transforming-growth-factor-beta and Hyperplasia

Revascularization surgeries such as coronary artery bypass grafting (CABG) are sometimes necessary to manage coronary heart disease (CHD). However, more than half of these surgeries fail within 10 years due to the development of intimal hyperplasia (IH) among others. The cytokine transforming growth factor-beta (TGFß) and its signaling components have been found to be upregulated in diseased or injured vessels, and to promote IH after grafting. Interventions that globally inhibit TGFß in CABG have yielded contrasting outcomes in

Topics: Activin Receptors, Type II; Coronary Artery Bypass; Coronary Disease; Graft Occlusion, Vascular; Humans; Hyperplasia; Myocytes, Smooth Muscle; Receptor, Transforming Growth Factor-beta Type I; Signal Transduction; Smad2 Protein; Transforming Growth Factor beta

Topics: Animals; Bile Acids and Salts; Cell Division; Chimera; G1 Phase; Hepatectomy; Hepatocytes; Humans; Hyperplasia; Liver; Liver Regeneration; Mice; Receptors, Transforming Growth Factor beta; S Phase; Transforming Growth Factor beta

Transforming growth factor (TGF)-beta is a multifunctional cytokine involved in the regulation of proliferation, differentiation, migration, and survival of many different cell types. The role of TGF-beta in atherosclerosis has been intensively studied, but the precise function of the downstream signals in this disease entity remains unclear. We recently discovered that mice lacking Smad3, a major downstream mediator of TGF-beta, show enhanced neointimal hyperplasia with decreased matrix deposition in response to vascular injury. This review summarizes the current view on involvement of TGF-beta in atherosclerotic vascular disease and discusses the role of Smad3-dependent TGF-beta signal in vascular response to injury.

Topics: Animals; Homeostasis; Humans; Hyperplasia; Matrix Metalloproteinases; Myocytes, Smooth Muscle; Signal Transduction; Smad3 Protein; Transforming Growth Factor beta; Tunica Intima; Vascular Diseases

Topics: Animals; Body Weight; Cell Cycle; Cyclin-Dependent Kinase Inhibitor p21; Cyclin-Dependent Kinases; Cyclins; Diet Therapy; Hyperplasia; Hypertrophy; Kidney; Mice; Mice, Knockout; Nephrons; Renal Insufficiency; Transforming Growth Factor beta; Uremia

The enlarged muscles of certain breeds of cattle, such as the Belgian Blue, have been shown to result from a marked increase in the number of normal sized muscle fibers. Originally insulin-like growth factors (IGFs) were implicated in this myofiber hyperplasia, as IGFs have been shown to stimulate myoblast proliferation as well as maintain fiber differentiation. Recently it has been reported that mice lacking a myostatin gene, a member of the TGFbeta superfamily, have enhanced skeletal mass resulting from increased muscle fiber number and size. Mutations in this gene have been found in double-muscled cattle, indicating that myostatin is an inhibitor of muscle growth. Myostatin is expressed early in gestation and then maintained to adulthood in certain muscles. Myostatin expression in bovine muscle is highest during gestation when muscle fibers are forming and some of the myogenic regulatory factors have elevated expression over the same period as myostatin. Molecular expression of the IGF axis does not differ between Belgian Blue and normal muscled cattle, and IGF-II mRNA is increased throughout formation of secondary fibers in both breeds. However, myostatin and MyoD expression in muscle differ between normal and hypertrophied muscle cattle breeds. This evidence strongly suggests that lack of myostatin is associated with an increase in fiber number which then results in a marked increase in potential muscle mass in double-muscled cattle.

Topics: Amino Acid Sequence; Animals; Cattle; Female; Fetus; Growth Substances; Hyperplasia; Insulin-Like Growth Factor II; Molecular Sequence Data; Muscle Development; Muscle, Skeletal; MyoD Protein; Myostatin; Point Mutation; Pregnancy; Sequence Alignment; Transforming Growth Factor beta

Asthma is a chronic inflammatory disease of the airways, with associated repair processes. Both inflammatory and repair processes appear to be strictly related, and can lead to several histopathological alterations of the bronchial mucosa, such as the shedding of epithelium and increased thickness of the basement membrane. The integrity as well as the alterations of the bronchial structure are the consequence of several biological events, such as cell proliferation and death, cell activation and inhibition, and extracellular matrix (ECM) production and degradation. These events are critically regulated by polypeptides called growth factors (GFs), which are able, functioning in an autocrine and paracrine fashion, to affect and modulate cell functions and ECM turnover. Although the importance of GFs has been widely demonstrated in other pulmonary conditions, such as lung fibrotic diseases, their possible involvement in the pathogenesis of inflammatory and postinflammatory processes in asthma is still not completely clear. The aim of the present review was to discuss the biological evidence concerning the role of several growth factors, such as transforming growth factor-beta (TGF-beta), epidermal growth factor (EGF), granulocyte/macrophage colony-stimulating factor (GM-CSF), platelet-derived growth factor (PDGF) and endothelin, in asthma and chronic bronchitis.

Topics: Asthma; Bronchi; Endothelial Growth Factors; Granulocyte-Macrophage Colony-Stimulating Factor; Growth Substances; Humans; Hyperplasia; Inflammation; Lung; Platelet-Derived Growth Factor; Respiratory System; Transforming Growth Factor beta

Agnogenic myeloid metaplasia (AMM) carries the worst prognosis among the chronic myeloproliferative disorders. Substantial bone marrow fibrosis, extramedullary hematopoiesis, anemia and hepatosplenomegaly are the characteristic features of the disease. AMM is currently incurable and the available treatment agents are mostly palliative and do not prolong life. Two pathogenetic processes are responsible for the impaired hematopoiesis and the clinical manifestations. The primary disease process is a clonal hematopoietic stem cell disorder which results in chronic myeloproliferation and atypical megakaryocytic hyperplasia. The secondary process of bone marrow fibrosis is the result of non-clonal fibroblastic proliferation and hyperactivity induced by growth factors abnormally shed from clonal megakaryocytes. Therefore, experimental treatment strategies may be directed towards either one or both of these disease processes. This report summarizes the current management options and new therapeutic endeavours.

