tissue hypoxia within inflammatory lesions ("inflammatory hypoxia"). As part of the adaptive immune response, T cells are recruited to sites of inflammatory hypoxia. Given the profound effects of hypoxia on gene regulation, we hypothesized that T-cell differentiation is controlled by hypoxia. To pursue this hypothesis, we analyzed the transcriptional consequences of ambient hypoxia (1% oxygen) on a broad panel of T-cell differentiation factors. Surprisingly, these studies revealed selective, robust induction of FoxP3, a key transcriptional regulator for regulatory T cells (Tregs). Studies of promoter binding or loss- and gain-of-function implicated hypoxia-inducible factor (HIF)-1α in inducing FoxP3. Similarly, hypoxia enhanced Treg abundance in vitro and in vivo. Finally, Treg-intrinsic HIF-1α was required for optimal Treg function and Hif1a-deficient Tregs failed to control T-cell-mediated colitis. These studies demonstrate that hypoxia is an intrinsic molecular cue that promotes FoxP3 expression, in turn eliciting potent anti-inflammatory mechanisms to limit tissue damage in conditions of reduced oxygen availability.

Topics: Animals; Cell Hypoxia; Cell Proliferation; Cells, Cultured; Colitis; Female; Flow Cytometry; Forkhead Transcription Factors; Gene Expression Regulation; Humans; Hypoxia; Hypoxia-Inducible Factor 1, alpha Subunit; Inflammation; Interleukin-1; Intestinal Mucosa; Jurkat Cells; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Reverse Transcriptase Polymerase Chain Reaction; RNA Interference; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Several lines of evidence suggest nuclear factor of activated T-cells (NFAT) to control regulatory T cells: thymus-derived naturally occurring regulatory T cells (nTreg) depend on calcium signals, the Foxp3 gene harbors several NFAT binding sites, and the Foxp3 (Fork head box P3) protein interacts with NFAT. Therefore, we investigated the impact of NFAT on Foxp3 expression. Indeed, the generation of peripherally induced Treg (iTreg) by TGF-β was highly dependent on NFAT expression because the ability of CD4(+) T cells to differentiate into iTreg diminished markedly with the number of NFAT family members missing. It can be concluded that the expression of Foxp3 in TGF-β-induced iTreg depends on the threshold value of NFAT rather than on an individual member present. This is specific for iTreg development, because frequency of nTreg remained unaltered in mice lacking NFAT1, NFAT2, or NFAT4 alone or in combination. Different from expectation, however, the function of both nTreg and iTreg was independent on robust NFAT levels, reflected by less nuclear NFAT in nTreg and iTreg. Accordingly, absence of one or two NFAT members did not alter suppressor activity in vitro or during colitis and transplantation in vivo. This scenario emphasizes an inhibition of high NFAT activity as treatment for autoimmune diseases and in transplantation, selectively targeting the proinflammatory conventional T cells, while keeping Treg functional.

Topics: Adoptive Transfer; Animals; Autoimmune Diseases; Chromatin Immunoprecipitation; Colitis; Cyclosporine; Flow Cytometry; Fluorescent Antibody Technique; Forkhead Transcription Factors; Homeodomain Proteins; Humans; Immunoblotting; Lymphocyte Activation; Mice; NFATC Transcription Factors; T-Lymphocytes; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

SMAD4 is a common mediator of the TGF-beta signaling pathway. One of the members of this pathway, TGF-beta 1, has an important role in controlling gut inflammation in relation to the continuous stimulation of the intestinal microbiota. SMAD4 haploinsufficiency in humans has been linked to juvenile polyposis hereditary hemorrhagic telangiectasia syndrome (JP/HHT; OMIM#17505). Hematochezia and colonic mucosal inflammation suggestive of inflammatory bowel diseases (IBD) have been reported in JP/HHT. Stimulated by recent experience with two affected pediatric patients presented here, we explored the potential role of Smad4 haploinsufficiency in a murine model of colonic inflammation. Smad4(+/-) mice were maintained on a mixed C57/129SvEv background. Chronic colitis was induced with repeated administration of dextran sulfate sodium (DSS) in drinking water. The colonic mucosal microbiota was interrogated by massively parallel pyrosequencing of the bacterial 16S rRNA gene. 66.7% of Smad4(+/-) mice were sensitive to DSS colitis compared to 14.3% of wild type (Chi-Square p=0.036). The augmented colitis was associated with microbiota separation in the Smad4(+/-) mice. Enterococcus and Enterococcus faecalis specifically was increased in abundance in the colitis-prone animals. Smad4 haploinsufficiency can associate with increased susceptibility to large bowel inflammation in mammals with variable penetrance in association with the colonic mucosal microbiota. These findings may reveal implications not only towards colonic inflammation in the setting of SMAD4 haploinsufficiency, but for colorectal cancer as well.

Topics: Animals; Child; Colitis; Dextran Sulfate; Enterococcus; Female; Haploinsufficiency; High-Throughput Nucleotide Sequencing; Humans; Inflammatory Bowel Diseases; Male; Metagenome; Mice; RNA, Ribosomal, 16S; Signal Transduction; Smad4 Protein; Species Specificity; Transforming Growth Factor beta

After the recent discovery of Th17 cells, it was proposed that Th17 responses are involved in the pathogenesis of inflammatory bowel diseases (IBD). CD4(+)CD25(+) regulatory T cells (Treg) are considered to be an attractive tool for the treatment of IBD. Here, we investigated whether Treg are capable of suppressing Th17-mediated colitis.. Naive CD4(+) T cells were transferred into SCID mice with or without Treg. In some experiments, Treg were transferred into recipient mice with established colitis. Mice treated with Treg were injected with an anti-transforming growth factor (TGF)-β mAb or control IgG. Clinical symptoms of colitis, histological changes and cytokine expressions were investigated.. SCID mice transferred with naive CD4(+) T cells developed chronic colitis with significant increases in Th1 and Th17 cytokine expressions in the colon. When Treg were co-transferred with naive CD4(+) T cells, development of colitis was prevented, and Th17 cytokine expressions were markedly reduced. Similarly, when Treg were transferred into mice with established colitis, the colitis was significantly ameliorated in association with dramatic reductions in Th17 cytokine expressions. Injection of anti-TGF-β mAb abolished the Treg-mediated suppression with significant elevations in Th17 cytokine productions.. This adoptive transfer model of colitis was associated with augmented Th1 and Th17 responses, and Treg were capable of suppressing colonic inflammation by downregulating Th17 responses as well as Th1 responses via TGF-β. Consequently, Treg transfer therapy is expected to be efficacious for IBD even if Th17 is involved in the pathogenesis.

Topics: Animals; CD4-Positive T-Lymphocytes; Colitis; Gene Expression Regulation; Interleukin-2 Receptor alpha Subunit; Lymph Nodes; Mesentery; Mice; Mice, Inbred BALB C; Mice, SCID; RNA, Messenger; Spleen; Th1 Cells; Th17 Cells; Transforming Growth Factor beta

Attenuated innate immune responses to the intestinal microbiota have been linked to the pathogenesis of Crohn's disease (CD). Recent genetic studies have revealed that hypofunctional mutations of NLRP3, a member of the NOD-like receptor (NLR) superfamily, are associated with an increased risk of developing CD. NLRP3 is a key component of the inflammasome, an intracellular danger sensor of the innate immune system. When activated, the inflammasome triggers caspase-1-dependent processing of inflammatory mediators, such as IL-1β and IL-18.. In the current study we sought to assess the role of the NLRP3 inflammasome in the maintenance of intestinal homeostasis through its regulation of innate protective processes. To investigate this role, Nlrp3(-/-) and wildtype mice were assessed in the dextran sulfate sodium and 2,4,6-trinitrobenzenesulfonic acid models of experimental colitis.. Nlrp3(-/-) mice were found to be more susceptible to experimental colitis, an observation that was associated with reduced IL-1β, reduced antiinflammatory cytokine IL-10, and reduced protective growth factor TGF-β. Macrophages isolated from Nlrp3(-/-) mice failed to respond to bacterial muramyl dipeptide. Furthermore, Nlrp3-deficient neutrophils exhibited reduced chemotaxis and enhanced spontaneous apoptosis, but no change in oxidative burst. Lastly, Nlrp3(-/-) mice displayed altered colonic β-defensin expression, reduced colonic antimicrobial secretions, and a unique intestinal microbiota.. Our data confirm an essential role for the NLRP3 inflammasome in the regulation of intestinal homeostasis and provide biological insight into disease mechanisms associated with increased risk of CD in individuals with NLRP3 mutations.

Topics: Animals; Apoptosis; Carrier Proteins; Chemotaxis; Colitis; Disease Models, Animal; Furans; Homeostasis; Immunity, Innate; Inflammasomes; Interleukin-10; Interleukin-1beta; Intestines; Mice; NLR Family, Pyrin Domain-Containing 3 Protein; Thiophenes; Transforming Growth Factor beta

Regulatory T (Treg) cells play an important role in the pathogenesis of inflammatory bowel disease (IBD). In the present study, we found that a superagonistic CD28-specific monoclonal antibody (supCD28mAb, D665) could preferentially stimulate expansion of CD4+Foxp3+ Treg cells. Foxp3(EGFP) mice were orally administrated with 3.5% DSS for 5days, and intraperitoneally injected supCD28mAb 1mg/mice in treated group. All of the mice were sacrificed on day 8, and both clinical and histological parameters showed that the severity of colitis was significantly reduced in treated group compared to controls. In treated group, the proportion of CD103, CD152 and CD62L expression on Foxp3+Treg cells in the spleen and mesenteric lymph node were higher than controls. Furthermore, qRT-PCR analysis showed that expression of anti-inflammatory cytokines such as IL-10, TGF-β was significantly increased in treated group. Taken together, our data demonstrated that supCD28mAb targets CD4+Foxp3+Treg cells expansion in vivo, maintains and enhances their regulatory functions, to reduce the damage of colon in dextran sulfate sodium (DSS)-induced mouse colitis by secreting a large amount of IL-10. It represents a major advance towards the therapeutic use of polyclonally activated Treg cells as cellular therapy for treatment of IBD.

Topics: Animals; Antibodies, Monoclonal; CD28 Antigens; Cell Proliferation; Cells, Cultured; Colitis; Dextran Sulfate; Disease Models, Animal; Disease Progression; Forkhead Transcription Factors; Humans; Inflammatory Bowel Diseases; Interleukin-10; Mice; Mice, Inbred C57BL; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

CD4(+) T regulatory cells (T(regs)), which express the Foxp3 transcription factor, play a critical role in the maintenance of immune homeostasis. Here, we show that in mice, T(regs) were most abundant in the colonic mucosa. The spore-forming component of indigenous intestinal microbiota, particularly clusters IV and XIVa of the genus Clostridium, promoted T(reg) cell accumulation. Colonization of mice by a defined mix of Clostridium strains provided an environment rich in transforming growth factor-β and affected Foxp3(+) T(reg) number and function in the colon. Oral inoculation of Clostridium during the early life of conventionally reared mice resulted in resistance to colitis and systemic immunoglobulin E responses in adult mice, suggesting a new therapeutic approach to autoimmunity and allergy.

Topics: Animals; Anti-Bacterial Agents; Cecum; Cells, Cultured; Clostridium; Colitis; Colon; Feces; Forkhead Transcription Factors; Germ-Free Life; Immunity, Innate; Immunoglobulin E; Interleukin-10; Intestinal Mucosa; Intestine, Small; Metagenome; Mice; Mice, Inbred A; Mice, Inbred BALB C; Receptors, Pattern Recognition; Specific Pathogen-Free Organisms; T-Lymphocytes, Helper-Inducer; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Regulatory T (Treg) cells are plastic, but the in vivo mechanisms by which they are converted into foxhead box p3 (Foxp3+) interferon (IFN)-γ+ T cells and whether these converted cells retain the ability to inhibit colitis are not clear.. Foxp3+ Treg cells were generated by culture of naïve CD4+ T cells from Foxp3GFP CBir1 T-cell receptor (TCR) transgenic (Tg) (CBir1-Tg) mice, which are specific for CBir1 flagellin (an immunodominant microbiota antigen), with transforming growth factor-β. Foxp3GFP+ CBir1-Tg Treg cells were isolated by fluorescence-activated cell sorting and transferred into TCRβxδ-/- mice. Colitis was induced by transfer of naïve CBir1-Tg CD4+ T cells into immunodeficient mice.. Microbiota antigen-specific Foxp3+ Treg cells were converted, in the intestine, to IFN-γ+ T-helper (Th)1 cells, interleukin (IL)-17+ Th17 cells, and Foxp3+ T cells that coexpress IFN-γ and/or IL-17. Conversion of Treg cells into IFN-γ-producing Th1 cells and Foxp3+IFN-γ+ T cells required innate cell production of IL-12 in the intestine; blocking IL-12 with an antibody inhibited their conversion to Th1 and Foxp3+IFN-γ+ T cells in the intestines of mice that were recipients of Treg cells. Addition of IL-12, but not IL-23, promoted conversion of Treg cells into Th1 and Foxp3+IFN-γ+ T cells, in vitro. Foxp3+IFN-γ+ T cells had regulatory activity because they suppressed proliferation of naïve T cells, in vitro, and inhibited induction of colitis by microbiota antigen-specific T cells. IFN-γ+ Th1 cells were not converted into Treg cells; Foxp3+IFN-γ+ T cells differentiated into IFN-γ+ but not Foxp3+ T cells.. IL-12 promotes conversion of Treg cells into IFN-γ-expressing cells; Foxp3+IFN-γ+ T cells retain their regulatory functions and develop during the transition of Foxp3+ Treg cells into IFN-γ+ Th1 cells.

Topics: Animals; CD4-Positive T-Lymphocytes; Cell Differentiation; Cell Separation; Cells, Cultured; Colitis; Disease Models, Animal; Flagellin; Flow Cytometry; Forkhead Transcription Factors; Genes, RAG-1; Genes, T-Cell Receptor beta; Genes, T-Cell Receptor delta; Interferon-gamma; Interleukin-12; Interleukin-17; Intestinal Mucosa; Intestines; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Spleen; T-Lymphocytes, Regulatory; Th1 Cells; Th17 Cells; Transforming Growth Factor beta

Mϕs promote tissue injury or repair depending on their activation status and the local cytokine milieu. It remains unclear whether the immunosuppressive effects of transforming growth factor β (TGF-β) serve a nonredundant role in Mϕ function in vivo. We generated Mϕ-specific transgenic mice that express a truncated TGF-β receptor II under control of the CD68 promoter (CD68TGF-βDNRII) and subjected these mice to the dextran sodium sulfate (DSS) model of colitis. CD68TGF-βDNRII mice have an impaired ability to resolve colitic inflammation as demonstrated by increased lethality, granulocytic inflammation, and delayed goblet cell regeneration compared with transgene negative littermates. CD68TGF-βDNRII mice produce significantly less IL-10, but have increased levels of IgE and numbers of IL-33+ Mϕs than controls. These data are consistent with associations between ulcerative colitis and increased IL-33 production in humans and suggest that TGF-β may promote the suppression of intestinal inflammation, at least in part, through direct effects on Mϕ function.

Topics: Animals; Antigens, CD; Antigens, Differentiation, Myelomonocytic; Blotting, Southern; Colitis; Colitis, Ulcerative; Dextran Sulfate; Disease Models, Animal; Flow Cytometry; Immunoglobulin E; Interleukin-33; Interleukins; Macrophages; Mice; Mice, Transgenic; Promoter Regions, Genetic; Receptors, Transforming Growth Factor beta; Transforming Growth Factor beta

Oral thiopurines are effective and widely used in treatment of inflammatory bowel disease (IBD) in humans, although their use is limited due the development of adverse events. Here, we examine the efficacy and toxicity of oral treatment with 6-tioguanine (6-TG) and azathioprine (AZA) in a murine model of IBD.. We induced acute or chronic colitis in BALB/c mice by one or four cycles of 3% dextran sulphate sodium (DSS), respectively. Mice were treated by daily gavages of various dosages of 6-tioguanine, azathioprine, or by phosphate buffered saline (PBS) starting the first day of DSS or after two cycles of DSS, respectively. We monitored the efficacy and toxicity by measuring the weight change and serum alanine aminotransferase (ALT) activity and by disease severity and histology, at the end of the experiment. Moreover, we measured cytokine production after colon fragment cultivation by enzyme-linked immunoabsorbent assay and numbers of apoptotic cells in the spleen by flow cytometry.. 6-TG is effective in the treatment of acute DSS-induced colitis in a dose-dependent manner and 40 μg of 6-TG is significantly more effective in the treatment of acute colitis than both AZA and PBS. This effect is accompanied by decrease of IL-6 and IFN-γ production in colon. We did not observe histological abnormalities in liver samples from control (PBS) or 6-TG treated mice. However, liver samples from most mice treated with AZA showed mild, yet distinct signs of hepatotoxicity. In chronic colitis, all thiopurine derivatives improved colitis, 20 μg of 6-TG per dose was superior. High doses of 6-TG led to significant weight loss at the end of the therapy, but none of the thiopurine derivatives increased levels of serum ALT. Both thiopurine derivatives reduced the proportion of apoptotic T helper cells, but a high production of both IL-6 and TGF-β was observed only in colon of AZA-treated mice.. Use of 6-TG in the treatment of experimental colitis in mice appears superior to AZA administration and placebo. In contrast to 6-TG, the use of AZA resulted in histological liver abnormalities.

Topics: Acute Disease; Alanine Transaminase; Analysis of Variance; Animals; Apoptosis; Azathioprine; Chronic Disease; Colitis; Colon; Dextran Sulfate; Female; Interferon-gamma; Interleukin-6; Liver; Mice; Mice, Inbred BALB C; Models, Animal; T-Lymphocytes, Helper-Inducer; Thioguanine; Transforming Growth Factor beta; Weight Loss

Recently, we reported the identification of a novel class of pregnane-X-receptor (PXR) agonists, solomonsterols A and B, isolated from the marine sponge Theonella swinhoei. Preliminary pharmacological studies demonstrated that these natural compounds are potential leads for the treatment of human disorders characterized by dysregulation of innate immunity. In this article, we describe the first total synthesis of solomonsterol A and its in vivo characterization in animal models of colitis. Using transgenic mice expressing the human PXR, we found that administration of synthetic solomonsterol A effectively protects against development of clinical signs and symptoms of colitis and reduced the generation of TNFα, a signature cytokine for this disorder. In addition, we have provided the first evidence that solomonsterol A might act by triggering the expression of TGFβ and IL-10, potent counter-regulatory cytokines in inflammatory bowel diseases (IBD). Finally, we have shown that solomonsterol A inhibits NF-κB activation by a PXR dependent mechanism. In summary, solomonsterol A is a marine PXR agonist that holds promise in the treatment of inflammation-driven immune dysfunction in clinical settings.