Topics: Bone Marrow Examination; Clone Cells; Diagnosis, Differential; Disease Progression; Fibroblasts; Hematopoiesis; Humans; Hyperplasia; Leukemia; Megakaryocytes; Palliative Care; Platelet-Derived Growth Factor; Primary Myelofibrosis; Transforming Growth Factor beta

Hepatocyte growth factor (HGF) is the most potent mitogen for mature hepatocytes and seems to act as a hepatotropic factor that has not been purified over the past 30 years. HGF was first purified from rat platelets in 1986. HGF is a hetrodimer molecule composed of 69-kDa alpha-subunit and 34-beta-subunit. In 1989, cDNAs of both human and rat HGF were cloned and primary structure of HGF was determined. HGF is derived from preproprecursor of of 728 amino acids, which is proteolytically processed to form mature HGF. The alpha-chain contains four kringle domains and it has 38% homology with plasmin. HGF mRNA and HGF activity increase markedly in the liver of rats after various liver injuries such as hepatitis, ischemia, physical crush, and partial hepatectomy. Production of HGF in the liver occurs in Kupffer cells and sinusoidal endothelial cells, but not in parenchymal hepatocytes. HGF mRNA is also markedly increased even in the intact lung, kidney, and spleen after injuries of the liver. Therefore, HGF may act as a trigger for liver regeneration through two mechanisms: a paracrine mechanism and an endocrine mechanism. Moreover, HGF mRNA increases markedly in the kidney after various renal injuries, thus it suggests that HGF may act not only as a hepatotropic factor but also as a renotropic factor. HGF receptor with a Kd of 20 to 30 pM is widely distributed in various epithelial cells including hepatocytes. HGF receptor was recently identified as the product of c-met protooncogene, which encodes a 190-kDa transmembrane protein possessing tyrosine kinase domain. HGF has recently been shown to be a pleiotropic factor. HGF stimulates growth of various epithelial cells, including renal tubular cells (Mitogen). It is worth noting that HGF strongly enhances motility of epithelial cells (Motogen) and induces epithelial tubule formation (Morphogen), while it strongly inhibits growth of several tumor cells. All these findings indicate that HGF may have important roles in organogenesis, morphogenesis, carcinogenesis, as well as in organ regeneration.

Topics: Animals; Base Sequence; Cell Communication; Cell Division; Chromosome Mapping; Gene Expression Regulation; Growth Substances; Hepatocyte Growth Factor; Humans; Hyperplasia; Kidney; Liver Regeneration; Molecular Sequence Data; Molecular Structure; Platelet-Derived Growth Factor; Protein Precursors; Proto-Oncogene Proteins c-met; Rats; Receptors, Cell Surface; Sequence Homology, Nucleic Acid; Transforming Growth Factor beta

In order to investigate the biologic processes underlying and resulting from the megakaryocytic hyperplasia that characterizes idiopathic myelofibrosis (IMF), peripheral blood CD34+ cells isolated from patients with IMF, polycythemia vera (PV), and G-CSF-mobilized healthy volunteers were cultured in the presence of stem cell factor and thrombopoietin. IMF CD34+ cells generated 24-fold greater numbers of megakaryocytes (MKs) than normal CD34+ cells. IMF MKs were also shown to have a delayed pattern of apoptosis and to overexpress the antiapoptotic protein bcl-xL. MK hyperplasia in IMF is, therefore, likely a consequence of both the increased ability of IMF progenitor cells to generate MKs and a decreased rate of MK apoptosis. Media conditioned (CM) by CD61+ cells generated in vitro from CD34+ cells were then assayed for the levels of growth factors and proteases. Higher levels of transforming growth factor-beta (TGF-beta) and active matrix metalloproteinase-9 (MMP9) were observed in media conditioned with IMF CD61+ cells than normal or PV CD61+ cells. Both normal and IMF CD61+ cells produced similar levels of VEGF. MK-derived TGF-B and MMP-9, therefore, likely contribute to the development of many pathological epiphenomena associated with IMF.

Topics: Adult; Aged; Antigens, CD34; Apoptosis; bcl-X Protein; Cells, Cultured; Culture Media, Conditioned; Female; Granulocyte Colony-Stimulating Factor; Hematopoietic Stem Cell Mobilization; Hematopoietic Stem Cells; Humans; Hyperplasia; Integrin beta3; Male; Matrix Metalloproteinase 9; Megakaryocytes; Middle Aged; Polycythemia Vera; Primary Myelofibrosis; Thrombopoietin; Transforming Growth Factor beta; Vascular Endothelial Growth Factor A

HGF (Hepatocyte Growth Factor), TGFbeta1 (Transforming Growth Factor beta1) and IGF-I (Insulin Like Growth Factor I) are cytokines that are involved in the parathyroid tumors formation and growth. We tried to determine, if there are changes and relationships in the production of these cytokines by tumor cells of parathyroid tumors.. We determined concentrations of HGF, TGFbeta1 and IGF-I in serum from peripheral blood of 16 patients with parathyroid adenoma and of 8 patients with parathyroid secondary hyperplasia before and after parathyroidectomy. Results were compared with serum levels in healthy people.. Both preoperative and postoperative HGF serum levels in patients with parathyroid adenoma and secondary hyperplasia are significantly higher than in healthy people. Preoperative and postoperative serum levels of TGFbeta1 in parathyroid adenoma and postoperative TGFb1 serum levels in parathyroid secondary hyperplasia are higher, compared with those in the healthy population and in parathyroid secondary hyperplasia preoperatively. There are no significant differences of IGF-I serum levels among the all investigated groups of patients.. Changes in the growth factors production by parathyroid tumor cells are reflected by their concentrations in peripheral blood. The elevation of HGF serum levels in patients with parathyroid adenoma and hyperplasia can be explained by very high HGF production by tumor cells. Nevertheless, there is no decrease of HGF serum levels after the parathyroidectomy. That may be the result of the extratumoral production of this cytokine. Also TGFbeta1 and IGF-I serum levels indicate high possibility of the extratumoral production of these cytokines. Higher postoperative IGF-I serum levels (but not significantly) in parathyroid secondary hyperplasia are in accordance with its bone production.

Topics: Adenoma; Cytokines; Hepatocyte Growth Factor; Humans; Hyperplasia; Insulin-Like Growth Factor I; Parathyroid Glands; Parathyroid Neoplasms; Reference Values; Transforming Growth Factor beta

Topics: Animals; Cicatrix; Epidermal Growth Factor; Hyperplasia; Male; Rabbits; Stem Cells; Transforming Growth Factor beta; Vascular Endothelial Growth Factor A; Wound Healing

Benign prostate hyperplasia (BPH) is the most commonly seen disease among aging males. Transforming growth factor(TGF)-β-mediated epithelial-mesenchymal transition (EMT) and epithelial overproliferation might be central events in BPH etiology and pathophysiology. In the present study, long noncoding RNA MIR663AHG, miR-765, and FOXK1 formed a competing endogenous RNAs network, modulating TGF-β-mediated EMT and epithelial overproliferation in BPH-1 cells. miR-765 expression was downregulated in TGF-β-stimulated BPH-1 cells; miR-765 overexpression ameliorated TGF-β-mediated EMT and epithelial overproliferation in BPH-1 cells. MIR663AHG directly targeted miR-765 and negatively regulated miR-765; MIR663AHG knockdown also attenuated TGF-β-induced EMT and epithelial overproliferation in BPH-1 cells, whereas miR-765 inhibition attenuated MIR663AHG knockdown effects on TGF-β-stimulated BPH-1 cells. miR-765 directly targeted FOXK1 and negatively regulated FOXK1. FOXK1 knockdown attenuated TGF-β-induced EMT and epithelial overproliferation and promoted autophagy in BPH-1 cells, and partially attenuated miR-765 inhibition effects on TGF-β-stimulated BPH-1 cells. In conclusion, this study provides a MIR663AHG/miR-765/FOXK1 axis modulating TGF-β-induced epithelial proliferation and EMT, which might exert an underlying effect on BPH development and act as therapeutic targets for BPH treatment regimens.

Topics: Cell Movement; Cell Proliferation; Epithelial Cells; Epithelial-Mesenchymal Transition; Forkhead Transcription Factors; Humans; Hyperplasia; Male; MicroRNAs; Prostate; Prostatic Hyperplasia; Transforming Growth Factor beta; Transforming Growth Factor beta1

The primary underlying defect in cystic fibrosis (CF) is disrupted ion transport in epithelia throughout the body. It is unclear if symptoms such as airway hyperreactivity (AHR) and increased airway smooth muscle (ASM) volume in people with CF are due to inherent abnormalities in smooth muscle or are secondary to epithelial dysfunction. Transforming Growth Factor beta 1 (TGFβ) is an established genetic modifier of CF lung disease and a known driver of abnormal ASM function. Prior studies have demonstrated that CF mice develop greater AHR, goblet cell hyperplasia, and ASM hypertrophy after pulmonary TGFβ exposure. However, the mechanism driving these abnormalities in CF lung disease, specifically the contribution of CFTR loss in ASM, was unknown.. In this study, mice with smooth muscle-specific loss of CFTR function (Cftr. Cftr. These results demonstrate a direct smooth muscle contribution to CF airway obstruction mediated by TGFβ. Dysfunction in non-epithelial tissues should be considered in the development of CF therapeutics, including potential genetic therapies.

Topics: Animals; Asthma; Cystic Fibrosis; Cystic Fibrosis Transmembrane Conductance Regulator; Hyperplasia; Mice; Muscle, Smooth; Transforming Growth Factor beta

M-LP/Mpv17L is a protein that was initially identified during screening of age-dependently expressed genes in mice. We have recently demonstrated that M-LP/Mpv17L-knockout (M-LP/Mpv17L-KO) in human hepatoma cells leads to a reduction of cellular cyclic nucleotide phosphodiesterase (PDE) activity, and that in vitro-synthesized M-LP/Mpv17L possesses PDE activity. These findings suggest that M-LP/Mpv17L functions as an atypical PDE, even though it has none of the well-conserved catalytic region or other structural motifs characteristic of the PDE family. In this study, we found that M-LP/Mpv17L-KO mice developed β-cell hyperplasia and improved glucose tolerance. Deficiency of M-LP/Mpv17L in islets from KO mice at early postnatal stages or siRNA-mediated suppression of M-LP/Mpv17L in rat insulinoma cells led to marked upregulation of lymphoid enhancer binding factor 1 (Lef1) and transcription factor 7 (Tcf7), key nuclear effectors in the Wnt signaling pathway, and some of the factors essential for the development and maintenance of β-cells. Moreover, at the protein level, increases in the levels of phosphorylated β-catenin and glycogen synthase kinase-3β (GSK-3β) were observed, indicating activation of the Wnt and TGF-β signaling pathways. Taken together, these findings suggest that protein kinase A (PKA)-dependent phosphorylations of β-catenin and GSK-3β, the key mediators of the Wnt and/or TGF-β signaling pathways, are the most upstream events triggering β-cell hyperplasia and improved glucose tolerance caused by M-LP/Mpv17L deficiency.

Topics: Animals; Cell Proliferation; Glucose Intolerance; Hyperplasia; Insulin-Secreting Cells; Male; Membrane Proteins; Mice; Mice, Inbred C57BL; Mice, Knockout; Phosphorylation; Transforming Growth Factor beta; Wnt Signaling Pathway

To evaluate dynamic tissue changes after airway stenting (AS) with a newly designed metal brachytherapy stent (BS) loaded with radioactive. Forty-five normal New Zealand white rabbits were divided into 3 groups (group A: stent without seeds; group B: stent with 0.4 mCi active seeds; group C: stent with 0.8 mCi active seeds) and underwent AS under C-arm guidance. Then, five rabbits were killed from each group at 2, 4, and 8 weeks for further examination. Laboratory tests (including routine blood tests, liver function, kidney function, and electrolytes), gross observations, and tissue changes of Masson/hematoxylin-eosin staining, plus immunohistochemistry of α-SMA, NOX4, and TGF-β were performed at each time point.. All animals underwent AS successfully without procedure-related death, but one animal died at 6 weeks due to severe pulmonary infection in group C. Apart from a transient increase in white blood cells (P < 0.05) and a gradual increase in ROS levels (P < 0.05), other blood test items showed no significant changes (P > 0.05). The brachytherapy injury score increased with irradiation dose accumulation (P < 0.05), but tissue hyperplasia at the stent end in group C was less severe than that in groups A and B (P < 0.05). Airway lateral fibrosis was observed in all groups by histopathologic analysis; however, fibrosis in group C was more severe than that in groups A and B (P < 0.05).. The brachytherapy injury score increased with irradiation dose accumulation, while granulation tissue hyperplasia at the stent end was inhibited by

Topics: Animals; Brachytherapy; Eosine Yellowish-(YS); Fibrosis; Hematoxylin; Hyperplasia; Iodine Radioisotopes; Rabbits; Reactive Oxygen Species; Stents; Transforming Growth Factor beta

The online version contains supplementary material available at 10.1007/s00531-022-02219-9.

Topics: Accidents, Occupational; Adult; Animals; Anxiety; beta Catenin; Chromatography, High Pressure Liquid; Chronic Disease; Cities; Depression; Drugs, Chinese Herbal; Flavonoids; Heat Stroke; Hesperidin; Humans; Hyperplasia; Hypoxia-Inducible Factor 1, alpha Subunit; Interleukin-10; Interleukin-6; Kruppel-Like Factor 4; Macrophages, Alveolar; Medical Staff; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Occupational Health; Occupational Injuries; Occupational Stress; Occupations; PPAR gamma; Pulmonary Fibrosis; RNA, Messenger; Sanitation; Silicon Dioxide; Sinusitis; Stress, Psychological; Surveys and Questionnaires; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha; X-Box Binding Protein 1; Young Adult