Topics: Animals; Anti-Inflammatory Agents; Aquatic Organisms; Cholanes; Colitis; Colon; Humans; Immunologic Factors; Interleukin-10; Intestinal Mucosa; Lipopolysaccharides; Mice; Mice, Inbred C57BL; Mice, Knockout; NF-kappa B; Pregnane X Receptor; Receptors, Steroid; Sulfuric Acid Esters; Theonella; Transcriptional Activation; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

Mice that are deficient in interleukin (IL)-10 develop colitis, mediated by T-helper (Th)1 and Th17 cells, and IL-10-producing regulatory T (Treg) cells suppress colitis, implicating IL-10 in maintaining mucosal homeostasis. We assessed the relative importance of immunoregulatory IL-10 derived from T cells or from antigen presenting cells (APCs) in development of intestinal inflammation.. CD4(+) cells from germ-free (GF) or specific pathogen-free (SPF) IL-10(-/-) or wild-type mice were injected into IL-10(-/-), Rag2(-/-) mice or Rag2(-/-) mice that express IL-10. After 6-8 weeks, we evaluated inflammation, spontaneous secretion of cytokines from colonic tissue, and mRNA levels of the transcription factor T-bet and the immunoregulatory cytokine transforming growth factor (TGF)-β. CD4(+) T cells were co-cultured with bacterial lysate-pulsed APCs and assayed for cytokine production, FoxP3 expression, and TGF-β-mediated Smad signaling.. CD4(+) cells from GF or SPF IL-10(-/-) or wild-type mice induced more severe colitis and higher levels of inflammatory cytokines in IL-10(-/-), Rag2(-/-) mice than in IL-10-replete, Rag2(-/-) mice. Co-cultures of IL-10(-/-) or wild-type CD4(+) T cells plus bacterial lysate-pulsed APCs from IL-10(-/-) mice contained more interferon (IFN)-γ, IL-12/23p40, and IL-17 than co-cultures of the same T cells plus APCs from wild-type mice. CD11b(+) APCs were required for these effects. Blocking IL-10 receptors increased production of IFN-γ and IL-12/23p40 whereas exogenous IL-10 suppressed these cytokines. IL-10-producing APCs induced TGF-β-mediated, retinoic acid-dependent, differentiation of FoxP3(+) Treg cells, whereas blocking the retinoic acid receptor, in vitro and in vivo, reduced proportions of FoxP3(+) Treg cells.. IL-10 produced by APCs regulates homeostatic T-cell responses to commensal bacteria.

Topics: Adaptive Immunity; Animals; Antigen-Presenting Cells; Antigens, Bacterial; CD11b Antigen; CD4-Positive T-Lymphocytes; Cells, Cultured; Colitis; DNA-Binding Proteins; Forkhead Transcription Factors; Germ-Free Life; Immunity, Innate; Interferon-gamma; Interleukin-10; Interleukin-12; Interleukin-17; Mice; Mice, Knockout; RNA, Messenger; Signal Transduction; Smad Proteins; T-Box Domain Proteins; T-Lymphocytes, Regulatory; Th1 Cells; Th17 Cells; Transforming Growth Factor beta; Tretinoin

Mycobacterium bovis Bacillus Calmette-Guérin (BCG), killed by extended freeze-drying (EFD), induces secretion of interleukin-10 and reduces lung inflammation in a mouse model of asthma. We investigated the effects of EFD BCG in mouse models of inflammatory bowel disease.. EFD BCG was administered subcutaneously to mice with colitis induced by dextran sodium sulfate (DSS), oxazolone, or adoptive transfer of CD4(+)CD45RB(high)Foxp3(-) T cells from C57Bl/6 Foxp3GFP mice to RAG2(-/-) mice.. EFD BCG, administered either before induction of DSS and oxazolone colitis or after development of acute or chronic DSS-induced colitis, reduced symptom scores, loss of body weight, and inflammation. Although transfer of CD4(+)CD45RB(high)Foxp3(-) cells induced colitis in RAG2(-/-) mice, administration of EFD BCG at the time of the transfer converted Foxp3(-) T cells to Foxp3(+) T cells and the mice did not develop colitis. EFD BCG protected mice from colitis via a mechanism that required expansion of T regulatory cells and production of interleukin-10 and transforming growth factor β. EFD BCG activated the retinoid X receptor (RXR)-α-peroxisome proliferator-activated receptor (PPAR)-γ heterodimer, blocked translocation of nuclear factor κB to the nucleus, and reduced colonic inflammation; it did not increase the number of colon tumors that formed in mice with chronic DSS-induced colitis.. EFD BCG controls severe colitis in mice by expanding T regulatory cell populations and PPAR-γ and might be developed to treat patients with inflammatory bowel disease.

Topics: Animals; BCG Vaccine; Colitis; Colon; Dextran Sulfate; Forkhead Transcription Factors; Freeze Drying; Interleukin-1; Interleukin-10; Interleukin-6; Male; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mycobacterium bovis; NF-kappa B; Oxazolone; Peroxidase; PPAR gamma; T-Lymphocytes, Regulatory; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha; Weight Loss

To study the proinflammatory and anti-inflammatory cytokines present in the acute phase of trinitrobenzene sulfonic acid (TNBS)-induced experimental colitis treated with Saccharomyces boulardii.. Thirty male Wistar rats were divided into three groups: (1) treated group--received Saccharomyces boulardii for 14 days; (2) non-treated group--received sodium chloride solution for 14 days; (3) control group. Colitis was induced on the seventh day of the study in the treated and the non-treated groups using TNBS (10 mg) dissolved in 50% ethanol. Quantification of cytokines, including interleukin (IL)-1beta (IL-1beta), IL-6, transforming growth factor-beta (TGF-beta), IL-10 and tumor necrosis factor-alpha (TNF-alpha), in the serum and colonic tissue collected on day 14 were carried out using an enzyme-linked immunosorbent assay (ELISA).. The mean concentrations of TGF-beta in both the serum and the colonic tissue of the treated group were statistically higher than that of the control group. The mean concentration of TGF-beta in the colonic tissue of the non-treated group was also statistically higher than the control group.. The group treated with Saccharomyces boulardii showed increased amounts of TGF-beta, an anti-inflammatory cytokine, during the acute phase of colitis. There were no differences in the amount of TNF-alpha, IL-1beta, IL-6, and IL-10 between the treated and the non-treated or the control groups during the acute phase of experimental colitis induced by TNBS.

Topics: Acute Disease; Animals; Colitis; Colon; Cytokines; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Inflammation Mediators; Interleukin-10; Interleukin-1beta; Interleukin-6; Male; Probiotics; Rats; Rats, Wistar; Saccharomyces; Severity of Illness Index; Time Factors; Transforming Growth Factor beta; Trinitrobenzenesulfonic Acid; Tumor Necrosis Factor-alpha

Epithelial barrier disturbance is thought to contribute to the pathogenesis of inflammatory bowel diseases; however, it remains unclear whether it is a primary defect participating to the onset of inflammation or only a consequence of sustained inflammation.. A time course study of epithelial barrier functions and immune mediators was performed in the CD4(+)CD45RB(hi) T cell transfer model of colitis using Ussing chambers.. In nonreconstituted severe combined immunodeficiency (SCID) mice, no epithelial dysfunction was observed. However, after transfer of CD4(+)CD45RB(hi) T cells or total CD4(+) T cells, colon of SCID mice displayed a decreased epithelial resistance, even before overt microscopic inflammation had occurred. Sustained colitis of CD4(+)CD45RB(hi) T cell reconstituted mice was also associated with enhanced subepithelial resistance, enhanced paracellular permeability, and decreased net ion transport. All these reflect a disturbance of barrier function and may contribute to diarrhea. Epithelial resistance was positively correlated with interleukin 10 (IL-10) and transforming growth factor beta (TGF-beta) levels and net ion transport inversely correlated with tumor necrosis factor alpha (TNF-alpha) levels, pointing to the protective effect of IL-10 and TGF-beta and to a damaging effect of TNF-alpha. Indomethacin, a nonselective COX inhibitor, decreased epithelial resistance independent of T cells and inflammation, but its effect was more pronounced in inflamed colon.. Induction of colitis by transfer of CD4(+)CD45RB(hi) T cells in SCID mice leads to changes in the colonic epithelium before colitis develops. Decreased epithelium resistance might contribute to the development of colitis; however, it is not sufficient to lead to chronic inflammation.

Topics: Adoptive Transfer; Animals; CD4-Positive T-Lymphocytes; Colitis; Female; Indomethacin; Interleukin-10; Intestinal Mucosa; Ion Transport; Leukocyte Common Antigens; Mice; Mice, Inbred BALB C; Mice, SCID; Spleen; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

TGFβ is the quintessential cytokine of T cell homeostasis. TGFβ signaling is required for the efficient differentiation and maintenance of CD4(+)FOXP3(+) T cells that inhibit immune responses. Conversely, in conjunction with the inflammatory cytokine IL-6, TGFβ promotes Th17 cell differentiation. The mechanism by which TGFβ signals synergize with IL-6 to generate inflammatory versus immunosuppressive T cell subsets is unclear. TGFβ signaling activates receptor SMADs, SMAD2 and SMAD3, which associate with a variety of nuclear factors to regulate gene transcription. Defining relative contributions of distinct SMAD molecules for CD4 T cell differentiation is critical for mapping the versatile intracellular TGFβ-signaling pathways that tailor TGFβ activities to the state of host interaction with pathogens. We show here that SMAD2 is essential for Th17 cell differentiation and that it acts in part by modulating the expression of IL-6R on T cells. Although mice lacking SMAD2 specifically in T cells do not develop spontaneous lymphoproliferative autoimmunity, Smad2-deficient T cells are impaired in their response to TGFβ in vitro and in vivo, and they are more pathogenic than controls when transferred into lymphopenic mice. These results demonstrate that SMAD2 is uniquely essential for TGFβ signaling in CD4(+) T effector cell differentiation.

Topics: Animals; CD4-Positive T-Lymphocytes; Cell Differentiation; Cells, Cultured; Colitis; Female; Flow Cytometry; Forkhead Transcription Factors; Interleukin-17; Interleukin-6; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Phosphorylation; Receptors, Interleukin-6; Reverse Transcriptase Polymerase Chain Reaction; Smad2 Protein; STAT3 Transcription Factor; T-Lymphocytes, Helper-Inducer; Transforming Growth Factor beta

Inflammatory bowel disease (IBD) is the most common and serious chronic inflammatory condition of the gut. Among the distinct T helper (Th) cell subsets, a Th1 type response is associated predominantly with Crohn's disease (CD) while helminth infections generate a strong Th2 type response. IBD is most prevalent in developed countries but rare in countries where infections with helminths are common. Thus, it has been hypothesized that infection with helminth infection influence the development of CD and recent clinical and experimental studies suggest strongly a beneficial role of helminth infection in IBD. In the present study we examined the effects of rectal submucosal administration of helminth antigens on subsequent experimental colitis. Mice were treated with Trichinella spiralis antigens prior to the induction of dinitrobenzenesulphonic acid (DNBS)-induced colitis and were killed 3 days post-DNBS to assess colonic damage macroscopically, histologically and by myeloperoxidase (MPO) activity, inducible nitric oxide synthase (iNOS) and cytokine levels. Previous treatment with T. spiralis antigens reduced the severity of colitis significantly, as assessed macroscopically and histologically, and reduced the mortality rate. This benefit was correlated with a down-regulation of MPO activity, interleukin (IL)-1beta production and iNOS expression and an up-regulation of IL-13 and transforming growth factor-beta production in colon. These results clearly show a beneficial role of local treatment with helminth antigens for experimental colitis and prompt consideration of helminth antigen-based therapy for IBD instead of infection with live parasites.

Topics: Animals; Antigens, Helminth; Colitis; Colon; Dinitrofluorobenzene; Injections; Interleukin-13; Interleukin-1beta; Male; Mice; Mice, Inbred C57BL; Models, Animal; Nitric Oxide Synthase Type II; Peroxidase; Rectum; Transforming Growth Factor beta; Trichinella spiralis; Trichinellosis; Vaccination

Cow milk contains a large amount of an immunoregulatory cytokine, transforming growth factor-beta (TGFbeta). The present study investigated whether commercially available pasteurized cow milk retains TGFbeta activity both in vitro and in vivo. Some commercial cow milk increased TGFbeta/Smad-responsive reporter activity and induced Smad2 phosphorylation and the transcription of the TGFbeta/Smad target genes TGFbeta itself and Smad7 in vitro. Mice treated orally with 500 microL of cow milk containing TGFbeta (3 microg/L) daily for 2 wk had increased phosphorylation of Smad2 and TGFbeta and Smad7 mRNA expression in the intestine. These mice also had significantly greater serum TGFbeta concentrations than the mice treated orally with PBS. Furthermore, oral administration of 500 microL of cow milk containing TGFbeta (3 microg/L) daily for 2 wk before the induction of dextran sodium sulfate colitis and lipopolysaccharide-induced endotoxemia ameliorated tissue damage and mortality, respectively, in mice. These in vivo effects of cow milk were abrogated by the simultaneous administration of TGFbeta type I receptor kinase inhibitor with the cow milk, and they were not observed after the oral administration of cow's milk containing little TGFbeta. In humans, 1 oral challenge of 10 mL/kg cow milk containing TGFbeta (3 microg/L) increased the plasma TGFbeta concentrations at 4 h after the challenge. Thus, some commercially available pasteurized cow milk retains TGFbeta activity, which may be able to provide protection against experimental colitis and endotoxemia associated with increased intestinal and circulating TGFbeta levels.

Topics: Adult; Animals; Cattle; Cell Line, Tumor; Colitis; Dextran Sulfate; Female; Humans; Inflammation; Male; Mice; Mice, Inbred BALB C; Middle Aged; Milk; Transforming Growth Factor beta

Foxp3-expressing regulatory T cells (Tregs) play a key role in the maintenance of the gut immune homeostasis, and an intact transforming growth factor (TGF)-beta signaling is required for their function. In inflammatory bowel disease (IBD), the TGF-beta signaling is impaired because of high expression of the inhibitory molecule Smad7. Although no intrinsic defects in Tregs function have been shown in IBD, it is still unknown whether colitogenic T cells are susceptible to Treg-mediated suppression. In this study, we have investigated whether IBD mucosal CD4+ T cells are resistant to Tregs and whether Smad7 is involved in this process.. IBD lamina propria mononuclear cells (LPMC) were cultured with or without Tregs, and proliferation was assessed by flow cytometry. Proliferation of IBD LPMC was also evaluated after Smad7 antisense oligonuclotide treatment. Treg-mediated suppression of T-cell proliferation and proinflammatory cytokine expression was investigated in murine Smad7 transgenic cells. In vivo, the Smad7-dependent resistance of colitogenic naïve T cells to Tregs was studied in the adoptive transfer model of colitis.. IBD LPMC were resistant to Treg-mediated suppression, and this phenomenon was reverted by Smad7 antisense treatment. Consistently, CD4+ T cells isolated from Smad7 transgenic mice showed high proliferation, produced considerable amount of inflammatory cytokines following activation, and induced a severe colitis when transferred in immunodeficient RAG1 knockout mice even in the presence of wild-type Tregs.. Smad7 makes CD4+ T cells resistant to Tregs-mediated suppression thus fine-tuning their proinflammatory potential.

Topics: Animals; CD4-Positive T-Lymphocytes; Cell Communication; Cell Proliferation; Cells, Cultured; Colitis; Colon; Crohn Disease; Disease Models, Animal; Homeodomain Proteins; Humans; Intestinal Mucosa; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Signal Transduction; Smad7 Protein; T-Lymphocytes, Regulatory; Transforming Growth Factor beta; Up-Regulation

Intestinal dendritic cells (DCs) have been shown to display specialized functions, including the ability to promote gut tropism to lymphocytes, to polarize noninflammatory responses, and to drive the differentiation of adaptive Foxp3(+) regulatory T (T(reg)) cells. However, very little is known about what drives the mucosal phenotype of DCs. Here, we present evidence that the local microenvironment, and in particular intestinal epithelial cells (ECs), drive the differentiation of T(reg)-cell-promoting DCs, which counteracts Th1 and Th17 development. EC-derived transforming growth factor-beta (TGF-beta) and retinoic acid (RA), but not thymic stromal lymphopoietin (TSLP), were found to be required for DC conversion. After EC contact, DCs upregulated CD103 and acquired a tolerogenic phenotype. EC-conditioned DCs were capable of inducing de novo T(reg) cells with gut-homing properties that when adoptively transferred, protected mice from experimental colitis. Thus, we have uncovered an essential mechanism in which EC control of DC function is required for tolerance induction.

Topics: Animals; Cell Communication; Cell Differentiation; Colitis; Cytokines; Dendritic Cells; Female; Immune Tolerance; Interleukin-17; Intestinal Mucosa; Lymphocyte Activation; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; T-Lymphocyte Subsets; T-Lymphocytes, Regulatory; Thymic Stromal Lymphopoietin; Transforming Growth Factor beta; Tretinoin

Colonization with helminthic parasites induces mucosal regulatory cytokines, like IL-10 or TGF-beta, that are important in suppressing colitis. Helminths induce mucosal T cell IL-10 secretion and regulate lamina propria mononuclear cell (LPMC) Th1 cytokine generation in an IL-10-dependent manner in WT mice. Helminths also stimulate mucosal TGF-beta release. As TGF-beta exerts major regulatory effects on T lymphocytes, we investigated the role of T lymphocyte TGF-beta signaling in helminthic modulation of intestinal immunity. T cell TGF-beta signaling is interrupted in TGF-beta receptor II dominant negative (TGF-betaRII DN) mice by T-cell-specific over-expression of a TGF-betaRII DN. We studied LPMC responses in WT and TGF-betaRII DN mice that were uninfected or colonized with the nematode, Heligmosomoides polygyrus. Our results indicate an essential role of T cell TGF-beta signaling in limiting mucosal Th1 and Th2 responses. Furthermore, we demonstrate that helminthic induction of intestinal T cell IL-10 secretion requires intact T cell TGF-beta-signaling pathway. Helminths fail to curtail robust, dysregulated intestinal Th1 cytokine production and chronic colitis in TGF-betaRII DN mice. Thus, T cell TGF-beta signaling is essential for helminthic stimulation of mucosal IL-10 production, helminthic modulation of intestinal IFN-gamma generation and H. polygyrus-mediated suppression of chronic colitis.