Cigarette smoke (CS) is one of the major risk factors for chronic obstructive pulmonary disease (COPD) and increases the risk of lung cancer (LC). Anemoside B4 (B4) is the main bioactive ingredient in Pulsatilla chinensis (P. chinensis), a traditional medicinal herb for various diseases. It has a wide range of anti-inflammatory, anti-oxidation and anti-cancer activities. However, in recent years, there is no relevant literature report on the therapeutic effect of B4 on COPD, and the anti-inflammatory and inhibitory effects of anemoside B4 on basal cell hyperplasia in CS-induced COPD have not been clearly established.. In the present study, we investigated whether anemoside B4 could alleviate CS or cigarette smoke extract (CSE) induced inflammation of COPD and further prevent basal cell hyperplasia, hoping to find its possible mechanism.. In this study, a COPD mouse model was established in C57BL mice by CS exposure 3 months. Bronchial pathology and basal cell hyperplasia were observed by HE staining and immunostaining. The contents of glutathione peroxidase catalase (GSH-PX), malondialdehyde (MDA) and superoxide dismutase (MPO) were determined by GSH-PX, MDA and SOD assay kits, respectively. 16HBE cells were cultured with 5% CSE with or without treatment with B4 (1, 10, 100 μM) or DEX (20 μM) in vitro. Cell viability was assessed by a cell counting kit 8 (CCK-8). Reactive oxygen species (ROS) generation was tested by DCFH-DA. Moreover, anti-inflammatory mechanism of anemoside B4 was further determined by pro-inflammatory cytokines production using RT-PCR. Protein expression levels of MAPK/AP-1/TGF-β signaling pathway were measured by western blot.. Anemoside B4 improved the lung function of mice, relieved lung inflammation and reduced the MDA, MPO and GSH-Px in the plasma. At the same time, B4 repressed the oxidative stress response and played a role in balancing the levels of protease and anti-protease. During the process of bronchial basal cell hyperplasia, B4 alleviated the degree of cell hyperplasia, and prevented further deterioration of hyperplasia through increased P53 and inhibited FHIT protein. In addition, B4 reduced ROS levels in human bronchial epithelial cells stimulated by CSE in vitro study. Meanwhile, B4 treatment also significantly attenuated increased IL-1β, TGF-β, IL-8 and TNF-α from CSE treated human bronchial epithelial cells. The expression of p-P38, AP-1(c-fos, and c-Jun), TGF-β proteins in MAPK/AP-1/TGF-β signaling pathway were decreased and the signal cascade reaction was blocked.. Anemoside B4 protects against CS-induced COPD. These findings indicated that B4 may have therapeutic potential for the prevention and treatment of COPD.

Topics: Animals; Anti-Inflammatory Agents; Catalase; Cigarette Smoking; Glutathione Peroxidase; Humans; Hyperplasia; Inflammation; Interleukin-8; Malondialdehyde; Mice; Mice, Inbred C57BL; Pulmonary Disease, Chronic Obstructive; Reactive Oxygen Species; Saponins; Superoxide Dismutase; Transcription Factor AP-1; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha; Tumor Suppressor Protein p53

Epithelial-mesenchymal transition (EMT) of the retinal pigment epithelium (RPE) is associated with several blinding retinal diseases. Using proteomics and phosphoproteomics studies of human induced pluripotent stem cell-derived RPE monolayers with induced EMT, we capture kinase/phosphatase signaling cascades 1 h and 12 h after induction to better understand the pathways mediating RPE EMT. Induction by co-treatment with transforming growth factor β and tumor necrosis factor alpha (TGNF) or enzymatic dissociation perturbs signaling in many of the same pathways, with striking similarity in the respective phosphoproteomes at 1 h. Liver hyperplasia and hepatocyte growth factor (HGF)-MET signaling exhibit the highest overall enrichment. We also observe that HGF and epidermal growth factor signaling, two cooperative pathways inhibited by EMT induction, regulate the RPE transcriptional profile.

Topics: Cell Line; Epithelial Cells; Epithelial-Mesenchymal Transition; ErbB Receptors; Hepatocyte Growth Factor; Humans; Hyperplasia; Induced Pluripotent Stem Cells; Liver; Phosphorylation; Proteome; Proteomics; Proto-Oncogene Proteins c-met; Retinal Pigment Epithelium; Signal Transduction; Transcriptome; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

Transforming growth factor β (TGFβ) signaling plays critical roles in reproductive development and function. TGFβ ligands signal through the TGFβ receptor type 2 (TGFBR2)/TGFBR1 complex. As TGFBR2 and TGFBR1 form a signaling complex upon ligand stimulation, they are expected to be equally important for propagating TGFβ signaling that elicits cellular responses. However, several genetic studies challenge this concept and indicate that disruption of TGFBR2 or TGFBR1 may lead to contrasting phenotypic outcomes. We have shown that conditional deletion of Tgfbr1 using anti-Mullerian hormone receptor type 2 (Amhr2)-Cre causes oviductal and myometrial defects. To determine the functional requirement of TGFBR2 in the female reproductive tract and the potential phenotypic divergence/similarity resulting from conditional ablation of either receptor, we generated mice harboring Tgfbr2 deletion using the same Cre driver that was previously employed to target Tgfbr1. Herein, we found that conditional deletion of Tgfbr2 led to a similar phenotype to that of Tgfbr1 deletion in the female reproductive tract. Furthermore, genetic removal of Tgfbr1 in the Tgfbr2-deleted uterus had minimal impact on the phenotype of Tgfbr2 conditional knockout mice. In summary, our results reveal the functional similarity between TGFBR2 and TGFBR1 in maintaining the structural integrity of the female reproductive tract.

Topics: Animals; Endometrium; Fallopian Tubes; Female; Gene Knockout Techniques; Genitalia, Female; Hyperplasia; Mice; Mice, Inbred C57BL; Myometrium; Phenotype; Receptor, Transforming Growth Factor-beta Type I; Receptor, Transforming Growth Factor-beta Type II; Signal Transduction; Transforming Growth Factor beta

Cystic fibrosis (CF) is a lethal genetic disease characterized by progressive lung damage and airway obstruction. The majority of patients demonstrate airway hyperresponsiveness (AHR), which is associated with more rapid lung function decline. Recent studies in the neonatal CF pig demonstrated airway smooth muscle (ASM) dysfunction. These findings, combined with observed CF transmembrane conductance regulator (CFTR) expression in ASM, suggest that a fundamental defect in ASM function contributes to lung function decline in CF. One established driver of AHR and ASM dysfunction is transforming growth factor (TGF) β1, a genetic modifier of CF lung disease. Prior studies demonstrated that TGFβ exposure in CF mice drives features of CF lung disease, including goblet cell hyperplasia and abnormal lung mechanics. CF mice displayed aberrant responses to pulmonary TGFβ, with elevated PI3K signaling and greater increases in lung resistance compared with controls. Here, we show that TGFβ drives abnormalities in CF ASM structure and function through PI3K signaling that is enhanced in CFTR-deficient lungs. CF and non-CF mice were exposed intratracheally to an adenoviral vector containing the TGFβ1 cDNA, empty vector, or PBS only. We assessed methacholine-induced AHR, bronchodilator response, and ASM area in control and CF mice. Notably, CF mice demonstrated enhanced AHR and bronchodilator response with greater ASM area increases compared with non-CF mice. Furthermore, therapeutic inhibition of PI3K signaling mitigated the TGFβ-induced AHR and goblet cell hyperplasia in CF mice. These results highlight a latent AHR phenotype in CFTR deficiency that is enhanced through TGFβ-induced PI3K signaling.

Topics: Adrenergic beta-Agonists; Albuterol; Animals; Bronchoconstriction; Cystic Fibrosis; Goblet Cells; Hyperplasia; Lung; Mice, Inbred C57BL; Muscle, Smooth; Phosphatidylinositol 3-Kinases; Phosphoinositide-3 Kinase Inhibitors; Respiratory Hypersensitivity; Signal Transduction; Transforming Growth Factor beta

Allergic asthma (AA) is characterized as a Th2-biased airway inflammation that can develop lung inflammation and remodeling of the respiratory tract.

Topics: Animals; Asthma; Cells, Cultured; Disease Models, Animal; Disease Resistance; Goblet Cells; Humans; Hyperplasia; Hypersensitivity; Interleukin-6; Mice; Mice, Knockout; Pneumonia; Pneumonia, Pneumococcal; Respiratory Mucosa; RNA, Small Interfering; Signal Transduction; Streptococcus pneumoniae; Tight Junctions; Transforming Growth Factor beta

Growth differentiation factor-15 (GDF-15) is a member of TGF-β superfamily. Among hematopoietic cells, this factor is mainly produced by erythroid series and is recently considered a biomarker of ineffective erythropoiesis (IE). Whether IE induces enhanced GDF-15 expression or is prompted by it, has remained elusive. In this study we investigated how high levels of GDF-15 contribute to IE-associated erythroid dysplasia. We assessed mRNA levels of GDF-15 during erythroid maturation as well as in patients with IE using qRT-PCR. Later, the erythroid colony-forming capacity of GDF-15-treated hematopoietic stem cells (HSCs) was evaluated by CFC assay. Any effect of elevated levels of GDF-15 on erythroid maturation was ultimately examined by expression analysis of erythroid-associated transcription factors and flow cytometry analysis of CD235a expression. GDF-15 mRNA expression increased during erythroid differentiation and also in β-thalassemia and MDS patients which was directly correlated with erythropoiesis severity. Treating the cells with high GDF-15 concentration (50 ng/ml) resulted in an approximate 30% decline in the capacity of erythroid colony formation of HSCs and CD235a positive cells. Additionally, erythroid-specific transcription factors showed significant down-regulation in the early stages of erythroid differentiation. According to the expression level of GDF-15 and the role it plays in the erythroid system, high-levels of this factor could be an auto-modulatory mechanism to control the excessive production of erythroid cells.

Topics: beta-Thalassemia; Case-Control Studies; Cell Differentiation; Erythroid Precursor Cells; Erythropoiesis; Growth Differentiation Factor 15; Hematopoietic Stem Cells; Humans; Hyperplasia; Stem Cell Factor; Transforming Growth Factor beta

To investigate the mechanism of bladder nerve hyperplasia and fibrosis in the patients with ketamine-associated cystitis (KC).. Sixteen patients with severe KC, six patients with mild KC, and five patients with localized invasive bladder cancer served as control patients. Bladder mucosa specimens were taken during the operations, and the specimens were stained for nerve growth factor (NGF) and S-100 to evaluated nerve hyperplasia. The quantitative Western blot analysis was performed for NGF, brain-derived neurotrophic factor (BDNF), growth-associated protein 43 (GAP-43), tropomyosin receptor kinase A and B (TrkA and TrkB), transforming growth factor-β (TGF-β), phosphorylated extracellular signal-regulated kinases (p-ERK), protein kinase B (p-Akt), and glycogen synthase kinase 3β (p-GSK-3β).. The results demonstrated diffuse NGF expression in KC bladder epithelium, lamina propria, and muscle. The GAP-43, NGF, BDNF, TrkA, TGF-β, p-ERK, P-AKT, and p-GSK-3β expression in the bladder mucosa specimens of patients with severe KC was significantly higher than in patients with mild KC and control patients. Expression of neurotrophins was significantly correlated with bladder capacity and pain. NGF and BDNF expression were significantly higher in the KC bladder specimens with strongly positive S-100 staining. TGF-β expression in the bladder specimens was significantly correlated with neurotrophins, p-ERK, P-AKT, and p-GSK-3β levels.. Our findings indicate upregulation of neurotrophins, TGF-β, and activation of the cell proliferation kinases plays an important role in nerve hyperplasia and fibrosis mechanisms in severe KC bladders. The neurotrophins and TGF-β interact as cause and effect, leading to bladder hypersensitivity and fibrosis in severe KC.

Topics: Adult; Aged; Brain-Derived Neurotrophic Factor; Cystitis; Excitatory Amino Acid Antagonists; Female; Fibrosis; Humans; Hyperplasia; Ketamine; Male; Middle Aged; Mucous Membrane; Nerve Growth Factors; Transforming Growth Factor beta; Up-Regulation; Urinary Bladder; Urinary Bladder Neoplasms; Young Adult

Asthma is a common obstructive airway disease characterized by inflammation and remodeling with a progressive decline in lung function. Fangxiao Formula (FXF) is an herbal medicine that has achieved significant clinical benefits toward asthma patients, but the relevant mechanism has not yet been clarified. The aim of this study was to determine the inhibitory effects of FXF on airway inflammation and remodeling, and investigate the activities of TGF‑β/Smads signaling pathway in the rat asthma model. Rats were sensitized by ovalbumin (OVA) for six weeks to establish the asthma experimental model. OVA-challenged animals were randomly divided into 5 groups and received different concentrations of FXF or dexamethasone. The animals in blank control group received saline only. Lung tissues were collected and analyzed for determining the inflammatory cells infiltration, HE and PAS staining, airway wall thickness and collagen deposition. The productions of inflammatory cytokine productions were analyzed by ELISA in the bronchoalveolar lavage (BAL) fluid. Immunohistochemical analysis was performed to measure the expression of α-SMA and PCNA in lung tissue after the treatment of FXF. The levels of TGF-β were assessed by both immunohistology and western blotting, and the expression of p-Smad2/3 proteins were determined by western blotting analysis. Our results indicated that FXF attenuated the infiltration of inflammatory cells, decreased the production of Th2 cytokines and simultaneously increased the levels of Th1 cytokine in the asthma rat model. In addition, FXF reduced allergen-induced increased airway wall thickness, goblet cell hyperplasia and collagen deposition. Furthermore, the expression levels of TGF-β and p-Smad3 were obviously reduced after the treatment of FXF. These results indicate that FXF alleviates airway inflammation and remodeling by restoring the balance of Th1/Th2 cytokines and the TGF-β/Smad-3 pathway, therefor providing potential therapeutic approach for asthmatic patients.