Topics: Animals; Cells, Cultured; Colitis; Cytokines; Enzyme-Linked Immunosorbent Assay; Flow Cytometry; Host-Parasite Interactions; Interferon-gamma; Interleukin-10; Intestinal Diseases, Parasitic; Intestine, Small; Mice; Mice, Inbred C57BL; Mice, Transgenic; Mutant Proteins; Nematospiroides dubius; Protein Serine-Threonine Kinases; Receptor, Transforming Growth Factor-beta Type II; Receptors, Transforming Growth Factor beta; Signal Transduction; Strongylida Infections; T-Lymphocytes; Transforming Growth Factor beta

Cyclosporine is a potent immunomodulator and has a beneficial effect in the treatment of ulcerative colitis (UC). We analyzed the mechanism of the effects of cyclosporine on the regulation of epithelial apoptosis via TGF-beta-related signaling, because the balance between the apoptosis and regeneration of epithelial cells seems to be a key factor to maintain the intestinal homeostasis. For this purpose, colitis was induced by treatment of 4% dextran sulfate sodium (DSS), and the effect of treatment with cyclosporine and anti-TGF-beta antibody was assessed. Treatment with cyclosporine ameliorated body weight loss, mucosal destruction, and epithelial apoptosis in DSS-induced colitis. Cyclosporine was shown to upregulate the expression of TGF-beta in the colonic tissue, enhance the expression of p-Smad2 and cFLIP in epithelial cells, and inhibit caspase-8 activity but not caspase-1 or -9. Upregulation of cFLIP in the colonic epithelial cells, amelioration of body weight loss, and mucosal destruction by cyclosporine were attenuated by anti-TGF-beta antibody treatment. These results indicated that cyclosporine could have a protective role against epithelial apoptosis associated with upregulation of TGF-beta-related signaling.

Topics: Animals; Antibodies; Apoptosis; Body Weight; CASP8 and FADD-Like Apoptosis Regulating Protein; Caspase 8; Colitis; Colon; Cyclosporine; Dextran Sulfate; Disease Models, Animal; Female; Immunosuppressive Agents; Intestinal Mucosa; Mice; Mice, Inbred C57BL; Phosphorylation; Signal Transduction; Smad2 Protein; Time Factors; Transforming Growth Factor beta; Up-Regulation

TV-5010 is a higher molecular weight version of glatiramer acetate (GA), the active ingredient of Copaxone - an approved drug for the treatment of multiple sclerosis (MS). GA has been shown to ameliorate colitis in several experimental models when administered by daily injection. The present study aimed to explore the effect of TV-5010 in a murine model of inflammatory bowel disease (IBD) and the possibility for its administration in tapered frequency. As demonstrated here, TV-5010 ameliorated the various pathological manifestations of Dextran (DSS)-induced colitis, i.e. weight loss, intestinal bleeding and diarrhea, resulting in substantial reduction of disease activity, colonic damage and mortality. In contrast to GA, which was more effective when administered daily by injection of 2.0 mg/mouse, TV-5010 was most effective when administered once a week, at dose of 0.2-1.0 mg/mouse. TV-5010 treatment abrogated the characteristic inflammation in the diseased organ as demonstrated by reduction in the colonic mRNA expression of TNF-alpha, IFN-gamma and the Th1 transcription factor T-bet, as well as by augmentation of the regulatory cytokine TGF-beta. Injection of naive mice with TV-5010 generated lymphocyte population of the Th2/3 subtype that effectively reduced disease manifestations upon adoptive transfer to mice with DSS-colitis. Moreover, TV-5010-specific T-cells, either exogenously labeled or genetically marked, adoptively transferred to colitis-induced mice, localized in the inner layers of the injured colon and expressed TGF-beta in situ. Thus, TV-5010 is effective in the suppression of experimental colitis similarly to GA, with the advantage of less frequent administration, possibly via immunomodulation at the site of pathological damage.

Topics: Animals; Colitis; Dextrans; Glatiramer Acetate; Inflammatory Bowel Diseases; Interferon-gamma; Lymphocytes; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Transgenic; Peptides; RNA, Messenger; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

Probiotics, defined as live or attenuated bacteria or bacterial products, confer a significant health benefit to the host. Recently, they have been shown to be useful in the treatment of chronic inflammatory bowel disease and infectious colitis. In this study, we investigated the effect of probiotics on the development of experimental colitis using Toll-like receptor 4 (TLR-4) mutant (lps-/lps-) mice. TLR-4(lps-/lps-) and wild-type (WT) mice were given 2.5% dextran sulphate sodium (DSS) in drinking water to induce colitis with or without Lactobacillus casei pretreatment. Clinical and histological activity of DSS-colitis was attenuated markedly both in TLR-4(lps-/lps-) and WT mice pretreated with L. casei. Interestingly, histological activity was less severe in TLR-4(lps-/lps-) mice than in WT mice. The levels of myeloperoxidase activity and interleukin (IL)-12p40 were attenuated in pretreated TLR-4(lps-/lps-) mice after DSS administration. By contrast, transforming growth factor (TGF)-beta and IL-10 mRNA and protein expressions were increased markedly in pretreated TLR-4(lps-/lps-) mice. The current results suggest that L. casei has a preventive effect in the development of acute DSS-induced colitis and its action depends largely upon TLR-4 status. L. casei modulates the expression of inflammatory cytokines and down-regulates neutrophilic infiltration in the case of incomplete TLR-4 complex signalling.

Topics: Animals; Biomarkers; Colitis; Colon; Dextran Sulfate; Enzyme-Linked Immunosorbent Assay; Interleukin-10; Interleukin-12 Subunit p40; Intestinal Mucosa; Lacticaseibacillus casei; Mice; Mice, Inbred BALB C; Mice, Knockout; Models, Animal; Peroxidase; Probiotics; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Toll-Like Receptor 4; Transforming Growth Factor beta

We have reported that commensal luminal bacterial components induce an active in vitro IFN-gamma response in mesenteric lymph node (MLN) and intestinal cells from specific pathogen-free (SPF) HLA-B27 transgenic (TG) rats with chronic colitis but not in cells from non-diseased SPF non-TG, germ-free (GF) non-TG or GF TG rats.. The study examined IL-12 stimulation of MLN IFN-gamma responses to luminal bacteria and regulation of these responses by suppressive cytokines.. Exogenous IL-12 significantly increased the bacterial lysate-induced IFN-gamma response in SPF TG MLN cells, while bacterial lysate and IL-12 synergistically induced IFN-gamma from low baseline levels in cells obtained from both SPF and GF non-TG rats, and in GF TG cells. TGF-beta fully counteracted the effects of IL-12 and bacterial lysate on non-TG cells by almost completely inhibiting IFN-gamma production. In contrast, TG cells were less responsive to TGF-beta-mediated downregulation with a substantial residual IFN-gamma response to IL-12 plus bacterial lysate. Further experiments showed that CD4+/CD25+ cells had no inhibitory effect on the IFN-gamma production and were not required for TGF-beta-mediated suppression. Addition of exogenous IL-10 also partially inhibited IFN-gamma production by non-TG cells but did not affect TG cells. Conversely, exogenous IL-12 preferentially suppressed bacterial lysate-induced TGF-beta and IL-10 production in TG rat cells.. An attenuated response to regulatory signals leads to uncontrolled potentiated induction of effector IFN-gamma responses to commensal bacteria in HLA-B27 TG rats that spontaneously develop chronic intestinal inflammation.

Topics: Animals; Animals, Genetically Modified; Bacteria; Blotting, Western; Cells, Cultured; Colitis; Down-Regulation; Enzyme-Linked Immunosorbent Assay; HLA-B27 Antigen; Interferon-gamma; Interleukin-10; Interleukin-12; Lymph Nodes; Mesentery; Rats; Specific Pathogen-Free Organisms; Transforming Growth Factor beta

Interleukin-23 (IL-23) is an inflammatory cytokine that plays a key role in the pathogenesis of several autoimmune and inflammatory diseases. It orchestrates innate and T cell-mediated inflammatory pathways and can promote T helper 17 (Th17) cell responses. Utilizing a T cell transfer model, we showed that IL-23-dependent colitis did not require IL-17 secretion by T cells. Furthermore, IL-23-independent intestinal inflammation could develop if immunosuppressive pathways were reduced. The frequency of naive T cell-derived Foxp3+ cells in the colon increased in the absence of IL-23, indicating a role for IL-23 in controlling regulatory T cell induction. Foxp3-deficient T cells induced colitis when transferred into recipients lacking IL-23p19, showing that IL-23 was not essential for intestinal inflammation in the absence of Foxp3. Taken together, our data indicate that overriding immunosuppressive pathways is an important function of IL-23 in the intestine and could influence not only Th17 cell activity but also other types of immune responses.

Topics: Adoptive Transfer; Animals; Colitis; Forkhead Transcription Factors; Immune Tolerance; Inflammation Mediators; Interleukin-10; Interleukin-23; Lymphopenia; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Chronic inflammation predisposes to cancer. Transforming growth factor (TGF)-beta, a multifunctional protein, suppresses the growth of normal colonic epithelial cells, whereas it stimulates the proliferation of cancer cells. Interleukin (IL)-10-deficient mice, which develop colitis and colorectal cancer, show an increased level of plasma TGF-beta. Although TGF-beta may be a key molecule in the development of colon cancer arising from chronic colitis in IL-10-deficient mice, the role of TGF-beta still remains unclear. TGF-beta activates not only TGF-beta type I receptor (TbetaRI) but also c-Jun N-terminal kinase (JNK), which converts the mediator Smad3 into two distinctive phosphoisoforms: C-terminally phosphorylated Smad3 (pSmad3C) and linker-phosphorylated Smad3 (pSmad3L). We studied C57BL/6-IL-10-deficient mice (n=18) at 4 to 32 weeks of age. We investigated histology, and pSmad2/3L, pSmad2/3C, and p53 by immunohistochemistry. pSmad3L staining was detected in the cancer cells in all 10 mice with colonic cancer and in the epithelial cells in 7 of 12 mice with colonic dysplasia, but not in the normal or colitic mice. pSmad3c was detected without any significant difference between stages. p53 was weakly stained in a few cancer cells in 5 out of 10 mice. Smad3L signaling plays an important role in the carcinogenesis of chronic colitis in IL-10-deficient mice.

Topics: Animals; Chronic Disease; Colitis; Colorectal Neoplasms; Immunoenzyme Techniques; Interleukin-10; Intestine, Large; JNK Mitogen-Activated Protein Kinases; Mice; Mice, Inbred C57BL; Mice, Knockout; Phosphorylation; Protein Serine-Threonine Kinases; Receptor, Transforming Growth Factor-beta Type II; Receptors, Transforming Growth Factor beta; Signal Transduction; Smad3 Protein; Transforming Growth Factor beta; Tumor Suppressor Protein p53

Filamentous haemagglutinin (FHA) of Bordetella pertussis subverts host immune responses by inhibiting interleukin (IL)12 and enhancing IL10 production by macrophages and dendritic cells, and promoting the induction of regulatory T cells.. Injection of FHA would ameliorate disease in a T cell-dependent model of colitis via the induction of anti-inflammatory cytokines and regulatory T cells.. Colitis was induced by injection of CD4CD45RB(high) naive T cells into severe combined immunodeficient (SCID) mice. Mice were treated with four subcutaneous injections of FHA or buffer alone.. Parenteral injection of FHA stimulated IL10 and/or transforming growth factor beta production in local and mesenteric lymph nodes and Peyer's patches of mice 2-6 h after administration. Compared with phosphate-buffered saline-treated mice, FHA-treated SCID mice had significantly (p<0.01) less weight loss, lower colon weight, less colon shrinkage and reduced inflammatory lesions. The therapeutic effect of FHA was associated with enhanced IL10 and reduced type 1 and type 2 T helper cytokine production by spleen cells. Finally, FHA also attenuated the symptoms of colitis in SCID mice transferred with CD4CD45RB(high) T cells from IL10-deficient mice.. Our finding shows that FHA suppresses type 1 T helper and pro-inflammatory cytokines, and ameliorates disease activity in a chronic T cell-dependent model of colitis, an effect that was not dependent on IL10 production by T cells, but was associated with induction of anti-inflammatory cytokines in vivo. Having already been used as a pertussis vaccine component in children, FHA is a promising candidate for clinical testing in patients with Crohn's disease.

Topics: Adhesins, Bacterial; Animals; Bordetella pertussis; Colitis; Cytokines; Disease Models, Animal; Interleukin-10; Leukocyte Common Antigens; Lymph Nodes; Lymphocyte Transfusion; Mice; Mice, Inbred BALB C; Mice, SCID; Peyer's Patches; Protein Tyrosine Phosphatase, Non-Receptor Type 1; Th1 Cells; Th2 Cells; Transforming Growth Factor beta; Virulence Factors, Bordetella

The propensity of a range of parasitic helminths to stimulate a Th2 or regulatory cell-biased response has been proposed to reduce the severity of experimental inflammatory bowel disease. We examined whether infection with Schistosoma mansoni, a trematode parasite, altered the susceptibility of mice to colitis induced by dextran sodium sulfate (DSS). Mice infected with schistosome worms were refractory to DSS-induced colitis. Egg-laying schistosome infections or injection of eggs did not render mice resistant to colitis induced by DSS. Schistosome worm infections prevent colitis by a novel mechanism dependent on macrophages, and not by simple modulation of Th2 responses, or via induction of regulatory CD4+ or CD25+ cells, IL-10, or TGF-beta. Infected mice had marked infiltration of macrophages (F4/80+CD11b+CD11c(-)) into the colon lamina propria and protection from DSS-induced colitis was shown to be macrophage dependent. Resistance from colitis was not due to alternatively activated macrophages. Transfer of colon lamina propria F4/80+ macrophages isolated from worm-infected mice induced significant protection from colitis in recipient mice treated with DSS. Therefore, we propose a new mechanism whereby a parasitic worm suppresses DSS-induced colitis via a novel colon-infiltrating macrophage population.

Topics: Animals; Colitis; Dextran Sulfate; Immune Tolerance; Interleukin-10; Macrophages; Mice; Mice, Inbred Strains; Models, Animal; Mucous Membrane; Ovum; Schistosoma mansoni; Schistosomiasis mansoni; T-Lymphocytes, Regulatory; Th2 Cells; Transforming Growth Factor beta

The cytokine transforming growth factor-beta (TGF-beta) converts naïve T cells into regulatory T (Treg) cells that prevent autoimmunity. However, in the presence of interleukin-6 (IL-6), TGF-beta has also been found to promote the differentiation of naïve T lymphocytes into proinflammatory IL-17 cytokine-producing T helper 17 (T(H)17) cells, which promote autoimmunity and inflammation. This raises the question of how TGF-beta can generate such distinct outcomes. We identified the vitamin A metabolite retinoic acid as a key regulator of TGF-beta-dependent immune responses, capable of inhibiting the IL-6-driven induction of proinflammatory T(H)17 cells and promoting anti-inflammatory Treg cell differentiation. These findings indicate that a common metabolite can regulate the balance between pro- and anti-inflammatory immunity.

Topics: Animals; Cell Differentiation; Cells, Cultured; Colitis; Dendritic Cells; Dibenzazepines; Forkhead Transcription Factors; Interleukin-17; Interleukin-2; Interleukin-6; Intestinal Mucosa; Listeriosis; Lymphocyte Activation; Mice; Mice, Inbred C57BL; Mice, Transgenic; Spleen; T-Lymphocyte Subsets; T-Lymphocytes, Helper-Inducer; T-Lymphocytes, Regulatory; Transforming Growth Factor beta; Tretinoin

Previous studies have identified a counterinflammatory vagal reflex in the context of endotoxic shock. We have extended this observation to show that the vagus confers protection against acute (5 days) colitis induced by dextran sodium sulfate (DSS) or by dinitrobenzene sulfonic acid (DNBS). We have shown that this is mediated via macrophages and involves the suppression of proinflammatory cytokines. In this study, we have examined whether the vagal integrity confers long-lasting protection by studying DNBS- and DSS-induced inflammatory responses in the colon at 9 to 61 days postvagotomy. The integrity of vagotomy was confirmed at all time points using CCK-induced satiety. As previously described in a DNBS and DSS model, vagotomy associated with the pyloroplasty increased all indices of inflammation. Vagotomy increased the disease activity index as well as the macroscopic and histological scores by 75 and 41%, respectively. In addition, myeloperoxidase (MPO) activity, serum levels of C-reactive protein (CRP), and colonic tissue levels of proinflammatory cytokine increased when colitis was induced 9 days postvagotomy. However, these increases in inflammatory indices were substantially diminished in mice with colitis induced 21, 33, and 61 days postvagotomy. This was accompanied by an increased production of interleukin-10, transforming growth factor-beta, Forkhead Box P3 (FOXP3) staining in colonic tissue, and serum corticosterone. These findings indicate that although vagal integrity is an important protective factor, other counterinflammatory mechanisms come into play if vagal integrity is compromised beyond 2 wk.

Topics: Animals; C-Reactive Protein; Colitis; Colon; Corticosterone; Dextran Sulfate; Dinitrofluorobenzene; Disease Models, Animal; Eating; Forkhead Transcription Factors; Interleukins; Male; Mice; Mice, Inbred C57BL; Peroxidase; Reflex; Severity of Illness Index; Sincalide; T-Lymphocytes, Regulatory; Time Factors; Transforming Growth Factor beta; Vagotomy, Truncal; Vagus Nerve

Chronic inflammatory diseases may develop when regulatory T cells (Tregs) fail to control the balance between tolerance and immunity. Alternatively, activated immune cells might prevent the induction or activation of Tregs in such diseases. In this study, we demonstrate that trans-signaling into T cells via the soluble IL-6 receptor completely abrogates the de novo induction of adaptive Tregs. Mechanistically, IL-6 trans-signaling augmented the expression of the TGF-beta signaling inhibitor SMAD7. Consequently, SMAD7 overexpression in T cells using newly created transgenic mice rendered CD4(+)CD25(-) T cells resistant to the induction of FoxP3. Finally, IL-6 trans-signaling inhibited Treg-mediated suppression in a murine model of colitis. In summary, IL-6 trans-signaling into T cells emerges as a key pathway for blockade of the development of adaptive Tregs and thus may play a pivotal role in shifting the balance between effector and regulatory T cell numbers in chronic inflammatory and autoimmune diseases.

Topics: Animals; Autoimmune Diseases; Chronic Disease; Colitis; Disease Models, Animal; Forkhead Transcription Factors; Gene Expression Regulation; Humans; Inflammation; Interleukin-6; Mice; Mice, Inbred BALB C; Mice, SCID; Mice, Transgenic; Receptors, Interleukin-6; Signal Transduction; Smad7 Protein; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

The cytokine transforming growth factor-beta (TGF-beta) is an important negative regulator of adaptive immunity. TGF-beta is secreted by cells as an inactive precursor that must be activated to exert biological effects, but the mechanisms that regulate TGF-beta activation and function in the immune system are poorly understood. Here we show that conditional loss of the TGF-beta-activating integrin alpha(v)beta8 on leukocytes causes severe inflammatory bowel disease and age-related autoimmunity in mice. This autoimmune phenotype is largely due to lack of alpha(v)beta8 on dendritic cells, as mice lacking alpha(v)beta8 principally on dendritic cells develop identical immunological abnormalities as mice lacking alpha(v)beta8 on all leukocytes, whereas mice lacking alpha(v)beta8 on T cells alone are phenotypically normal. We further show that dendritic cells lacking alpha(v)beta8 fail to induce regulatory T cells (T(R) cells) in vitro, an effect that depends on TGF-beta activity. Furthermore, mice lacking alpha(v)beta8 on dendritic cells have reduced proportions of T(R) cells in colonic tissue. These results suggest that alpha(v)beta8-mediated TGF-beta activation by dendritic cells is essential for preventing immune dysfunction that results in inflammatory bowel disease and autoimmunity, effects that are due, at least in part, to the ability of alpha(v)beta8 on dendritic cells to induce and/or maintain tissue T(R) cells.

Topics: Aging; Animals; Autoimmunity; Colitis; Colon; Dendritic Cells; Immunoglobulins; Immunologic Memory; Integrins; Interferon-gamma; Interleukin-4; Leukocytes; Lymphocyte Activation; Lymphocyte Count; Mice; Phenotype; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Regulatory T (T(reg)) cells play a key role in the maintenance of the immune system homeostasis. T(reg) cells can be generated in the periphery under control of TGF-beta, a cytokine involved in the negative control of the immune system. However, TGF-beta cooperates with IL-6 in the generation of Th17 cells, a novel class of effector cells involved in numerous inflammatory diseases, including colitis. Therefore, TGF-beta emerges as a mediator of both anti-inflammatory and pro-inflammatory processes, depending on the local cytokine milieu. Here we demonstrate that IL-21, a type-1 cytokine produced by T cells and involved in the pathogenesis of immune-mediated diseases, prevents the TGF-beta-dependent expression of FoxP3, the master regulator of T(reg) cell commitment, and the induction of suppressive capacity in naive CD4(+) T cells, while promoting the differentiation of Th17 cells. In vivo, CD4(+) naive T cells activated in the presence of TGF-beta and IL-21 failed to suppress colitis while inducing an inflammatory response characterized by high levels of IL-17 and RORgammat, the transcription factor expressed by Th17 cells. Therefore, IL-21 emerges as a key modulator of TGF-beta signaling, leading to the reciprocal differentiation of T(reg) and Th17 cells.

Topics: Adoptive Transfer; Animals; Colitis; Disease Models, Animal; Flow Cytometry; Forkhead Transcription Factors; Gene Expression; Interleukin-17; Interleukins; Mice; Mice, Inbred BALB C; Mice, SCID; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; T-Lymphocyte Subsets; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

The imbalance between effector and regulatory T cells plays a central role in the pathogenesis of inflammatory bowel diseases. In addition to the thymus, CD4+CD25+ regulatory T cells can be induced in the periphery from a population of CD25- T cells by treatment with transforming growth factor beta (TGF-beta). Here, we analysed the in vivo function of TGF-beta induced regulatory T (Ti-Treg) cells in experimental colitis.. Ti-Treg cells were generated in cell culture in the presence or absence of TGF-beta and tested for their regulatory potential in experimental colitis using the CD4+CD62L+ T cell transfer model.. Ti-Treg cells significantly suppressed Th1 mediated colitis on CD4+CD62L+ T cell transfer in vivo, as shown by high resolution endoscopy, histology, immunohistochemistry, and cytokine analysis. Further analysis of in vivo and in vitro expanded Ti-Treg cells showed that exogenous interleukin 2 (IL-2) was crucial for survival and expansion of these cells.. Our data suggest that regulatory Ti-Treg cells expand by TGF-beta and exogenous IL-2 derived from effector T cells at the site of inflammation. In addition to Tr1 and thymic CD4+CD25+ T cells, peripheral Ti-Treg cells emerge as a class of regulatory T cells with therapeutic potential in T cell mediated chronic intestinal inflammation.

Topics: Adoptive Transfer; Animals; CD4-Positive T-Lymphocytes; Cell Proliferation; Colitis; Cytokines; Forkhead Transcription Factors; Immunohistochemistry; L-Selectin; Lymphocyte Activation; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, SCID; Reverse Transcriptase Polymerase Chain Reaction; T-Lymphocytes, Regulatory; Th1 Cells; Transforming Growth Factor beta

The phosphodiesterase-4 (PDE4) inhibitors may be an important target in the treatment of several inflammatory conditions. The anti-inflammatory effect of PDE4 inhibitors bears similarities with that of steroids, without interfering with the hypophysary-adrenal-axis. We compared the effect of rolipram, a selective PDE4 inhibitor, with steroids on the clinical course of experimental colitis induced by 2,4,6-trinitrobenzenesulfonic acid (TNBS). Three groups of rats (n = 20) received TNBS. One group received methylprednisolone from day 7, another group received rolipram from the same day, and control group received no further treatment. On days 14 and 21 after TNBS instillation, sets of 10 rats underwent colonic dialysis to measure eicosanoid release. Colonic lesions were blindly scored, and colons were homogenized for quantification of myeloperoxidase (MPO) activity and collagen content. Concentration of tumor necrosis factor alpha (TNF-alpha) and transforming growth factor beta1 (TGF-beta1) in colonic tissue was also measured. Both treatments reduced significantly the eicosanoid release and MPO activity. On day 14, both rolipram and methylprednisolone significantly reduced TNF-alpha content, but TGF-beta1 was only inhibited by rolipram. On day 21, lesion scores and collagen content were significantly reduced only in rolipram-treated group. In conclusion, PDE4 inhibition by rolipram markedly ameliorates the course of chronic colitis and it is superior to methylprednisolone in preventing late collagen deposition.

Topics: 3',5'-Cyclic-AMP Phosphodiesterases; Animals; Chronic Disease; Colitis; Colon; Cyclic Nucleotide Phosphodiesterases, Type 4; Disease Models, Animal; Fibrosis; Male; Methylprednisolone; Peroxidase; Phosphodiesterase Inhibitors; Rats; Rats, Sprague-Dawley; Rolipram; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tumor Necrosis Factor-alpha

Interleukin (IL)-13 is a major inducer of fibrosis in many chronic infectious and autoimmune diseases. In studies of the mechanisms underlying such induction, we found that IL-13 induces transforming growth factor (TGF)-beta(1) in macrophages through a two-stage process involving, first, the induction of a receptor formerly considered to function only as a decoy receptor, IL-13Ralpha(2). Such induction requires IL-13 (or IL-4) and tumor necrosis factor (TNF)-alpha. Second, it involves IL-13 signaling through IL-13Ralpha(2) to activate an AP-1 variant containing c-jun and Fra-2, which then activates the TGFB1 promoter. In vivo, we found that prevention of IL-13Ralpha(2) expression reduced production of TGF-beta(1) in oxazolone-induced colitis and that prevention of IL-13Ralpha(2) expression, Il13ra2 gene silencing or blockade of IL-13Ralpha(2) signaling led to marked downregulation of TGF-beta(1) production and collagen deposition in bleomycin-induced lung fibrosis. These data suggest that IL-13Ralpha(2) signaling during prolonged inflammation is an important therapeutic target for the prevention of TGF-beta(1)-mediated fibrosis.

Topics: Animals; Bleomycin; Blotting, Western; Cell Lineage; Colitis; Collagen; Cytokines; Down-Regulation; Enzyme-Linked Immunosorbent Assay; Etanercept; Fibrosis; Flow Cytometry; Gene Silencing; Genetic Vectors; Humans; Immunoglobulin G; Inflammation; Interleukin-13; Interleukin-13 Receptor alpha1 Subunit; Luciferases; Lung; Macrophages; Mice; Mice, Inbred C57BL; Monocytes; NF-kappa B; Oxazolone; Promoter Regions, Genetic; Receptors, Interleukin; Receptors, Interleukin-13; Receptors, Tumor Necrosis Factor; Reverse Transcriptase Polymerase Chain Reaction; RNA, Small Interfering; Signal Transduction; Time Factors; Transcription Factor AP-1; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tumor Necrosis Factor-alpha; Up-Regulation

Topics: Colitis; Humans; Immunosuppression Therapy; Interleukin-2; T-Lymphocytes, Regulatory; Thymus Gland; Transforming Growth Factor beta

While the impact of Bifidobacterium infantis 35624 and other probiotics on cytokines has been shown in established colitis, the effects of B. infantis consumption in pre-inflammation of interleukin (IL)-10 knock-out (KO) mice and on the wild-type (WT) C57Bl/6 mice have not been well demonstrated. The objective of this study was to examine cytokine responses in mucosal and systemic lymphoid compartments of IL-10 KO mice early in disease and to compare with control WT mice. Mice were fed B. infantis or placebo for 5 weeks and culled prior to the onset of chronic intestinal inflammation (12-14 weeks). The spleen, Peyer's patches and intestinal mucosa were removed and stimulated with various bacterial stimuli. Cytokine levels were measured by enzyme-linked immunosorbent assay. While basal intestinal and systemic cytokine profiles of WT and IL-10 KO mice were similar, transforming growth factor (TGF)-beta was reduced in the spleen of IL-10 KO mice. Following probiotic consumption, interferon (IFN)-gamma was reduced in the Peyer's patch of both WT and IL-10 KO mice. Alterations in IFN-gamma in the Peyer's patches of WT mice (enhancement) versus IL-10 KO (reduction) were observed following in vitro stimulation with salmonella. Differential IL-12p40, CCL2 and CCL5 responses were also observed in IL-10 KO mice and WT mice. The cytokine profile of IL-10 KO mice in early disease was similar to that of WT mice. The most pronounced changes occurred in the Peyer's patch of IL-10 KO mice, suggesting a probiotic mechanism of action independent of IL-10. This study provides a rationale for the use of B. infantis 35624 for the treatment of gastrointestinal inflammation.

Topics: Animals; Bifidobacterium; Colitis; Cytokines; Female; Interferon-gamma; Interleukin-10; Intestinal Mucosa; Mice; Mice, Inbred BALB C; Mice, Knockout; Peyer's Patches; Probiotics; Spleen; Transforming Growth Factor beta

Dynamic and reciprocal epithelial-mesenchymal interactions are critical for the normal morphogenesis and maintenance of epithelia. Epimorphin has been identified as a unique molecule expressed by mesenchymal cells and myofibroblasts and has putative morphogenetic effects in multiple epithelial tissues, including intestine, skin, mammary gland, lung, gallbladder, and liver. To define the in vivo role of epimorphin, we created epimorphin-null mice by targeted inactivation of the epimorphin gene. Male epimorphin-/- mice are sterile due to abnormal testicular development and impaired spermatogenesis. Intestinal growth is increased in epimorphin-/- mice due to augmented crypt cell proliferation and crypt fission during the neonatal (suckling) period, mediated at least in part by changes in bone morphogenetic protein (Bmp) and Wnt/beta-catenin signaling pathways. Colonic mucosal injury and colitis induced by dextran sodium sulfate (DSS) are ameliorated in epimorphin-/- mice, probably due to the increased proliferative capacity of the epimorphin-/- colon. These in vivo findings support the notion that epimorphin is a key stromal regulator of epithelial cell proliferation and growth in the intestine. In addition, our studies demonstrate a novel and critical role for epimorphin in regulating testicular development and growth as well as spermatogenesis.

Topics: Animals; beta Catenin; Bone Morphogenetic Proteins; Cell Line; Colitis; Dextran Sulfate; Female; Gene Targeting; Indicators and Reagents; Intestines; Male; Membrane Glycoproteins; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Nude; Morphogenesis; Spermatogenesis; Testis; Transforming Growth Factor beta; Wnt Proteins

Specific pathogen-free (SPF), but not germfree (GF), interleukin (IL)-2-deficient (IL-2-/-) mice develop inflammatory bowel disease (IBD) at 10 to 15 weeks of age. Gnotobiotic IL-2-/- mice monocolonized with E. coli mpk develop IBD at 25 to 33 weeks of age but not B. vulgatus mpk, E. coli Nissle 1917, or mice cocolonized with both E. coli mpk and B. vulgatus.. To determine genes regulated by these commensal bacteria, host gene expression in the colon of 8-week-old IL-2-/- mice was compared by using microarrays and semiquantitative reverse-transcription polymerase chain reaction. Colonization with E. coli mpk/B. vulgatus or SPF microbiota altered the gene expression profile more profoundly than monocolonization with either B. vulgatus, E. coli mpk or E. coli Nissle indicating that the complexity of the gene expression pattern is influenced by the diversity of the microbiota.. A small but distinct group of genes could be defined which might be associated with colitis development. Thus, 8 week old E. coli mpk IL-2-/- mice rone to colitis compared to E. coli Nissle, B. vulgatus and E. coli mpk/B. vulgatus IL-2-/- mice displayed a lower expression of the anti-inflammatory RegIII family genes such as RegIII[gamma] and pancreatitis associated protein (PAP) and peroxisome proliferator-activated receptor-[gamma] regulated genes such as adipsin and adiponectin.. The increased expression of these genes in B. vulgatus colonized mice might be associated with prevention of E. coli mpk triggered colitis in E. coli mpkM/B. vulgatus IL-2-/- mice.

Topics: Adipocytes; Animals; Bacteroides; Colitis; Colon; Escherichia coli; Gene Expression; Gene Expression Profiling; Gene Expression Regulation; Genes, Immunoglobulin; Genetic Predisposition to Disease; Germ-Free Life; Interleukin-2; Mice; Mice, Inbred C57BL; Pancreatitis-Associated Proteins; Transforming Growth Factor beta

Interleukin (IL)-6 is a proinflammatory cytokine implicated in the pathogenesis of inflammatory bowel disease. IL-6 is locally upregulated in inflammatory bowel disease patients and in murine models of experimental colitis. Treatment with anti-IL-6 receptor antibody ameliorates intestinal inflammation.. It was the aim of this study to investigate the role of genetic IL-6 deficiency in trinitrobenzene sulfonic acid (TNBS)-mediated colitis, an experimental model inflammation that shares several features with Crohn's disease in humans.. Acute colitis was induced in wild-type and IL-6-deficient (Il-6(-/-)) mice by intracolonic administration of TNBS. Forty-eight hours after treatment, the local and systemic features of inflammation, i.e. food intake, weight loss, histological markers of colitis, cytokine expression by real-time PCR, food intake and body weight changes, were assessed.. In wild-type mice, TNBS administration increased both IL-6 serum levels and local expression of IL-6 by 36 and 9 fold, respectively, compared with control, vehicle-injected mice. Compared with the wild-type mice, the Il-6(-/-) mice had significantly reduced intestinal inflammation as evidenced by epithelial damage, neutrophil infiltration, colon thickness and proinflammatory cytokine expression, following treatment with TNBS. Moreover, baseline levels of the antiinflammatory cytokines IL-10 and tumor growth factor-beta were significantly elevated in Il-6(-/-)compared with the wild-type mice.. Our results demonstrate that Il-6(-/-)are partially protected from the development of TNBS-induced acute experimental colitis, most likely via their significantly elevated baseline levels of antiinflammatory cytokines.

Topics: Animals; Body Weight; Chemotaxis, Leukocyte; Colitis; Colon; Disease Models, Animal; Eating; Female; Interleukin-10; Interleukin-6; Mice; Mice, Inbred C57BL; Mice, Knockout; Transforming Growth Factor beta; Trinitrobenzenesulfonic Acid; Up-Regulation

Crohn's disease is a chronic debilitating disease characterized by severe T helper cell (Th)1-driven inflammation of the colon partially caused by a loss of immune tolerance against mucosal antigens. The use of regulatory dendritic cells (DCs) with the capacity to induce regulatory T cells has been proposed recently for the treatment of Crohn's disease in a strategy to restore immune tolerance. Vasoactive intestinal peptide is an immunomodulatory neuropeptide that induces regulatory DCs. The aim of this study was to investigate the therapeutic effect of vasoactive intestinal peptide-induced regulatory DCs (DC(VIP)) in a murine model of colitis.. We examined the therapeutic action of DC(VIP) in the colitis induced by intracolonic administration of trinitrobenzene sulfonic acid, evaluating diverse clinical signs of the disease including weight loss, diarrhea, colitis, and histopathology. We also investigated the mechanisms involved in the potential therapeutic effect of DC(VIP), such as inflammatory cytokines and chemokines, Th1-type response, and the generation of regulatory T cells.. DC(VIP) injection significantly ameliorated the clinical and histopathologic severity of colitis, abrogating body weight loss, diarrhea, and inflammation, and increasing survival. The therapeutic effect was associated with down-regulation of both inflammatory and Th1-driven autoimmune response, by regulating a wide spectrum of inflammatory mediators directly through activated macrophages, and by generating interleukin-10-secreting regulatory T cells with suppressive capacity on autoreactive T cells.. The possibility to generate/expand ex vivo regulatory DC(VIP) opens new therapeutic perspectives for the treatment of Crohn's disease in human beings, and may minimize the dependence on nonspecific immunosuppressive drugs used currently for autoimmune disorders.

Topics: Animals; Autoimmunity; CD4-Positive T-Lymphocytes; Cell Transplantation; Cells, Cultured; Colitis; Dendritic Cells; Disease Models, Animal; Immune Tolerance; Immunotherapy; Interleukin-10; Mice; Mice, Inbred BALB C; Th1 Cells; Transforming Growth Factor beta; Trinitrobenzenesulfonic Acid; Vasoactive Intestinal Peptide

The ectopeptidases Dipeptidylpeptidase IV and Alanyl-Aminopeptidase N, strongly expressed by both, activated and regulatory T cells were shown to co-operate in T cell regulation. Based on the findings that DPIV and APN inhibitors induce the TGF-beta1 and IL-10 production and a suppression of T helper cell proliferation in parallel, and that particularly APN inhibitors amplify the suppressing activity of regulatory T cells, both peptidases represent a promising target complex for treatment of diseases associated with an imbalanced T cell response, such as inflammatory bowel diseases (IBD). The aim of the present study was to analyze the therapeutic potential of DPIV and APN inhibitors in vivo in a mouse model of colitis. Balb/c mice received 3% (w/v) dextran sulphate sodium with the drinking water for 7 days. After onset of colitis symptoms, inhibitor treatment started at day 3. Disease activity index (DAI) was assessed daily, supplemented by histological and immunological analysis. While the DPIV inhibitor Lys-[Z(NO])(2)]-pyrrolidide or the APN-inhibitor Actinonin alone had marked but no significant therapeutic effects, the simultaneous administration of both inhibitors reduced colitis activity in comparison to placebo treated mice, significantly (DAI 4.8 vs. 7.7, p<0.005). A newly developed compound IP12.C6 with inhibitory capacity toward both enzymes significantly attenuated the clinical manifestation of colitis (DAI 3.2 vs. 7.6, p<0.0001). TGF-beta mRNA was found to be up-regulated in colon tissue of inhibitor-treated animals. In summary our results strongly suggest that combined DPIV and APN inhibition by synthetic inhibitors represents a novel and efficient approach for the pharmacological therapy of IBD by triggering endogenous immunosuppressive mechanisms.