Topics: Airway Remodeling; Animals; Asthma; Bronchoalveolar Lavage Fluid; Collagen; Cytokines; Disease Models, Animal; Drugs, Chinese Herbal; Hyperplasia; Inflammation; Lung; Male; Proliferating Cell Nuclear Antigen; Rats, Sprague-Dawley; Signal Transduction; Smad3 Protein; Th1 Cells; Th2 Cells; Transforming Growth Factor beta

Laparoscopic resection of the pancreatic body and tail is the predominant methodology to remove lesions in these locations; its safety and surgical planning are relatively mature, but it remains a complex and high-precision surgical operation, requiring abundant experience and skills in laparoscopic surgery, with a 10% rate of complications.. To verify the feasibility and safety, as well as to examine the complications of endoscopic pancreatectomy and healing mechanisms of pancreatic wounds after endoscopic resection.. Transgastric endoscopic resections of varying sizes of pancreases were performed in 15 healthy Bama miniature pigs. The technical success rate, the incidence of serious complications, and the survival of the animals were studied. The healing of the wounds was evaluated by sacrificing the animals at various time points. Finally, the expression of transforming growth factor-β1 and Smad3/Smad7 in the surgical site was examined by immunohistochemistry to explore the role of these factors in wound healing of the pancreas.. Partial and total resections were successfully performed in two groups of animals, respectively. The technical success rate and the survival rate of the pigs were both 100%. We obtained 12 pancreatic tissue samples by endoscopic resection. The pancreatic wounds were closed with metal clips in one group and the wounds healed well by forming scars. There was a small amount of pancreatic leakage in the other group, but it can be fully encapsulated. The level of transforming growth factor-β1 (TGF-β1) in the wounds increased during the inflammatory and fibrous hyperplasia phases, and decreased in the scar phase. The expression of Smad3 paralleled that of TGF-β1, while the expression of Smad7 had an inverse relationship with the expression of TGF-β1.. Purely transgastric endoscopic resection of the pancreas is a safe, effective, and feasible procedure, but the incidence of pancreatic leakage in total pancreatic tail resection is high. The expression of TGF-β1 and Smad3/Samd7 is related to the progression of pancreatic wound healing.

Topics: Animals; Disease Models, Animal; Endoscopy; Feasibility Studies; Humans; Hyperplasia; Incidence; Pancreas; Pancreatectomy; Postoperative Complications; Smad3 Protein; Smad7 Protein; Survival Rate; Swine; Swine, Miniature; Transforming Growth Factor beta; Wound Healing

α7-nAChR is a nicotinic acetylcholine receptor [specifically expressed on hepatic stellate cells (HSCs), Kupffer cells, and cholangiocytes] that regulates inflammation and apoptosis in the liver. Thus, targeting α7-nAChR may be therapeutic in biliary diseases. Bile duct ligation (BDL) was performed on wild-type (WT) and α7-nAChR-/- mice. We first evaluated the expression of α7-nAChR by immunohistochemistry (IHC) in liver sections. IHC was also performed to assess intrahepatic bile duct mass (IBDM), and Sirius Red staining was performed to quantify the amount of collagen deposition. Immunofluorescence was performed to assess colocalization of α7-nAChR with bile ducts (costained with CK-19) and HSCs (costained with desmin). The mRNA expression of α7-nAChR, Ki-67/PCNA (proliferation), fibrosis genes (TGF-β1, fibronectin-1, Col1α1, and α-SMA), and inflammatory markers (IL-6, IL-1β, and TNF-α) was measured by real-time PCR. Biliary TGF-β1 and hepatic CD68 (Kupffer cell marker) expression was assessed using IHC. α7-nAChR immunoreactivity was observed in both bile ducts and HSCs and increased following BDL. α7-nAChR-/- BDL mice exhibited decreased (i) bile duct mass, liver fibrosis, and inflammation, and (ii) immunoreactivity of TGF-β1 as well as expression of fibrosis genes compared to WT BDL mice. α7-nAChR activation triggers biliary proliferation and liver fibrosis and may be a therapeutic target in managing extrahepatic biliary obstruction.

Topics: alpha7 Nicotinic Acetylcholine Receptor; Animals; Bile Ducts; Cell Line, Tumor; Cholestasis, Extrahepatic; Cytokines; Humans; Hyperplasia; Ki-67 Antigen; Liver Cirrhosis; Male; Mice; Mice, Inbred C57BL; Proliferating Cell Nuclear Antigen; Transforming Growth Factor beta

The phenomena involved in regression of arterial myointimal hyperplasia have not been analyzed in detail.. In 24 Lewis rats, a 1-cm-long venous graft, obtained from syngenic Lewis rats, was implanted in the infrarenal aorta. After 4 wk, the grafts were removed and analyzed using scanning electron microscopy and histochemistry. The grafts showed evidence of myointimal hyperplasia; 16 of these explanted grafts were reimplanted in the vein circulation of syngenic Lewis rats. These grafts were harvested 2 wk (8 animals) and 8 wk (8 animals) later, showing complete regression of myointimal hyperplasia.. Regression of experimental myointimal hyperplasia was correlated with the simultaneous and complementary action of Transforming Growth Factor beta and Tumor Necrosis Factor alfa. Inflammatory cytokines (IL1, IL2, and IL6) inhibit Tumor Necrosis Factor alfa-induced apoptosis.. Regression of myointimal hyperplasia is an active process, which implies the action of several inhibitory factors. The analysis of these phenomena can lead to new therapeutic approaches to prevent myointimal hyperplasia progression.

Topics: Animals; Aorta, Abdominal; Apoptosis; Disease Models, Animal; Fibroblast Growth Factor 2; Hyperplasia; Interleukin-1; Interleukin-2; Interleukin-6; Male; Microscopy, Electron, Scanning; Muscle, Smooth, Vascular; Platelet-Derived Growth Factor; Rats; Rats, Inbred Lew; Replantation; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha; Tunica Intima; Veins

Benign prostatic hyperplasia (BPH) is the overgrowth of prostate tissues with high prevalence in older men. BPH pathogenesis is not completely understood, but it is believed to be a result of de novo overgrowth of prostatic stroma. In this study, we show that aberrant activation of transforming growth factor-β (TGF-β) mobilizes mesenchymal/stromal stem cells (MSCs) in circulating blood, which are recruited for the prostatic stromal hyperplasia. Elevated levels of active TGF-β were observed in both a phenylephrine-induced prostatic hyperplasia mouse model and human BPH tissues. Nestin lineage tracing revealed that 39.6% ± 6.3% of fibroblasts and 73.3% ± 4.2% smooth muscle cells were derived from nestin

Topics: Animals; Antibodies, Neutralizing; Case-Control Studies; Cell Lineage; Cell Movement; Cell Proliferation; Cells, Cultured; Disease Models, Animal; Fibroblasts; Humans; Hyperplasia; Male; Mesenchymal Stem Cells; Mice, Inbred C57BL; Mice, Transgenic; Nestin; Parabiosis; Phenotype; Prostate; Prostatic Hyperplasia; Receptor, Transforming Growth Factor-beta Type II; Signal Transduction; Transforming Growth Factor beta