Topics: Animals; Body Weight; CD13 Antigens; Colitis; Colon; Cytokines; Dextran Sulfate; Dipeptidyl-Peptidase IV Inhibitors; Drug Therapy, Combination; Female; Forkhead Transcription Factors; Gene Expression; Hydroxamic Acids; Immunosuppressive Agents; Inflammatory Bowel Diseases; Lysine; Mice; Mice, Inbred BALB C; Protease Inhibitors; Pyrrolidines; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Transforming Growth Factor beta

The aim of this study was to ascertain factors that might be protective of the appearance of gross blood in the stools of breast-fed infants.. Logistic regression models were formed to search for variables possibly explaining the condition. In addition to the analyzed breast milk factors, mother's allergic disease was introduced into the models to control for its possible confounding effect. The breast milk samples, collected from mothers of infants with gross blood in stools (n = 23) and from mothers of healthy age-matched infants (n = 71), were analyzed for concentrations of transforming growth factor-beta2, tumor necrosis factor-alpha, interleukin (IL)-4, IL-10, prostaglandin (PG)E2, cysteinyl leukotrienes (Cys-LTs) and fatty acid composition.. Increase in the concentrations of PGE2 and Cys-LTs in the breast milk together with mother's allergic disease reduced the likelihood of gross blood in stools in the breast-fed infant. The results suggest that no single factor, but a combination of immunomodulatory factors may protect the child from gross blood in the stools of breast-fed infants. Allergic disease was not a risk factor as mother's allergic disease appeared to counterbalance the gross blood in stools. Due to the preliminary nature of the study, the results need to be verified in a larger setting. The challenge for the future lies in identifying of such active compounds for dietary modification to enforce particularly the properties of the breast milk which are immunoprotective for the infant and to reduce the likelihood of intestinal disorders in at risk infants.

Topics: Blood; Breast Feeding; Colitis; Cysteine; Dinoprostone; Fatty Acids; Feces; Female; Humans; Hypersensitivity; Immunoglobulin E; Infant; Infant, Newborn; Leukotrienes; Lipids; Logistic Models; Male; Milk, Human; Transforming Growth Factor beta; Transforming Growth Factor beta2; Tumor Necrosis Factor-alpha

In previous studies, we have shown that the oral administration of colonic proteins that have been haptenated (i.e., haptenated colonic proteins [HCPs]) with trinitrophenol (TNP) can protect mice from the subsequent induction of trinitrobenzene sulfonic acid colitis. Inasmuch as this protection was mediated by regulatory cells that express the antigen-non-specific suppressor factors transforming growth factor-beta and interleukin-10, we reasoned that TNP-HCP feeding would also "cross-protect" mice from colitis induced by a different hapten, oxazolone. Indeed, we found that feeding TNP-HCP protected mice from the development of oxazolone-colitis, albeit to a lesser extent than it protected mice from trinitrobenzene sulfonic acid colitis. In addition, we showed that protection was associated with the appearance of mononuclear cells producing regulatory cytokines. These data strongly imply that the cells induced by feeding 1 type of haptenated protein are capable of cross-reacting with antigens present in colitis produced by a second type of haptenated protein. The cross-protection demonstrated in this study holds promise for the treatment of humans with inflammatory bowel disease because it shows that an appropriate fed antigen can induce regulatory cells that have the potential to suppress an inflammation induced by the unknown antigens causing this disease.

Topics: Adjuvants, Immunologic; Administration, Oral; Animal Feed; Animals; Antigens; Colitis; Colon; Disease Models, Animal; Haptens; Interleukin-10; Male; Mice; Monocytes; Oxazolone; Picrates; Proteins; Transforming Growth Factor beta; Trinitrobenzenesulfonic Acid

Nonpathogenic enteric bacterial species initiate and perpetuate experimental colitis in IL-10 gene-deficient mice (IL-10(-/-)). Bacteria-specific effects on the epithelium are difficult to dissect due to the complex nature of the gut microflora. We showed that IL-10(-/-) mice compared with wild-type mice fail to inhibit proinflammatory gene expression in native intestinal epithelial cells (IEC) after the colonization with colitogenic Gram-positive Enterococcus faecalis. Interestingly, proinflammatory gene expression was transient after 1 wk of E. faecalis monoassociation in IEC from wild-type mice, but persisted after 14 wk of bacterial colonization in IL-10(-/-) mice. Accordingly, wild-type IEC expressed phosphorylated NF-kappaB subunit RelA (p65) and phosphorylated Smad2 only at day 7 after bacterial colonization, whereas E. faecalis-monoassociated IL-10(-/-) mice triggered persistent RelA, but no Smad2 phosphorylation in IEC at days 3, 7, 14, and 28. Consistent with the induction of TLR2-mediated RelA phosphorylation and proinflammatory gene expression in E. faecalis-stimulated cell lines, TLR2 protein expression was absent after day 7 from E. faecalis-monoassociated wild-type mice, but persisted in IL-10(-/-) IEC. Of note, TGF-beta1-activated Smad signaling was associated with the loss of TLR2 protein expression and the inhibition of NF-kappaB-dependent gene expression in IEC lines. In conclusion, E. faecalis-monoassociated IL-10(-/-), but not wild-type mice lack protective TGF-beta/Smad signaling and fail to inhibit TLR2-mediated proinflammatory gene expression in the intestinal epithelium, suggesting a critical role for IL-10 and TGF-beta in maintaining normal epithelial cell homeostasis in the interplay with commensal enteric bacteria.

Topics: Animals; Cell Line; Cells, Cultured; Colitis; DNA-Binding Proteins; Enterococcus faecalis; Gene Expression Regulation; Gram-Positive Bacterial Infections; Inflammation Mediators; Interleukin-10; Intestinal Mucosa; Mice; Mice, Inbred C3H; Mice, Knockout; NF-kappa B; Phosphorylation; Receptors, Cell Surface; Signal Transduction; Smad Proteins; Toll-Like Receptor 2; Trans-Activators; Transcription Factor RelA; Transforming Growth Factor beta; Transforming Growth Factor beta1

Recent studies of murine models of mucosal inflammation suggest that, whereas some kinds of bacterial microflora are inducers of disease, others, known as probiotics, prevent disease. In the present study, we analyzed the regulatory cytokine and cell response to probiotic (VSL#3) administration in the context of the Th1 T cell colitis induced by trinitrobenzene sulfonic acid treatment of SJL/J mice. Daily administration of probiotics for 3 wk to mice during a remission period between a first and second course of colitis induced by trinitrobenzene sulfonic acid, resulted in a milder form of recurrent colitis than observed in mice administered PBS during this same period. This protective effect was attributable to effects on the lamina propria mononuclear cell (LPMC) population, because it could be transferred by LPMC from probiotic-treated mice to naive mice. Probiotic administration was associated with an early increase in the production of IL-10 and an increased number of regulatory CD4+ T cells bearing surface TGF-beta in the form of latency-associated protein (LAP) (LAP+ T cells). The latter were dependent on the IL-10 production because administration of anti-IL-10R mAb blocked their appearance. Finally, the LAP+ T cells were essential to the protective effect of probiotics because administration of anti-IL-10R or anti-TGF-beta at the initiation of recurrent colitis induction or depletion of LAP+ T cells from LPMC abolished the latter's capacity to transfer protection to naive recipients. These studies show that probiotic (VSL#3) administration during a remission period ameliorates the severity of recurrent colitis by inducing an immunoregulatory response involving TGF-beta-bearing regulatory cells.

Topics: Adoptive Transfer; Animals; CD4-Positive T-Lymphocytes; Colitis; Interleukin-10; Male; Mice; Peptide Fragments; Probiotics; Protein Precursors; Recurrence; T-Lymphocyte Subsets; Th1 Cells; Transforming Growth Factor beta; Transforming Growth Factor beta1; Trinitrobenzenesulfonic Acid

CD4(+)CD25(+) regulatory T (T reg) cells play a pivotal role in control of the immune response. Transforming growth factor-beta (TGF-beta) has been shown to be required for T reg cell activity; however, precisely how it is involved in the mechanism of suppression is poorly understood. Using the T cell transfer model of colitis, we show here that CD4(+)CD45RB(high) T cells that express a dominant negative TGF-beta receptor type II (dnTbetaRII) and therefore cannot respond to TGF-beta, escape control by T reg cells in vivo. CD4(+)CD25(+) T reg cells from the thymus of dnTbetaRII mice retain the ability to inhibit colitis, suggesting that T cell responsiveness to TGF-beta is not required for the development or peripheral function of thymic-derived T reg cells. In contrast, T reg cell activity among the peripheral dnTbetaRII CD4(+)CD25(+) population is masked by the presence of colitogenic effector cells that cannot be suppressed. Finally, we show that CD4(+)CD25(+) T reg cells develop normally in the absence of TGF-beta1 and retain the ability to suppress colitis in vivo. Importantly, the function of TGF-beta1(-/-) T reg cells was abrogated by anti-TGF-beta monoclonal antibody, indicating that functional TGF-beta can be provided by a non-T reg cell source.

Topics: Animals; CD4-Positive T-Lymphocytes; Cell Differentiation; Colitis; Cytokines; Immune Tolerance; Mice; Mice, Knockout; Receptors, Interleukin-2; Spleen; Th1 Cells; Thymus Gland; Transforming Growth Factor beta; Transforming Growth Factor beta1

n-3 polyunsaturated fatty acids (PUFAs) reduce the severity of chronic inflammatory bowel disease, probably by means of reduction of immune cell activation or enhancement of the epithelial barrier. Using the severe combined immunodeficient (SCID) mouse model of colitis, this study examined the effect of dietary n-3 PUFAs on development of colitis and on immunologic, epithelial, and matrix parameters in the intestines of control and colitic animals.. SCID mice were fed n-3-enriched or control diet for 3 weeks before colitis induction by transplantation of CD45RB T cells and maintained on the same diet for 4 to 8 weeks. Phenotype of infiltrating cells, epithelial ZO-1 protein, and mucosal type I collagen were assessed by immunohistology and tissue cytokines by ELISA.. Transplanted n-3-fed animals had significantly reduced pathology scores, colonic tumor necrosis factor-alpha, interleukin-12, and interleukin-1beta compared with animals fed standard diet. Proinflammatory cytokines were reduced despite a similar level of immune cell infiltration by T cells, CD11c cells, and CD11b cells. Neutrophil infiltration was significantly reduced in n-3-fed control and colitic mice, and other myeloid populations were reduced in mice on the n-3 diet. Epithelial ZO-1 expression was increased, and myofibroblast activation significantly decreased in transplanted n-3-fed animals compared with standard diet mice. Submucosal collagen synthesis was enhanced in n-3-fed mice.. Dietary n-3 PUFAs reduced clinical colitis and colonic immunopathology in this model of colonic inflammation by decreasing proinflammatory cytokine synthesis, reducing myeloid cell recruitment and activation, and enhancing epithelial barrier function and mucosal wound healing mechanisms.

Topics: Animals; Chronic Disease; Colitis; Colon; Diet; Disease Models, Animal; Fatty Acids, Omega-3; Female; Interleukins; Mice; Mice, SCID; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

Taking advantage of the proximity of bowel mucosa to luminal bacteria, we have attempted to deliver a therapeutic gene to the colonic mucosa by oral administration of an invasive and non-pathogenic Escherichia coli. E. coli diamenopimelate (dap) auxotroph, harboring plasmid pGB2Omegainv-hly, express the inv gene from Yersinia pseudotubercolosis that confers the ability to invade nonprofessional phagocytic cells and the hly gene from Listeria monocytogenes that allows expression of lystreriolysin O, a perforin cytolysin able to perfore phagosomal membranes. This bacterial vector invades and transfers functional DNA to epithelial cells in vitro. We have shown that this strain carrying a therapeutic gene (pC1OmegaTGF-beta1) can significantly reduce the severity of experimental colitis in mice. However, as a consequence of mucosal barrier disruption during colitis, vector-specific mRNA transcripts could be recovered from the colon and also from extra-colonic tissues. We therefore replaced the constitutive CMV promoter in pC1OmegaTGF-beta1 by the inflammation-inducible interleukin-8 promoter generating plasmid pC1OmegaTGF-beta1IND. Plasmid-specific TGF-beta1 mRNA transcripts were detectable in mouse CMT-93 epithelial cells incubated with E. coli BM2710/pGB2Omegainv-hly carrying pC1OmegaTGF-beta1IND following exposure to inflammatory cytokines. Furthermore, the transcripts were detectable only within inflamed tissues and the therapeutic effects were comparable to those in animals treated with E. coli BM2710/pGB2Omegainv-hly+pC1OmegaTGF-beta1. In summary, engineered enteric bacteria can efficiently deliver in vivo therapeutic genes to the intact intestinal mucosa and regulation expression of the therapeutic gene by an inflammation-inducible promoter prevents its dissemination during colitis.

Topics: Administration, Oral; Animals; Bacterial Toxins; Colitis; Colon; Epithelial Cells; Escherichia coli Proteins; Gene Expression Regulation; Genetic Engineering; Genetic Therapy; Genetic Vectors; Heat-Shock Proteins; Hemolysin Proteins; Intestinal Absorption; Intestinal Mucosa; Listeria monocytogenes; Mice; Promoter Regions, Genetic; Transforming Growth Factor beta; Transforming Growth Factor beta1; Yersinia pseudotuberculosis

Null mutation of heterotrimeric G protein alpha2 inhibitory subunit (Galphai2) induces Th1-skewed hyperimmune responses in the colon, leading to chronic colitis and the development of colonic adenocarcinoma. However, the underlying molecular mechanisms and cellular basis, in particular, for the role of Galphai2 in regulating immune responses, are poorly understood. We show here that peripheral T cells from Galphai2-deficient mice do not respond normally to the inhibitory effects of TGF-beta on proliferation and cytokine production, revealing a previously unappreciated cross-talk between these two signaling pathways. Lack of Galphai2 resulted in decreased phosphorylation of Smad2 and Smad3 in T cells at the basal levels as well as at the late but not early phase of TGF-beta stimulation, which appears to be ascribed to differential expression of neither cell surface TGF-beta receptors nor Smad7. The altered phosphorylation of Smad proteins involves phospholipase C-mediated signaling, a downstream signaling molecule of Galphai2, because phospholipase C inhibitors could restore Smad2 and Smad3 phosphorylation in Galphai2(-/-) T cells at levels comparable to that in wild-type T cells. Moreover, adoptive transfer of Galphai2-deficient T cells into immunocompromised mice rendered an otherwise resistant mouse strain susceptible to trinitrobenzesulfonic acid-induced colitis, suggesting that an impaired response of Galphai2-deficient T cells to TGF-beta may be one of the primary defects accounting for the observed colonic Th1-skewed hyperimmune responses. These findings shed new lights on the molecular and cellular basis of how Galphai2 down-regulates immune responses, contributing to the maintenance of mucosal tolerance.

Topics: Adoptive Transfer; Animals; Colitis; DNA-Binding Proteins; Female; Genetic Predisposition to Disease; GTP-Binding Protein alpha Subunit, Gi2; GTP-Binding Protein alpha Subunits, Gi-Go; Immune Tolerance; Intestinal Mucosa; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Phosphorylation; Proto-Oncogene Proteins; Smad2 Protein; Smad3 Protein; T-Lymphocyte Subsets; Trans-Activators; Transforming Growth Factor beta; Trinitrobenzenesulfonic Acid

Enteropathogenic Escherichia coli (EPEC) is a common pathogen in infantile diarrhea, causing a characteristic histopathologic attaching and effacing (A/E) lesion in the intestinal mucosa. The mouse pathogen Citrobacter rodentium causes a similar A/E lesion in the murine intestine. Like EPEC, C. rodentium infection results in colonic crypt hyperplasia, goblet cell depletion, epithelial proliferation, and mucosal disruption. Using this murine model, we tested the hypothesis that preinoculation of murine gut with Lactobacillus acidophilus early in life can enhance host defense against enteric bacterial infection and attenuate bacteria-mediated colitis. Two-week old BALB/c mice were inoculated with L. acidophilus twice per week for 4 weeks before C. rodentium infection or concomitantly with the exposure to C. rodentium at 6-8 weeks of age. The probiotics were administered twice weekly thereafter. We observed that L. acidophilus inoculation in mice inhibits C. rodentium-induced colitis, which is associated with a decrease in C. rodentium colonization and translocation, an increase in its clearance, and a suppression of colonic myeloperoxidase (MPO) activity. Probiotic treatment also stimulates regulatory cytokine expression in the colon [transforming growth factor beta (TGF-beta), interleukin (IL)-10]. Preinoculation with L. acidophilus is more effective than concomitant use of probiotics in the induction of intestinal IgA secretion and in the downregulation of proinflammatory cytokine expression [tumor necrosis factor alpha (TNF-alpha), IL-6, and IL-12]. These observations suggest that inoculation with probiotics can effectively prevent bacteria-induced colitis by limiting enteric bacteria infection and promoting mucosal protective regulatory immune responses. This study may have ramifications for prevention of infectious diarrhea in human infants and children, particularly in developing countries.

Topics: Animals; Cell Proliferation; Citrobacter rodentium; Colitis; Colon; Enterobacteriaceae Infections; Interleukin-10; Intestinal Mucosa; Lactobacillus acidophilus; Mice; Mice, Inbred BALB C; Probiotics; Transforming Growth Factor beta

In previous studies, we have shown that murine CD4+CD25+ regulatory T cells produce high levels of TGF-beta1 in a cell surface and/or secreted form, and blockade of such TGF-beta1 by anti-TGF-beta curtails the ability of these cells to suppress CD25- T cell proliferation and B cell Ig production in in vitro suppressor assays. In further support for the role of TGF-beta1 in suppression by CD4+CD25+ T cells, we show in this study that another TGF-beta1-blocking molecule, recombinant latency-associated peptide of TGF-beta1 (rLAP), also reverses suppression by mouse CD4+CD25+ T cells as well as their human counterparts, CD4+CD25(high) T cells. In addition, we show that CD25- T cells exposed to CD4+CD25+ T cells in vitro manifest activation of Smad-2 and induction of CD103, the latter a TGF-beta-inducible surface integrin. In further studies, we show that while CD4+CD25+ T cells from TGF-beta1-deficient mice can suppress CD25- T cell proliferation in vitro, these cells do not protect recipient mice from colitis in the SCID transfer model in vivo, and, in addition, CD4+LAP+, but not CD4+LAP- T cells from normal mice protect recipient mice from colitis in this model. Together, these studies demonstrate that TGF-beta1 produced by CD4+CD25+ T cells is involved in the suppressor activity of these cells, particularly in their ability to regulate intestinal inflammation.

Topics: Adoptive Transfer; Animals; CD4-Positive T-Lymphocytes; Cell Membrane; Cells, Cultured; Colitis; Down-Regulation; Female; Humans; Inflammation Mediators; Intestinal Mucosa; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Mice, SCID; Peptide Fragments; Protein Binding; Protein Precursors; Receptors, Interleukin-2; Receptors, Transforming Growth Factor beta; Recombinant Proteins; Signal Transduction; T-Lymphocyte Subsets; T-Lymphocytes, Regulatory; Transforming Growth Factor beta; Transforming Growth Factor beta1

To determine the role of Smad3 in re-epithelialization and inflammation, experimental colitis was induced in Smad3 heterozygous mice and their wild-type littermates by single intrarectal administration of 2,4,6-trinitrobenzene sulfonic acid (TNBS) in ethanol. The area of epithelial deficiency was significantly reduced in the heterozygotes on the 4th-6th day after TNBS administration as compared to the controls although the number of inflammatory cells in the colonic mucosa in the heterozygotes and their wild-type littermates varied similarly throughout the course of colitis. Proliferation of the intestinal epithelium in the heterozygotes was significantly accelerated as compared to that in the wild-type controls on the 1st and 2nd days after TNBS administration. These results suggest that reduction of Smad3 significantly accelerates re-epithelialization of the intestinal mucosa without enhancing inflammation. Suppression of TGF-beta1 induction in the colonic mucosa of the heterozygotes may lead to a higher level of proliferation of intestinal epithelial cells.

Topics: Animals; Cell Division; Colitis; Disease Models, Animal; DNA-Binding Proteins; Inflammatory Bowel Diseases; Intestinal Mucosa; Male; Mice; Mice, Inbred C57BL; Regeneration; Severity of Illness Index; Smad3 Protein; Trans-Activators; Transforming Growth Factor beta; Trinitrobenzenesulfonic Acid; Wound Healing

There is a body of evidence to suggest that the local activation of angiotensin II (ANG II) plays a pivotal role in fibrogenic response involving the kidney, heart, lung, pancreas and liver. In such conditions, fibrosis is mediated, at least partially, through ANG II induction of the cytokine transforming growth factor-beta1 (TGF-beta1). Both ANG II and TGF-beta1 also seem to be involved in intestinal fibrosis and stenosis, particularly in Crohn's disease. The aim of the present study was, firstly, to determine the effects of the angiotensin-converting enzyme inhibitor, captopril, on colonic fibrosis in experimental colitis in rats and, secondly, to check the role of TGF-beta1 on these effects.. Colitis was induced in rats by intracolonic administration of TNBS. Colonic fibrosis was assessed 21 days later by macroscopic and microscopic evaluation. Levels of collagen alpha1 gene expression, hydroxyproline, angiotensin II and TGF-beta1 proteins, and TGF-beta1 mRNA were measured on the colonic tissue.. In chronic colitis, captopril significantly reduced the score of macroscopic and histologic lesions, as well as the colonic tissue levels of collagen alpha1, hydroxyproline, ANG II and TGF-beta1 proteins, and TGF-beta1 mRNA.. These data demonstrate for the first time that the prophylactic administration of captopril is effective in preventing colonic fibrosis in TNBS-induced colitis. The antifibrotic action of captopril could be due to the blockade of TGFbeta-1 overexpression, and/or to a direct down-regulation of TGFbeta-1 transcript.

Topics: Angiotensin-Converting Enzyme Inhibitors; Animals; Captopril; Colitis; Colon; Disease Models, Animal; Fibrosis; Male; Rats; Transforming Growth Factor beta; Transforming Growth Factor beta1

Data regarding the role of TGF-beta for the in vivo function of regulatory CD4(+)CD25(+) T cells (Treg) are controversial. A transgenic mouse model with impaired TGF-beta signaling specifically in T cells was used to assess the role of endogenous TGF-beta for the in vivo function of CD4(+)CD25(+) Treg in a murine model of colitis induced by dextran sulfate. Transfer of wild-type, but not transgenic CD4(+)CD25(+) Treg was found to suppress colitis in wild-type mice. In addition, by transferring CFSE-labeled CD4(+)CD25(+) Treg we could demonstrate that endogenous TGF-beta promotes the expansion of CD4(+)CD25(+) Treg in vivo. Transgenic mice themselves developed reduced numbers of peripheral CD4(+)CD25(+) Treg and were more susceptible to the induction of colitis, which could be prevented by the transfer of wild-type Treg. These data indicate that TGF-beta signaling in CD4(+)CD25(+) Treg is required for their in vivo expansion and suppressive capacity.

Topics: Adoptive Transfer; Animals; Cell Differentiation; Colitis; Genetic Predisposition to Disease; Humans; Lymphocyte Count; Mice; Receptors, Interleukin-2; Signal Transduction; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Transforming growth factor (TGF)-beta has a key role in intestinal homeostasis. Our present data suggest that TGF-beta, which was constitutively expressed by lamina propria mononuclear cells and epithelium, affected epithelial cells. Abnormal suppression of TGF-beta could enhance the sensitivity of epithelial cells to apoptosis associated with interferon-gamma in DSS-induced colitis.

Topics: Animals; Colitis; Dextran Sulfate; Disease Models, Animal; Immunity, Mucosal; Intestinal Mucosa; Mice; Transforming Growth Factor beta

In vitro studies suggest that transforming growth factor (TGF)-beta has potent effects on gastrointestinal mucosal integrity, wound repair, and neoplasia. However, the multiplicity of actions of this peptide on many different cell types confounds efforts to define the role of TGF-beta within the intestinal epithelium in vivo. To delineate these effects selective blockade of intestinal epithelial TGF-beta activity was undertaken through targeted expression of a dominant-negative (DN) TGF-beta RII to intestinal epithelial cells in vitro and in vivo. Stable intestinal epithelial cell (IEC)-6 lines overexpressing TGF-beta RII-DN (nucleotides -7 to 573) were established. Transgenic mice overexpressing TGF-beta RII-DN under the regulation of a modified liver fatty acid-binding promoter (LFABP-PTS4) were constructed. In vitro healing was assessed by wounding of confluent monolayers. Colitis was induced by the addition of dextran sodium sulfate (2.5 to 7.5% w/v) to their drinking water. Overexpression of TGF-beta RII-DN in intestinal epithelial cell-6 cells resulted in a marked reduction in cell migration and TGF-beta-stimulated wound healing in vitro. TGF-beta RII-DN transgenic mice did not exhibit baseline intestinal inflammation or changes in survival, body weight, epithelial cell proliferation, aberrant crypt foci, or tumor formation. TGF-beta RII-DN mice were markedly more susceptible to dextran sodium sulfate-induced colitis and exhibited impaired recovery after colonic injury. TGF-beta is required for intestinal mucosal healing and TGF-beta modulation of the intestinal epithelium plays a central role in determining susceptibility to injury.

Topics: Animals; Cell Division; Cell Line; Cell Movement; Colitis; Colon; Granulocytes; Humans; Intestinal Mucosa; Mice; Mice, Inbred DBA; Mice, Transgenic; Peroxidase; Promoter Regions, Genetic; Recombinant Proteins; Transforming Growth Factor beta; Wound Healing

Murine CD4(+)CD25(+) regulatory cells have been reported to express latency-associated peptide (LAP) and TGF-beta on the surface after activation, and exert regulatory function by the membrane-bound TGF-beta in vitro. We have now found that a small population of CD4(+) T cells, both CD25(+) and CD25(-), can be stained with a goat anti-LAP polyclonal Ab without being stimulated. Virtually all these LAP(+) cells are also positive for thrombospondin, which has the ability to convert latent TGF-beta to the active form. In the CD4(+)CD45RB(high)-induced colitis model of SCID mice, regulatory activity was exhibited not only by CD25(+)LAP(+) and CD25(+)LAP(-) cells, but also by CD25(-)LAP(+) cells. CD4(+)CD25(-)LAP(+) T cells were part of the CD45RB(low) cell fraction. CD4(+)CD25(-)LAP(-)CD45RB(low) cells had minimal, if any, regulatory activity in the colitis model. The regulatory function of CD25(-)LAP(+) cells was abrogated in vivo by anti-TGF-beta mAb. These results identify a new TGF-beta-dependent regulatory CD4(+) T cell phenotype that is CD25(-) and LAP(+).

Topics: Animals; Antibodies; Biotinylation; CD4-Positive T-Lymphocytes; Cell Membrane; Cells, Cultured; Coculture Techniques; Colitis; Cytokines; Disease Models, Animal; Female; Flow Cytometry; Goats; Immunophenotyping; Leukocyte Common Antigens; Male; Mice; Mice, Inbred BALB C; Mice, SCID; Peptide Fragments; Protein Precursors; Receptors, Interleukin-2; Staining and Labeling; T-Lymphocyte Subsets; Transforming Growth Factor beta; Transforming Growth Factor beta1

Regulatory T cells are critical in regulating the immune response, and therefore play an important role in the defense against infection and control of autoimmune diseases. However, a therapeutic role of regulatory T cells in an established disease has not been fully established. In this study, we provide direct evidence that CD4(+)CD25(+) regulatory T cells can cure an established, severe, and progressive colitis. SCID mice developed severe colitis when adoptively transferred with naive CD4(+)CD25(-) T cells and infected with the protozoan parasite Leishmania major. The disease development can be completely halted and symptoms reversed, with a healthy outcome, by transferring freshly isolated or activated CD4(+)CD25(+) T cells from syngeneic donors. The therapeutic effect of the regulatory T cells was completely blocked by treatment of the recipients with anti-IL-10R, anti-CTLA4, or anti-TGF-beta Ab. However, the resurgence of colitis under these treatments was not accompanied by the reactivation of Th1 or Th2 response nor was it correlated to the parasite load. These results therefore demonstrate that CD4(+)CD25(+) T cells are therapeutic and that the effect is mediated by both IL-10/TGF-beta-dependent and independent mechanisms. Furthermore, colitis can manifest independent of Th1 and Th2 responses.

Topics: Adoptive Transfer; Animals; Antigens, CD; Antigens, Differentiation; CD4-Positive T-Lymphocytes; Cells, Cultured; Colitis; CTLA-4 Antigen; Disease Models, Animal; Female; Interleukin-10; Lymph Nodes; Male; Mice; Mice, Inbred BALB C; Mice, SCID; Receptors, Interleukin-2; Spleen; Suppressor Factors, Immunologic; T-Lymphocyte Subsets; Transforming Growth Factor beta

Current treatments of inflammatory bowel diseases are limited either by their lack of efficacy or their potential toxicity. In recent years, major advances have been obtained by the development of biological therapies. However, these types of treatment are systemic and can lead to serious adverse events. The new venue of local biological treatments would be most welcome. In this issue of the Journal, Castagliuolo et al. show that lymphocytes engineered to produce TGF-beta1 can reverse dinitrobenzene sulphonic acid-induced colitis in mice. These engineered lymphocytes selectively accumulate in the intestinal mucosa due to the homing properties of their alpha4beta7 integrins, a ligand for MAdCAM1. A local treatment restricted to the inflamed mucosa can thus be obtained. This opens a brand new area of research with the hope of restoring the immunoregulatory balance selectively in the inflamed tissues.

Topics: Animals; Benzenesulfonates; Colitis; Genetic Therapy; Lymphocyte Transfusion; Lymphocytes; Mice; Transforming Growth Factor beta; Transforming Growth Factor beta1

Gene therapy is an attractive approach to the treatment of inflammatory diseases. However, the lack of tissue targeting of available vectors jeopardizes their clinical use.. Since alpha4beta7 integrin mediates lymphocyte homing to the intestinal mucosa, we tested the possibility of in-vitro engineering alpha4beta7-bearing lymphocytes to restrict the production of a therapeutic cytokine, transforming growth factor (TGF)-beta1, to within the colonic mucosa.. Lymphocytes were isolated from colonic lamina propria or spleen and transfected with either pC1 or pC1/TGF-beta1.. Transfected spleen and lamina propria cells released TGF-beta1 for up to 5 days in vitro and administration of 107 spleen cells, but not 106 lamina propria or spleen cells, to normal mice caused a significant rise in circulating TGF-beta1. Following intrarectal injection of dinitrobenzene sulphonic acid, intraperitoneal administration of lamina propria or spleen cells transfected with pC1/TGF-beta1, but not pC1, significantly reduced colitis-associated body weight loss, colonic myeloperoxidase (MPO) activity, interleukin-1beta levels, and macroscopic and microscopic inflammatory damage. Vector-specific TGF-beta1 mRNA transcripts were detectable in the colon and liver following injection of lamina propria lymphocytes, and in the spleen, liver and colon following administration of spleen lymphocytes. Incubation of pC1/TGF-beta1-transfected lamina propria lymphocytes with anti-alpha4beta7 integrin antibody blocked their protective effects and caused the disappearance of vector-specific TGF-beta1 transcripts from the colonic mucosa.. We conclude that lymphocytes are an efficient vehicle for transient gene therapy and that cells bearing alpha4beta7 integrins preferentially deliver therapeutic genes to the colonic mucosa.

Topics: Animals; Antibodies, Monoclonal; Benzenesulfonates; Colitis; Cytokines; Gene Targeting; Genetic Therapy; Integrins; Lymphocyte Transfusion; Lymphocytes; Male; Mice; Mice, Inbred BALB C; Mucous Membrane; Spleen; Transfection; Transforming Growth Factor beta; Transforming Growth Factor beta1

To elucidate extracellular matrix (ECM) changes underlying intestinal fibrosis, a frequent complication of inflammatory bowel disease, we developed a murine model of chronic colitis associated with intestinal fibrosis.. Chronic inflammation was established by weekly intrarectal administration of trinitrobenzene sulfonic acid (TNBS). In 2 variations of the model an antisense oligonucleotide for nuclear factor kappa B (NF-kappa B) p65 was given prophylactically or therapeutically to block chronic inflammation-associated fibrosis. Colonic inflammation and fibrosis were determined by histology. Total collagen level was estimated by hydroxyproline quantification. Colonic expression of collagens (Col1a2, Col3a2), ECM remodeling genes (matrix metalloproteinase [MMP]-1, -3, and tissue inhibitor of matrix metalloproteinase [TIMP]-1), and inflammation-modulating cytokines (tumor necrosis factor alpha [TNF-alpha], interferon gamma [IFN-gamma], transforming growth factor beta 1 [TGF-beta 1], and insulin-like growth factor 1 [IGF-1]) were assessed by semiquantitative reverse-transcription polymerase chain reaction. Control and TNBS-treated colonic mesenchymal cells were characterized by morphology, phenotype, and functional response to TNF-alpha and IFN-gamma.. Colons of TNBS-treated mice contained acute and chronic inflammatory infiltrates, increased collagen, fibrogenic tissue architecture, and increased expression of TNF-alpha, TGF-beta 1, IGF-1, Col1a2, MMP-1, and TIMP-1. Colonic mesenchymal cells from TNBS-treated mice were also morphologically distinct from those of the control mice, with increased TIMP-1 expression in response to IFN-gamma treatment. Fibrosis persisted for 2-4 weeks after cessation of the TNBS treatment. In mice given NF-kappa B antisense prophylactically, 67% were fibrosis-free, whereas of those treated after establishing chronic inflammation, 43% were free of fibrosis.. Extended TNBS treatment of mice yielded chronic intestinal inflammation-associated fibrosis with extensive fibrogenic ECM changes that could be counteracted by specific blockade of NF-kappa B.

Topics: Animals; Chronic Disease; Colitis; Collagen; Colon; Disease Models, Animal; Female; Fibrosis; Interferon-gamma; Matrix Metalloproteinase 1; Mice; Mice, Inbred BALB C; NF-kappa B; Oligonucleotides, Antisense; RNA, Messenger; Transcription Factor RelA; Transforming Growth Factor beta; Trinitrobenzenesulfonic Acid; Tumor Necrosis Factor-alpha

We have previously demonstrated that interleukin (IL)-10-deficient (IL-10 knockout [KO]) but not wild-type (WT) mice develop colitis after infection with Helicobacter hepaticus. Here, we show that infected recombination activating gene (RAG) KO mice develop intestinal inflammation after reconstitution with CD4(+) T cells from IL-10 KO animals and that the cotransfer of CD4(+) T cells from H. hepaticus-infected but not uninfected WT mice prevents this colitis. The disease-protective WT CD4(+) cells are contained within the CD45RB(low) fraction and unexpectedly were found in both the CD25(+) and the CD25(-) subpopulations of these cells, their frequency being higher in the latter. The mechanism by which CD25(+) and CD25(-) CD45RB(low) CD4(+) cells block colitis involves IL-10 and not transforming growth factor (TGF)-beta, as treatment with anti-IL-10R but not anti-TGF-beta monoclonal antibody abrogated their protective effect. In vitro, CD45RB(low) CD4(+) cells from infected WT mice were shown to produce IL-10 and suppress interferon-gamma production by IL-10 KO CD4(+) cells in an H. hepaticus antigen-specific manner. Together, our data support the concept that H. hepaticus infection results in the induction in WT mice of regulatory T cells that prevent bacteria-induced colitis. The induction of such cells in response to gut flora may be a mechanism protecting normal individuals against inflammatory bowel disease.

Topics: Adoptive Transfer; Animals; Antigens, Bacterial; Biomarkers; CD4-Positive T-Lymphocytes; Colitis; DNA-Binding Proteins; Female; Helicobacter; Helicobacter Infections; Interleukin-10; Interleukin-4; Leukocyte Common Antigens; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Receptors, Interleukin-2; Transforming Growth Factor beta

In the present study, we define the relation between TGF-beta and IL-10 in the regulation of the Th1-mediated inflammation occurring in trinitrobenzene sulfonic acid (TNBS)-colitis. In initial studies, we showed that the feeding of trinitrophenol-haptenated colonic protein to SJL/J mice induces CD4(+) regulatory T cells that transfer protection from induction of TNBS-colitis, and that such protection correlates with cells producing TGF-beta, not IL-10. Further studies in which SJL/J mice were fed haptenated colonic protein, and then administered either anti-TGF-beta or anti-IL-10 at the time of subsequent TNBS administration per rectum, showed that while both Abs abolished protection, anti-TGF-beta administration prevented TGF-beta secretion, but left IL-10 secretion intact; whereas anti-IL-10 administration prevented both TGF-beta secretion and IL-10 secretion. Thus, it appeared that the protective effect of IL-10 was an indirect consequence of its effect on TGF-beta secretion. To establish this point further, we conducted adoptive transfer studies and showed that anti-IL-10 administration had no effect on induction of TGF-beta producing T cells in donor mice. However, it did inhibit their subsequent expansion in recipient mice, probably by regulating the magnitude of the Th1 T cell response which would otherwise inhibit the TGF-beta response. Therefore, these studies suggest that TGF-beta production is a primary mechanism of counter-regulation of Th1 T cell-mediated mucosal inflammation, and that IL-10 is necessary as a secondary factor that facilitates TGF-beta production.