WISP2 is a novel adipokine, most highly expressed in the adipose tissue and primarily in undifferentiated mesenchymal cells. As a secreted protein, it is an autocrine/paracrine activator of canonical WNT signaling and, as an intracellular protein, it helps to maintain precursor cells undifferentiated. To examine effects of increased WISP2 in vivo, we generated an aP2-WISP2 transgenic (Tg) mouse. These mice had increased serum levels of WISP2, increased lean body mass and whole body energy expenditure, hyperplastic brown/white adipose tissues and larger hyperplastic hearts. Obese Tg mice remained insulin sensitive, had increased glucose uptake by adipose cells and skeletal muscle in vivo and ex vivo, increased GLUT4, increased ChREBP and markers of adipose tissue lipogenesis. Serum levels of the novel fatty acid esters of hydroxy fatty acids (FAHFAs) were increased and transplantation of Tg adipose tissue improved glucose tolerance in recipient mice supporting a role of secreted FAHFAs. The growth-promoting effect of WISP2 was shown by increased BrdU incorporation in vivo and Tg serum increased mesenchymal precursor cell proliferation in vitro. In contrast to conventional canonical WNT ligands, WISP2 expression was inhibited by BMP4 thereby allowing normal induction of adipogenesis. WISP2 is a novel secreted regulator of mesenchymal tissue cellularity.

Topics: Adipose Tissue; Adipose Tissue, Brown; Adipose Tissue, White; Animals; Autocrine Communication; Biomarkers; Body Composition; Body Weight; Bone Morphogenetic Protein 4; Cell Count; Cell Proliferation; Cell Size; Energy Metabolism; Gene Expression; Genotype; Glucose; Glucose Tolerance Test; Glucose Transporter Type 4; Hyperplasia; Insulin; Insulin Resistance; Intracellular Signaling Peptides and Proteins; Lipogenesis; Male; Mesenchymal Stem Cells; Mice; Mice, Transgenic; Myocardium; Transforming Growth Factor beta

Upregulation of WISP1 has been demonstrated in lung remodeling. Moreover, it has been recently found that some signaling components of WNT pathway can activate GSK3β signaling to mediate remodeling of airway smooth muscle (ASM) in asthma. Therefore, we hypothesized that WISP1, a signaling molecule downstream of the WNT signaling pathway, is involved in PI3K/GSK3β signaling to mediate ASM remodeling in asthma. Our results showed that WISP1 depletion partly suppressed OVA-induced ASM hypertrophy in vivo. In vitro, WISP1 could induce hBSMC hypertrophy and proliferation, accompanied by upregulation of levels of PI3K, p-Akt, p-GSK3β, and its own expression. TGF-β treatment could increase expression of PI3K, p-Akt, p-GSK3β, and WISP1. SH-5 treatment could partly suppress TGF-β-induced hypertrophy and proliferation of hBSMC, and depress expression of p-GSK3β and WISP1. In conclusion, WISP1 may be a potential inducer of ASM proliferation and hypertrophy in asthma. The pro-remodeling effect of WISP1 is likely due to be involved in PI3K-GSK3β-dependent noncanonical TGF-β signaling.

Topics: Airway Remodeling; Animals; Asthma; Bronchi; CCN Intercellular Signaling Proteins; Cell Line; Cell Movement; Cell Proliferation; Glycogen Synthase Kinase 3 beta; Humans; Hyperplasia; Hypertrophy; Male; Myocytes, Smooth Muscle; Ovalbumin; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-akt; Rats; Rats, Sprague-Dawley; Signal Transduction; Transforming Growth Factor beta

The production of growth factors from several experimental arterial conduits was determined.. We implanted 105 experimental arterial grafts that were 1 cm long in the abdominal aorta of Lewis rats (average weight, 250 g). Five different types of grafts were analyzed: arterial isografts, vein grafts, arterial allografts, and polytetrafluoroethylene (PTFE) grafts with normal or decreased compliance. Animals were killed humanely 4 weeks after surgery and the production of platelet-derived growth factor (PDGF), basic fibroblast growth factor (bFGF), transforming growth factor-β, tumor necrosis factor-α, and interleukin-1 was analyzed.. Myointimal hyperplasia (MH) was evident in vein grafts, arterial allografts, and PTFE grafts, but not in arterial isografts. Growth factor production was increased for grafts prone to develop MH like vein, PTFE grafts, and arterial allografts. PDGF and bFGF were increased significantly for PTFE and vein grafts, but not for arterial allografts. The importance of bFGF and PGDF was confirmed by the capability of antibody to PDGF and to bFGF to reduce the mitogenic activity of smooth muscle cells, in vivo and in vitro, for PTFE and vein grafts, but not for arterial allografts, in which a predominant role was played by interleukin-1 and tumor necrosis factor-α.. Agents able to neutralize this increased production of growth factors, either directly or by competition with their receptors, can prevent MH formation.

Topics: Allografts; Animals; Aorta, Abdominal; Arteries; Blood Vessel Prosthesis; Blood Vessel Prosthesis Implantation; Cell Proliferation; Cells, Cultured; Culture Media, Conditioned; Fibroblast Growth Factor 2; Hyperplasia; Intercellular Signaling Peptides and Proteins; Interleukin-1; Isografts; Models, Animal; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; Neointima; Platelet-Derived Growth Factor; Polytetrafluoroethylene; Prosthesis Design; Rats, Inbred Lew; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha; Veins

Mps One Binder Kinase Activator (MOB)1A/1B are core components of the Hippo pathway that coactivate large tumor suppressor homolog (LATS) kinases. Mob1a/1b double deficiency in mouse liver (LMob1DKO) results in hyperplasia of oval cells and immature cholangiocytes accompanied by inflammatory cell infiltration and fibrosis. More than half of mutant mice die within 3 wk of birth. All survivors eventually develop liver cancers, particularly combined hepatocellular and cholangiocarcinomas (cHC-CCs) and intrahepatic cholangiocellular carcinomas (ICCs), and die by age 60 wk. Because this phenotype is the most severe among mutant mice lacking a Hippo signaling component, MOB1A/1B constitute the critical hub of Hippo signaling in mammalian liver. LMob1DKO liver cells show hyperproliferation, increased cell saturation density, hepatocyte dedifferentiation, enhanced epithelial-mesenchymal transition and cell migration, and elevated transforming growth factor beta(TGF-β)2/3 production. These changes are strongly dependent on Yes-Associated Protein-1 (Yap1) and partially dependent on PDZ-binding motif (Taz) and Tgfbr2, but independent of connective tissue growth factor (Ctgf). In human liver cancers, YAP1 activation is frequent in cHC-CCs and ICCs and correlates with SMAD family member 2 activation. Drug screening revealed that antiparasitic macrocyclic lactones inhibit YAP1 activation in vitro and in vivo. Targeting YAP1/TAZ with these drugs in combination with inhibition of the TGF-β pathway may be effective treatment for cHC-CCs and ICCs.