Topics: Adjuvants, Immunologic; Administration, Oral; Administration, Rectal; Adoptive Transfer; Animals; Antibodies, Monoclonal; CD4-Positive T-Lymphocytes; Colitis; Colon; Down-Regulation; Haptens; Interleukin-10; Lymphocyte Activation; Male; Mice; Mice, Inbred Strains; Picrates; T-Lymphocyte Subsets; Transforming Growth Factor beta; Trinitrobenzenesulfonic Acid

Interleukin (IL)-10 is a cytokine with anti-inflammatory properties. The aim of this study was to explore the effect of a site-specific delivery of IL-10 on intestinal immune responses.. Transgenic mice were created in which IL-10 is expressed by the intestinal epithelial cells.. Transgenic mice showed a marked increase in the number of intraepithelial lymphocytes in the small intestine. Mucosal lymphocytes of transgenic animals produced fewer T helper type 1 cytokines than wild-type lymphocytes. By contrast, the production of transforming growth factor beta was increased. Moreover, the epithelial layer in transgenic mice was significantly enriched for CD4(+)CD25(+) T cells. Furthermore, transgenic mice had increased numbers of immunoglobulin A-producing B cells in the small intestine. These effects were local because splenic lymphocytes were not affected. Studies in models of inflammatory bowel disease showed that transgenic IL-10 was able to attenuate the acute colitis induced by dextran sodium sulfate administration or by adoptive transfer of CD4(+)CD45RB(high) splenocytes, with a modest effect on the chronic intestinal inflammation arising spontaneously in IL-10(-/-) mice.. These observations provide evidence for an in vivo lymphoepithelial cross talk, by which cytokines locally produced by epithelial cells can regulate immune responses in the intestine without systemic modifications.

Topics: Animals; Antibody Formation; Colitis; Cytokines; Gene Targeting; Immunoglobulin A; Interleukin-10; Intestinal Mucosa; Intestine, Small; Lymphocyte Count; Lymphocytes; Mice; Mice, Inbred Strains; Mice, Knockout; Mice, Transgenic; Rats; Recombinant Proteins; T-Lymphocytes, Helper-Inducer; Tissue Distribution; Transforming Growth Factor beta; Transforming Growth Factor beta1

The aim of this study was to investigate cytokine expression patterns in the large intestinal mucosa of horses, particularly in diseases associated with inflammation. Many cases of equine colitis remain without a definitive diagnosis and survival rates are poor. In humans, colitis is associated with increased expression of pro-inflammatory cytokines. To examine if similar responses occur in horses, we investigated il -2, il -4, il -5, il -10, tnfalpha, ifngamma and tgfbeta messenger rna expression in large intestinal mucosa. Samples were obtained from animals with large intestinal disease (n=15) or from horses which had different levels of cyathostomin infection (n=9) and analysed by reverse transcription-polymerase chain reaction. il -2 was detected at all sites, whilst il -4 was detected at all but one site. The presence of il -10, il -5, ifngamma and tgfbeta varied with no significant differences amongst groups (P>0.4). Detection of tnfalpha was significantly different between the group of horses that had infiltrative inflammatory bowel disease and those with larval cyathostominosis (P=0.028) and those that were helminth negative (P=0.014). These results indicate a possible role for tnfalpha in the pathogenesis of equine infiltrative inflammatory bowel disease.

Topics: Animals; Colitis; Cytokines; Electrophoresis, Agar Gel; Horse Diseases; Horses; Interferon-gamma; Interleukin-10; Interleukin-2; Interleukin-4; Interleukin-5; RNA, Messenger; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

Trinitrobenzene sulfonic acid (TNBS)-induced colitis is an IL-12-driven, Th1 T cell-mediated colitis that resembles human Crohn's disease. In the present study, we showed initially that the oral administration of recombinant subunit B of cholera toxin (rCT-B) at the time of TNBS-induced colitis by intrarectal TNBS instillation inhibits the development of colitis or, at later time when TNBS-induced colitis is well established, brings about resolution of the colitis. Dose-response studies showed that a majority of mice (68%) treated with rCT-B at a dose of 100 microg (times four daily doses) exhibited complete inhibition of the development of colitis, whereas a minority (30%) treated with rCT-B at a dose of 10 microg (times four daily doses) exhibited complete inhibition; in both cases, however, the remaining mice exhibited some reduction in the severity of inflammation. In further studies, we showed that rCT-B administration is accompanied by prevention/reversal of increased IFN-gamma secretion (the hallmark of a Th1 response) without at the same time causing an increase in IL-4 secretion. This decreased IFN-gamma secretion was not associated with the up-regulation of the secretion of counterregulatory cytokines (IL-10 or TGF-beta), but was associated with a marked inhibition of IL-12 secretion, i.e., the secretion of the cytokine driving the Th1 response. Finally, we showed that rCT-B administration results in increased apoptosis of lamina propria cells, an effect previously shown to be indicative of IL-12 deprivation. From these studies, rCT-B emerges as a powerful inhibitor of Th1 T cell-driven inflammation that can conceivably be applied to the treatment of Crohn's disease.

Topics: Administration, Oral; Administration, Rectal; Animals; Apoptosis; Cells, Cultured; Cholera Toxin; Colitis; Colon; Disease Progression; Interferon-gamma; Interleukin-10; Interleukin-12; Interleukin-4; Intestinal Mucosa; Leukocytes, Mononuclear; Male; Mice; Mice, Inbred Strains; Oxazolone; Recombinant Proteins; Transforming Growth Factor beta; Trinitrobenzenesulfonic Acid

Using a CD4+ T-cell-transplanted SCID mouse model of colitis, we have analyzed TGF-beta transcription and translation in advanced disease. By in situ hybridization, the epithelium of both control and inflamed tissues transcribed TGF-beta1 and TGF-beta3 mRNAs, but both were expressed significantly farther along the crypt axis in disease. Control lamina propria cells transcribed little TGF-beta1 or TGF-beta3 mRNA, but in inflamed tissues many cells expressed mRNA for both isoforms. No TGF-beta2 message was detected in either control or inflamed tissues. Immunohistochemistry for latent and active TGF-beta1 showed that all cells produced perinuclear latent TGF-beta1. The epithelial cell basal latent protein resulted in only low levels of subepithelial active protein, which co-localized with collagen IV and laminin in diseased and control tissue. Infiltrating cells expressed very low levels of active TGF-beta. By ELISA, very low levels (0-69 pg/mg) of soluble total or active TGF-beta were detected in hypotonic tissue lysates. TGF-beta1 and TGF-beta3 are produced by SCID mouse colon and transcription is increased in the colitis caused by transplantation of CD4+ T-cells, but this does not result in high levels of soluble active protein. Low levels of active TGF-beta may be a factor contributing to unresolved inflammation.

Topics: Animals; CD4-Positive T-Lymphocytes; Colitis; Colon; Connective Tissue; Enzyme-Linked Immunosorbent Assay; Fluorescent Antibody Technique, Indirect; Immunohistochemistry; In Situ Hybridization; Inflammatory Bowel Diseases; Mice; Mice, SCID; RNA, Messenger; Tissue Distribution; Transforming Growth Factor beta

Topics: Animals; Animals, Genetically Modified; Cell Communication; Colitis; Genes, MHC Class II; Homeostasis; Intestinal Mucosa; Matrix Metalloproteinases; Mice; Mice, Knockout; Mice, SCID; Transforming Growth Factor beta

Studies conducted over the past 10 years have provided ample evidence that many types of inflammations arising from basic abnormalities of immune regulation are ultimately 'funneled' through a Th1 or Th2 T cell-mediated immune reaction. Thus, by understanding these types of reactions and, in particular, by identifying their natural checkpoints, one can control the inflammation regardless of its more basic causes. A case in point is the inflammatory disease of the intestine known as Crohn disease, a disease now thought to be due to one or more abnormalities leading to an excessive immune response to elements of the bacterial microflora of the gut. Both in murine models and by study of Crohn disease itself, we have shown that Crohn inflammation is due to a Th1 T-cell abnormality involving overproduction of interleukin (IL)-12, interferon (IFN-gamma, and tumor necrosis factor (TNF)-alpha. In addition, we and others have shown that treatment of mice with anti-IL-12 or other agents that downregulate the level of IL- 12 secretion can have a dramatic effect on the inflammation. This is because anti-IL-12 administration leads to apoptosis of activated Th1 T cells. A second checkpoint of Th1 T-cell-mediated inflammation involves its downregulation by the suppressor cytokine, transforming growth factor (TGF)-beta. We have been delivering TGF-beta to mice with experimental intestinal inflammation, using several novel approaches. In particular, we have successfully treated such mice with intranasally administered DNA encoding active TGF-beta. Another approach currently under investigation is delivery of TGF-beta by gene therapy. These and other developments in the understanding of inflammation paint a bright future for cytokine-based therapeutic agents. It is now apparent that these therapies are not only effective and safe but also potentially long-lasting.

Topics: Animals; Colitis; Down-Regulation; Gene Transfer Techniques; Immunotherapy; Interleukin-10; Interleukin-12; Intestinal Mucosa; Mice; Mice, Inbred Strains; T-Lymphocyte Subsets; Th1 Cells; Transforming Growth Factor beta; Trinitrobenzenesulfonic Acid; Tumor Necrosis Factor-alpha

Rectal administration of trinitrobenzene sulfonic acid (TNBS) produces chronic colitis in experimental animals. However, the role of epithelial cellular protein(s) in this model is unknown. We examined whether oral tolerance can be induced in this model with colon epithelial cell proteins and whether it is organ specific. Rats were fed five times with extracts of LS-180 human colon cancer cells or HT 1080 human fibroblast cells. Syngeneic normal rat colon or small intestinal extracts were fed to separate groups of rats. After oral feedings, each rat received TNBS by enema. Rats were killed 15 days later, and the following were measured: gross and histologic disease score, weight, thickness, and myeloperoxidase values of colon and serum interferon-gamma (IFN-gamma) and transforming growth factor-beta (TGF-beta) levels. Rectal TNBS alone produced severe colitis with a 26% mortality rate. Rats fed LS-180 or rat colon extract before TNBS enema were protected, as evidenced by reductions in mortality rate, disease scores, and myeloperoxidase values. However, rats fed HT 1080 or small intestine extract lacked such protection. To examine the possible mechanism of the oral tolerance, T lymphocytes from mesenteric lymph nodes and spleen of LS-180 extract-fed rats were passively transferred to naive rats, and this was followed by TNBS enema. These rats showed clear protection. Protected animals had low IFN-gamma and high TGF-beta levels. This study demonstrates that cellular protein(s) from human colon epithelial cells, but not from human fibroblasts, can induce oral tolerance in experimental colitis. This oral tolerance is mediated by primed mesenteric and splenic T lymphocytes.

Topics: Administration, Oral; Animals; Cell Extracts; Colitis; Colon; Colonic Neoplasms; Epithelium; Female; Fibroblasts; Humans; Immune Tolerance; Immunization, Passive; Interferon-gamma; Intestine, Small; Lymph Nodes; Peroxidase; Proteins; Rats; Rats, Sprague-Dawley; Rectum; Spleen; Tissue Extracts; Transforming Growth Factor beta; Trinitrobenzenesulfonic Acid; Tumor Cells, Cultured

Transgenic rats expressing HLA-B27 and human beta2-microglobulin (HLA-B27 rats) spontaneously develop chronic colitis resembling human inflammatory bowel disease. We investigated the sequential changes in the luminal bacterial flora and mucosal cytokine mRNA expression in this model.. HLA-B27 rats were maintained in a specific pathogen-free environment, and luminal microflora was evaluated by standard bacterial culture technique. The expression of mucosal cytokine mRNA was analysed by RT-PCR methods.. Clinical symptoms of colitis appeared at 8 weeks of age. The total number of obligate anaerobes was higher than those of facultative anaerobes during the experimental period. At 6 weeks of age, the colonization of Bacteroides spp., Bifidobacterium spp. and Lactobacillus spp. was already detectable at high concentrations, whereas Clostridium spp. and Eubacterium spp. were not detected. The expression of proinflammatory cytokines (IL-Ibeta, IL-8 and TNF-alpha) appeared at 8 weeks of age, and these were detectable until 17 weeks. A similar pattern was observed in the expression of Th1 cytokines (IL-2, IL-12 and IFN-gamma). On the other hand, the expression of Th2 cytokines (IL-4, IL-10 and TGF-beta) was weak. IL-4 mRNA expression was weakly detectable only at 6 and 8 weeks of age. The expression of IL-10 and TGF-beta mRNA was scarcely detectable throughout the experimental period.. The development of colitis may be mediated by both the predominant expression of Th1 cytokines and the weakness of Th2 cytokine expression in the mucosa. The colonization of anaerobic bacteria, especially Bacteroides spp., may be initiating and promoting these cytokine responses.

Topics: Animals; Bacteroides; Bifidobacterium; Colitis; Cytokines; HLA-B27 Antigen; Interferon-gamma; Interleukin-10; Interleukin-12; Interleukin-2; Interleukin-4; Intestinal Mucosa; Lactobacillus; Mice; Mice, Transgenic; Polymerase Chain Reaction; RNA, Messenger; Specific Pathogen-Free Organisms; Transforming Growth Factor beta

Verotoxin-producing Escherichia coli (VTEC) cause hemorrhagic colitis (HC) and hemolytic uremic syndrome (HUS). The aim of this study was to compare the circulating levels of transforming growth factor-beta 1 (TGF-beta1), T helper (T(H))1 (interferon [IFN]-gamma, interleukin [IL]-2), and T(H)2-associated lymphokines (IL-4, IL-13) in children with uncomplicated Escherichia coli O157:H7 HC and patients who developed HUS. Circulating levels of IL-2, IL-4, and IL-13 were undetectable, and those of IFN-gamma were low and comparable among groups. Concentrations of TGF-beta1 were higher in children with uncomplicated O157:H7 HC than among those who developed HUS (934 +/- 680 versus 514 +/- 497 pg/mL, respectively; P < 0.04). The circulating levels of TGF-beta1 were also higher among children who did not take antidiarrheal agents (P < 0.008) and those who have been immediately discharged from the emergency room (P < 0.03). Our results did not show an imbalanced T(H)1/T(H)2-associated lymphokine response during the development of HUS. Increased circulating levels of TGF-beta1 in children with milder O157:H7 or uncomplicated HC most likely reflect appropriate intestinal tissue repair mechanisms rather than a remote systemic endocrine effect on the kidneys.

Topics: Adolescent; Child; Child, Preschool; Colitis; Escherichia coli Infections; Escherichia coli O157; Female; Hemolytic-Uremic Syndrome; Humans; Infant; Lymphocyte Count; Lymphokines; Male; Th1 Cells; Th2 Cells; Transforming Growth Factor beta

In this study, we show that a single intranasal dose of a plasmid encoding active transforming growth factor beta1 (pCMV-TGF-beta1) prevents the development of T helper cell type 1 (Th1)-mediated experimental colitis induced by the haptenating reagent, 2,4, 6-trinitrobenzene sulfonic acid (TNBS). In addition, such plasmid administration abrogates TNBS colitis after it has been established, whereas, in contrast, intraperitoneal administration of rTGF-beta1 protein does not have this effect. Intranasal pCMV-TGF-beta1 administration leads to the expression of TGF-beta1 mRNA in the intestinal lamina propria and spleen for 2 wk, as well as the appearance of TGF-beta1-producing T cells and macrophages in these tissues, and is not associated with the appearances of fibrosis. These cells cause marked suppression of interleukin (IL)-12 and interferon (IFN)-gamma production and enhancement of IL-10 production; in addition, they inhibit IL-12 receptor beta2 (IL-12Rbeta2) chain expression. Coadministration of anti-IL-10 at the time of pCMV-TGF-beta1 administration prevents the enhancement of IL-10 production and reverses the suppression of IL-12 but not IFN-gamma secretion. However, anti-IL-10 leads to increased tumor necrosis factor alpha production, especially in established colitis. Taken together, these studies show that TGF-beta1 inhibition of a Th1-mediated colitis is due to: (a) suppression of IL-12 secretion by IL-10 induction and (b) inhibition of IL-12 signaling via downregulation of IL-12Rbeta2 chain expression. In addition, TGF-beta1 may also have an inhibitory effect on IFN-gamma transcription.

Topics: Administration, Intranasal; Animals; Colitis; Colon; Cytokines; Cytomegalovirus; Genetic Therapy; Injections, Intraperitoneal; Intestinal Mucosa; Male; Mice; Mice, Inbred Strains; Plasmids; Recombinant Proteins; Swine; Th1 Cells; Transforming Growth Factor beta; Trinitrobenzenesulfonic Acid

It is now clear that functionally specialized regulatory T (Treg) cells exist as part of the normal immune repertoire, preventing the development of pathogenic responses to both self- and intestinal antigens. Here, we report that the Treg cells that control intestinal inflammation express the same phenotype (CD25(+)CD45RB(low)CD4(+)) as those that control autoimmunity. Previous studies have failed to identify how CD25(+) Treg cells function in vivo. Our studies reveal that the immune-suppressive function of these cells in vivo is dependent on signaling via the negative regulator of T cell activation cytotoxic T lymphocyte-associated antigen 4 (CTLA-4), as well as secretion of the immune-suppressive cytokine transforming growth factor beta. Strikingly, constitutive expression of CTLA-4 among CD4(+) cells was restricted primarily to Treg cells, suggesting that CTLA-4 expression by these cells is involved in their immune-suppressive function. These findings raise the possibility that Treg cell function contributes to the immune suppression characteristic of CTLA-4 signaling. Identification of costimulatory molecules involved in the function of Treg cells may facilitate further characterization of these cells and development of new therapeutic strategies for the treatment of inflammatory diseases.

Topics: Abatacept; Animals; Antigens, CD; Antigens, Differentiation; CD4-Positive T-Lymphocytes; Colitis; CTLA-4 Antigen; Immunoconjugates; Leukocyte Common Antigens; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Receptors, Interleukin-2; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Collagenous colitis is a disease characterized by chronic watery diarrhea, and on microscopic examination of colonic tissue, a typical thickening of the subepithelial collagen layer is seen. The etiology and pathophysiology behind this disease state are largely unknown.. We have used in situ hybridization and immunohistochemistry to study the expression of transforming growth factor (TGF) -beta1, a growth factor with the capacity to cause accumulation of collagen in tissues, in collagenous colitis. Colonic pinch biopsy specimens from a total of 34 patients were investigated: 17 patients with collagenous colitis and 17 controls.. In patients with collagenous colitis there was increased expression of the TGF-beta1 gene compared with controls, as visualized by in situ hybridization. The vast majority of the TGF-beta1-expressing cells were eosinophils, both in collagenous colitis and controls, but there were also scattered fibroblastic and histiocytic stromal cells. Immunohistochemistry showed the presence of TGF-beta1, mainly in eosinophils, in the colonic mucosa. Morphometric quantification showed 603 +/- 192 eosinophils/mm2, (mean +/- standard error of the mean) in the colonic mucosa of patients with collagenous colitis compared with 30 +/- 7 eosinophils/mm2 in the controls.. The present results suggest that eosinophils expressing TGF-beta1 may be of pathophysiologic importance in the connective tissue remodeling seen in collagenous colitis.

Topics: Adult; Aged; Aged, 80 and over; Colitis; Collagen; Colon; Eosinophils; Female; Humans; Immunohistochemistry; In Situ Hybridization; Intestinal Mucosa; Male; Middle Aged; Transforming Growth Factor beta; Transforming Growth Factor beta1

Bacteria and their products stimulate inflammatory responses. Certain mediators, such as transforming growth factor beta1 (TGF-beta1), induce collagen synthesis. Excess collagen deposition results in bowel strictures. The aim of this study was to investigate the role of bacteria and TGF-beta1 in the pathogenesis of intestinal fibrosis.. In rats with colitis, the effects of bowel decontamination with antibiotics on TGF-beta1, tumor necrosis factor alpha (TNF-alpha), and collagen content in colonic tissue were studied. In normal rats, bacteria of the predominant flora were inoculated into the colonic wall. The effect of neutralizing antibody to TGF-beta1 on tissue collagen deposition was studied.. Rats with chronic colitis showed increased levels of TGF-beta1, TNF-alpha, and collagen in the tissue and a high rate of bowel strictures. Antibiotic treatment significantly prevented the increase in TGF-beta1 and collagen and the formation of strictures. Inoculation of bacterial suspensions into the colonic wall increased tissue TGF-beta1 and collagen content. Neutralizing antibody to TGF-beta1 prevented collagen deposition. Colonic wall inoculations with single anaerobic strains (Clostridium ramosum, Bacteroides fragilis, and Bacteroides uniformis), but not with aerobes, induced collagen deposition.. Certain strains of the common flora stimulate TGF-beta1 and induce deposition of collagen in the colonic wall.

Topics: Animals; Anti-Bacterial Agents; Bacteria; Colitis; Collagen; Fibrosis; Intestinal Obstruction; Intestines; Male; Rats; Rats, Sprague-Dawley; Transforming Growth Factor beta

Intestinal epithelial cells may be actively involved in the immunoregulatory pathways leading to intestinal inflammation. The aim of this study was to assess expression by intestinal epithelial cells of cytokines with potential involvement in the development of intestinal inflammation in interleukin (IL)-2-deficient [(-/-)] mice. Wild-type mice, mice heterozygous for the disrupted IL-2 gene, and IL-2(-/-) mice were studied at 6, 16, and 24 wk of age. The mRNA levels of transforming growth factor-beta 1 (TGF-beta 1), tumor necrosis factor-alpha (TNF-alpha), IL-1 beta, IL-6, IL-15, KC, JE, and CD14 in colonic and small intestinal epithelial cells were assessed by Northern blot analysis. CD14 was also measured by Western blotting and reverse transcriptase polymerase chain reaction (RT-PCR). TGF-beta 1 mRNA was constitutively expressed in both colonic and small intestinal epithelial cells with increased expression in the colonic epithelium of colitic mice. CD14 was detected only in colonic epithelial cells, and mRNA levels increased severalfold in IL-2(-/-) mice with colitis. Northern analysis demonstrated increased levels of TGF-beta 1 and CD14 mRNA in colonic epithelial cells of IL-2(-/-) mice before the development of signs of colitis. CD14 mRNA and protein expression in the epithelial cells of colitic mice were confirmed by RT-PCR and Western blot analysis of isolated cells. In addition, IL-2(-/-) mice also expressed increased levels of IL-15 mRNA in small intestinal and colonic epithelial cells compared with heterozygous control mice. TNF-alpha, IL-1 beta, IL-6, KC, and JE mRNAs were only detectable in colonic epithelial cells of mice after the onset of colitis. Enhanced expression of TGF-beta 1, IL-15, and CD14 by colonic epithelial cells may play a role in the subsequent development of colitis in IL-2(-/-) mice.

Topics: Animals; Colitis; Colon; Gene Expression; Interleukin-15; Interleukin-2; Intestinal Mucosa; Intestine, Small; Lipopolysaccharide Receptors; Mice; Mice, Inbred C57BL; RNA, Messenger; Transforming Growth Factor beta

Recombinant human interleukin-11 (rhIL-11) is a pleiotropic cytokine with effects on multiple cell types. In addition to thrombopoietic activity, rhIL-11 has demonstrated anti-inflammatory activity in vitro and in vivo. rhIL-11 treatment reduces clinical signs and histologic lesions of colitis in transgenic rats expressing the human major histocompatibility complex (MHC) Class I allele, HLA-B27. We have investigated the effects of rhIL-11 at the molecular and cellular level in this model of inflammatory bowel disease. RT-PCR analysis of colonic RNA revealed that treatment with rhIL-11 down-regulated expression of proinflammatory cytokines including TNF-alpha, IL-1beta, and IFN-gamma. rhIL-11 also reduced the level of myeloperoxidase activity in the cecum indicating reduced inflammation. After stimulation in vitro with anti-CD3 antibody, spleen cell cultures derived from rhIL-11-treated rats produced less IFN-gamma, TNF-alpha, and IL-2 than cultures derived from vehicle-treated rats. These molecular and cellular effects correlated with amelioration of disease as measured by stool character and histologic lesion scores. These findings suggest that rhIL-11 acts to reduce inflammation through modulation of multiple proinflammatory mediators including products of activated T cells. This study has identified pharmacodynamic markers of rhIL-11 anti-inflammatory activity in vivo and supports rhIL-11 therapy to treat inflammatory bowel disease.

Topics: Animals; Animals, Genetically Modified; beta 2-Microglobulin; Cecum; Cells, Cultured; Colitis; Colon; Cytokines; Disease Models, Animal; Gene Expression Regulation; Genes, MHC Class I; HLA-B27 Antigen; Humans; Inflammatory Bowel Diseases; Interferon-gamma; Interleukin-11; Interleukins; Lymphocytes; Male; Rats; Rats, Inbred F344; Recombinant Proteins; Reference Values; Reverse Transcriptase Polymerase Chain Reaction; Spleen; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

Chronic colitis develops spontaneously in mice mutant for T-cell receptor alpha (TCR alpha) genes maintained in pathogen-free conditions. Our previous studies indicate that the cytokine imbalance caused by a lack of TCR alpha beta + T cells may be associated with the development of chronic colitis in TCR alpha-mutant (TCR alpha -/-) mice. The histologic changes of chronic colitis are recognizable by 3 to 4 months of age and are characterized by marked crypt cell hyperplasia and the presence of an inflammatory cell infiltrate. Because early events in the development of chronic colitis may be important in the pathogenesis of the disorder, we investigated the changes in the colon at a time when chronic colitis was not yet histologically recognizable. In vivo labeling with 5-bromo-2'-deoxyuridine revealed increased epithelial proliferation in the crypts by 6 to 8 weeks of age. There was an increase in number of T cells in the colon of TCR alpha -/- mice compared with that in TCR alpha +/- (heterozygous TCR alpha-mutant) mice after 6 weeks of age. The predominant T-cell subsets were CD4+, TCR alpha- beta +, and TCR gamma delta + cells. Reverse transcription-PCR and immunohistochemistry of the colonic mucosa obtained from TCR alpha -/- mice (> 6 weeks old) showed a marked increase of IL-1 alpha and IL-1 beta but not of TNF alpha or transforming growth factor beta-1 compared with TCR alpha +/- mice. In vivo neutralization of IL-1 alpha or IL-1 beta by specific mAb suppressed colonic epithelial proliferation and decreased colonic mucosal T-cell infiltration in 8-week-old TCR alpha -/- mice. These results provide direct evidence that overproduction of IL-1 alpha and IL-1 beta in the colonic mucosa may play an important role in the early stages of development of chronic colitis in TCR alpha -/- mice.

Topics: Animals; Antibodies, Monoclonal; Chronic Disease; Colitis; Colon; Cricetinae; Crosses, Genetic; Cytokines; Interleukin-1; Intestinal Mucosa; Lymphocytes; Mice; Mice, Inbred C57BL; Mice, Knockout; Polymerase Chain Reaction; Receptors, Antigen, T-Cell, alpha-beta; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

Superantigens (SAgs) are extremely potent stimulants of T cell activity that have been implicated in the etiopathophysiology of inflammatory disease. Here, we tested the hypothesis that Staphylococcus aureus enterotoxin B (SEB), a model SAg, can alter epithelial transport and/or barrier functions via immune stimulation. Confluent monolayers of the human colonic T84 epithelial cell line, grown on filter supports, were cocultured with SEB +/- PBMC. Subsequently, T84 transport (consisting of baseline short-circuit current (Isc, indicates net ion transport) and secretory responses to carbachol and forskolin) and barrier functions (consisting of transepithelial resistance and fluxes of 3H-labeled mannitol and 51Cr-EDTA) were examined in Ussing chambers. T84 monolayers cocultured with SEB-activated PBMC displayed a time- and dose-dependent decrease in secretory responses to carbachol and forskolin and a significant increase in permeability. These dramatic changes in epithelial function were not due to reduced epithelial viability. Neutralizing Abs to IFN-gamma partially prevented the transport abnormalities, and Abs to TNF-alpha inhibited the increase in epithelial permeability. Abs to IL-1beta and IL-6 did not modulate the SEB-activated PBMC-induced T84 pathophysiology. Addition of TGF-beta2 to conditioned medium from SEB-activated PBMC partially inhibited the increase in T84 permeability but did not affect the transport abnormalities. We conclude that SAgs can elicit epithelial irregularities characteristic of enteric inflammation and that IFN-gamma and TNF-alpha are key mediators in this coculture model of epithelial dysfunction. Additionally, we would highlight the role that TGF-beta2 may play in preventing prolonged increases in epithelial permeability.

Topics: Adult; Antibodies, Monoclonal; Antigens, Bacterial; Biological Transport; Carbachol; Cell Line; Cell Membrane Permeability; Coculture Techniques; Colforsin; Colitis; Colon; Culture Media, Conditioned; Enterotoxins; Epithelial Cells; Epithelium; Female; Humans; Interferon-gamma; Intestinal Absorption; Leukocytes, Mononuclear; Male; Middle Aged; Recombinant Proteins; Superantigens; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

A severe, Th1-mediated experimental colitis with similarities to inflammatory bowel disease in humans can be induced by a single injection of 2,4,6-trinitrophenol (TNP)-substituted protein plus adjuvant in IL-2-/- mice. To determine the early events involved in the pathogenesis of IL-2-/-colitis, we compared the function of lamina propria (LP) T cells from IL-2-/- and IL-2+/+ mice subjected to disease-inducing (TNP-conjugated keyhole limpet hemocyanin [TNP-KLH]) and disease-inhibiting (anti-CD3) immunization protocols. We show that LP T cells in TNP-KLH-immunized IL-2-/- mice fail to produce TGF-beta early (day 2), whereas LP T cells in TNP-KLH-immunized IL-2+/+ mice exhibit an approximately eightfold rise in TGF-beta secretion. The critical importance of local TGF-beta production was further substantiated by the following findings. 1) LP T cells from TNP-KLH-immunized IL-2-/- mice administered anti-CD3 (i.p.) exhibit a significant rise in TGF-beta, production but fail to produce IFN-gamma, and such mice do not develop colitis. 2) TNP-KLH-immunized IL-2-/- mice administered anti-CD3 and coadministered anti-TGF-beta mAb again give rise to IFN-gamma-producing LP cells, and such mice develop colitis. 3) TNP-KLH-immunized IL-2+/+ mice administered anti-TGF-beta mAb exhibit pockets of mononuclear cell infiltrates in the LP. These results indicate that the disposition of IL-2-/- mice to develop chronic colonic inflammation is due to a Th1 cell response in the LP that is not appropriately counter-regulated by the production of the suppressor cytokine, TGF-beta.

Topics: Animals; Antibodies, Monoclonal; CD3 Complex; Colitis; Female; Haptens; Hemocyanins; Interleukin-2; Interleukin-4; Male; Mice; Mice, Knockout; Picrates; Th1 Cells; Transforming Growth Factor beta

Polyunsaturated phosphatidylcholine stimulates collagen breakdown in experimental models of liver cirrhosis. Bowel strictures are characterized by excess deposition of collagen in the intestinal wall. The aim of this study was to investigate the effect of polyunsaturated phosphatidylcholine in the prevention of bowel strictures.. Colitis was induced by trinitrobenzenesulfonic acid. On day 21, the presence of strictures was assessed in control rats, rats with colitis, and phosphatidylcholine-fed (100 mg/day) rats with colitis. Furthermore, serum transforming growth factor beta1, collagen deposition, and collagenase activity in colonic tissue were measured in all groups.. None of the control rats but 12 of 16 rats with colitis developed colonic strictures. In contrast, only 2 of 15 phosphatidylcholine-fed rats with colitis showed strictures. Collagen content was much higher in rats with colitis than in phosphatidylcholine-fed rats with colitis and control rats. Phosphatidylcholine-fed rats showed significantly higher collagenase activity in colonic tissue than rats with colitis and control rats. In an ancillary study, free linoleic acid-fed rats showed no differences when compared with rats with colitis. Stimulation of transforming growth factor beta1 was similar in all rats with colitis.. Oral supplementation with polyunsaturated phosphatidylcholine prevents the accumulation of collagen in inflamed intestinal tissue and the formation of strictures. This effect is associated with an enhanced collagen catabolism.

Topics: Analysis of Variance; Animals; Chronic Disease; Colitis; Collagen; Collagenases; Colon; Colonic Diseases; Constriction, Pathologic; Dietary Fats, Unsaturated; Disease Models, Animal; Intestinal Obstruction; Male; Phosphatidylcholines; Rats; Rats, Sprague-Dawley; Transforming Growth Factor beta; Trinitrobenzenesulfonic Acid

In previous studies we showed that a chronic colitis associated with a Th1 T cell response can be induced by the rectal administration of the haptenizing reagent 2,4,6-trinitrobenzene sulfonic acid (TNBS). We report here that oral administration of haptenized colonic proteins (HCP) before rectal administration of TNBS effectively suppresses the ability of the latter to induce colitis. This suppression (oral tolerance) appears to be due to the generation of mucosal T cells producing TGF-beta and Th2-type cytokines after oral HCP administration. Peyer's patch and lamina propria CD4+ T cells from HCP-fed animals stimulated with anti-CD3/anti-CD28 had a 5-10-fold increase in their production of TGF-beta and secreted increased amounts of IL-4 and IL-10 but lower levels of IFN-gamma in comparison to T cells from ovalbumin-fed control animals. In addition, the colons of HCP-fed mice showed strikingly increased TGF-beta but decreased IL-12 expression by immunohistochemical studies and isolated mononuclear cells from HCP-fed animals secreted less IL-12 heterodimer. Finally, and most importantly, the suppressive effect of orally administered HCP was abrogated by the concomitant systemic administration of anti-TGF-beta or rIL-12 suggesting a reciprocal relationship between IL-12 and TGF-beta on tolerance induction in TNBS-induced colitis. In parallel studies we demonstrated that TNBS-induced colitis can be transferred to naive recipient animals with purified CD4+ T cells from the colon of TNBS-treated animals and that such animals develop lethal pancolitis when exposed to very low doses of TNBS. Feeding of HCP suppressed this sensitivity to TNBS, indicating that oral feeding can suppress the response of pre-committed T cells in vivo. These studies suggest for the first time that TGF-beta production can abrogate experimental granulomatous colitis even after such colitis is established, and thus, that regulation of TGF-beta levels may have relevance to the treatment of human inflammatory bowel disease.

Topics: Animals; Antibodies; CD28 Antigens; CD3 Complex; CD4-Positive T-Lymphocytes; Colitis; Cytokines; Female; Granulomatous Disease, Chronic; Haptens; Humans; Immune Tolerance; Immunotherapy, Adoptive; Interleukin-10; Interleukin-4; Intestinal Mucosa; Mice; Mice, Inbred Strains; Ovalbumin; Peyer's Patches; T-Lymphocytes; Th1 Cells; Transforming Growth Factor beta; Trinitrobenzenesulfonic Acid

A T helper type 1 (Th1)-mediated colitis with similarities to inflammatory bowel disease in humans developed in severe combined immunodeficiency mice reconstituted with CD45RB(high) CD4+ splenic T cells and could be prevented by cotransfer of CD45RB(low) CD4+ T cells. Inhibition of this Th1 response by the CD45RB(low) T cell population could be reversed in vivo by an anti-transforming growth factor (TGF) beta antibody. Interleukin (IL) 4 was not required for either the differentiation of function of protective cells as CD45RB(low) CD4+ cells from IL-4-deficient mice were fully effective. These results identify a subpopulation of peripheral CD4+ cells and TGF-beta as critical components of the natural immune regulatory mechanism, which prevents the development of pathogenic Th1 responses in the gut, and suggests that this immunoregulatory population is distinct from Th2 cells.

Topics: Animals; Antibodies, Monoclonal; CD4-Positive T-Lymphocytes; Colitis; Flow Cytometry; Humans; Interleukin-4; Leukocyte Common Antigens; Mice; Mice, Inbred BALB C; Mice, Mutant Strains; Mice, SCID; Protein Tyrosine Phosphatase, Non-Receptor Type 1; Spleen; T-Lymphocytes; Th1 Cells; Transforming Growth Factor beta

To modulate experimental colitis by the direct intramuscular injection of cDNA expression vector encoding transforming growth factor-beta 1.. Colitis was induced in rats by the intracolonic administration of 0.25 ml 50% ethanol containing 30 mg trinitrobenzene sulfonic acid. Three and 10 days later the rats were injected intramuscularly with 200 micrograms transforming growth factor-beta 1 cDNA subcloned to the pRSV expression vector (pRSVTGF-beta 1). Control rats were injected with pRSVP2. The rats were sacrificed 2 weeks after trinitrobenzene sulfonic acid treatment and a 10 cm long distal colonic segment was resected, weighted, the lesion area measured, sections obtained for histology and the mucosa extracted for determination of myeloperoxidase activity and leukotriene generation.. In pRSVP2-treated rats (n = 17) the colon was inflamed and ulcerated with many adhesions to adjacent structures; the pRSVTGF-beta 1-treated rats (n = 21) had minimal swelling and inflammation in the colon. On histological examination 50% of the pRSVTGF-beta 1-treated rats had minimal or no ulceration, whereas 83% of the pRSVP2-treated rats had a maximal damage score. In pRSVTG-beta 1-treated rats the lesion area and wet weight were 21 and 52.5%, respectively, of the values for pRSVP2-treated rats (P < 0.05). The amelioration of tissue injury was accompanied by a significant decrease in mucosal leukotriene C4 generation.. Direct intramuscular transforming growth factor-beta 1 gene delivery effectively ameliorates trinitrobenzene sulfonic acid-induced colitis, suggesting that gene therapy with immunosuppressive cytokines may be a novel approach for the treatment of inflammatory bowel disease.

Topics: Animals; Colitis; DNA, Complementary; Genetic Therapy; Genetic Vectors; Male; Rats; Rats, Inbred Strains; Transforming Growth Factor beta; Trinitrobenzenesulfonic Acid