Topics: Acyltransferases; Adaptor Proteins, Signal Transducing; Animals; Bile Duct Neoplasms; Carcinogenesis; Cell Line, Tumor; Cholangiocarcinoma; Connective Tissue Growth Factor; Epithelial-Mesenchymal Transition; Genes, Tumor Suppressor; Humans; Hyperplasia; Intracellular Signaling Peptides and Proteins; Liver; Liver Neoplasms; Mice; Mice, Knockout; Mice, Nude; Phosphoproteins; Protein Kinases; Protein Serine-Threonine Kinases; Receptor, Transforming Growth Factor-beta Type II; Receptors, Transforming Growth Factor beta; Signal Transduction; Transcription Factors; Transforming Growth Factor beta; Xenograft Model Antitumor Assays; YAP-Signaling Proteins

The traditional Chinese medicine Shensong Yangxin (SSYX) can improve the clinical symptoms of arrhythmia in an integrated manner. This study aimed to investigate the electrophysiological effect of SSYX on the hearts of myocardial-infarcted rabbits and further explore the mechanism by which SSYX alleviates myocardial fibrosis. Myocardial infarction (MI) was established in rabbits by ligation of the left circumflex coronary. The rabbits were treated with SSYX (0.5 g/kg/d) or saline for 8 weeks by oral administration. Microelectrode array (MEA) technology was used in vivo for extracellular electrophysiological recordings of the infarct border zone. Masson's trichrome staining was used to observe myocardial fibrosis. Western blotting was performed to evaluate the protein expression levels of collagen I (COL I) and collagen III (COL III). Quantitative real-time polymerase chain reaction (real-time PCR) was performed to evaluate the TGF-β1 and MMP-2 mRNA expression levels. The results showed that the total activation time (TAT) and the dispersion of TAT were significantly increased and the excitation propagation markedly disordered after MI. SSYX could significantly decrease TAT and the dispersion of TAT, and significantly ameliorate the chaotic spread pattern of excitation. Furthermore, SSYX treatment could significantly decrease COL I and COL III protein levels and down-regulate TGF-β1 and MMP-2 mRNA expression levels in MI rabbits. It was concluded that SSYX may ameliorate cardiac electrophysiological abnormalities in infarcted hearts by decreasing the protein levels of COL I and COL III, down-regulating the mRNA expression levels of TGF-β1 and MMP2, and thereby reducing adverse cardiac remodeling.

Topics: Animals; Collagen Type I; Collagen Type III; Drugs, Chinese Herbal; Female; Heart Rate; Hyperplasia; Male; Matrix Metalloproteinase 2; Myocardial Infarction; Myocardium; Rabbits; Transforming Growth Factor beta

Fibrosis of lung tissue is a disease where a chronic inflammatory process determines a pathological remodelling of lung parenchyma. The animal model obtained by intra-tracheal administration of bleomycin in C57BL/6 mice is one of the most validated murine model. Bleomycin stimulates oxidative stress and the production of pro-inflammatory mediators. Histamine H4R have recently been implicated in inflammation and immune diseases. This study was focused to investigate the effects of H4R ligands in the modulation of inflammation and in the reduction of lung fibrosis in C57BL/6 mice treated with bleomycin. C57BL/6 mice were treated with vehicle, JNJ7777120 (JNJ, selective H4R antagonist) or ST-1006 (partial H4R agonist), ST-994 (H4R neutral antagonist) and ST-1012 (inverse H4R agonist) at equimolar doses, released by micro-osmotic pumps for 21days. Airway resistance to inflation was assayed and lung samples were processed to measure malondialdehyde (TBARS); 8-hydroxy-2'-deoxyguanosine (8OHdG); myeloperoxidase (MPO); COX-2 expression and activity as markers of oxidative stress and inflammation. Fibrosis and airway remodelling were evaluated throughout transforming growth factor-β (TGF-β), percentage of positive Goblet cells, smooth muscle layer thickness determination. Our results indicated that JNJ, ST-994 and ST-1012 decreased inflammation and oxidative stress markers, i.e. the number of infiltrating leukocytes evaluated as lung tissue MPO, COX-2 expression and activity, TBARS and 8OHdG production. They also reduced the level of TGF-β, a pro-fibrotic cytokine, collagen deposition, thickness of smooth muscle layer, Goblet cells hyperplasia; resulting in a decrease of airway functional impairment. The results here reported clearly demonstrated that H4R ligands have a beneficial effect in a model of lung fibrosis in the mouse, thus indicating that H4R antagonists or inverse agonists could be a novel therapeutic strategy for lung inflammatory diseases.

Topics: Animals; Anti-Inflammatory Agents; Biomarkers; Bleomycin; Collagen; Cytoprotection; Disease Models, Animal; Drug Partial Agonism; Goblet Cells; Histamine Antagonists; Hyperplasia; Indoles; Inflammation Mediators; Ligands; Lung; Male; Mice, Inbred C57BL; Oxidative Stress; Piperazines; Pneumonia; Pulmonary Fibrosis; Pyrimidines; Receptors, Histamine H4; Signal Transduction; Transforming Growth Factor beta

Although indoleamine 2,3-dioxygenase (IDO)-mediated immune suppression of mesenchymal stem cells (MSCs) has been revealed in septic and tumor microenvironments, the role of IDO in suppressing allergic airway inflammation by MSCs is not well documented. We evaluated the effects of adipose-derived stem cells (ASCs) on allergic inflammation in IDO-knockout (KO) asthmatic mice or asthmatic mice treated with ASCs derived from IDO-KO mice.. ASCs were injected intravenously in wild-type (WT) and IDO-KO asthmatic mice. Furthermore, asthmatic mice were injected with ASCs derived from IDO-KO mice. We investigated the immunomodulatory effects of ASCs between WT and IDO-KO mice or IDO-KO ASCs in asthmatic mice. In asthmatic mice, ASCs significantly reduced airway hyperresponsiveness, the number of total inflammatory cells and eosinophils in bronchoalveolar lavage fluid (BALF), eosinophilic inflammation, goblet hyperplasia, and serum concentrations of total and allergen-specific IgE and IgG1. ASCs significantly inhibited Th2 cytokines, such as interleukin (IL)-4, IL-5, and IL-13, and enhanced Th1 cytokine (interferon-γ) and regulatory cytokines (IL-10, TGF-β) in BALF and lung draining lymph nodes (LLNs). ASCs led to significant increases in regulatory T-cells (Tregs) and IL-10+ T cell populations in LLNs. However, the immunosuppressive effects of ASCs did not significantly differ between WT and IDO-KO mice. Moreover, ASCs derived from IDO-KO mice showed immunosuppressive effects in allergic airway inflammation.. IDO did not play a pivotal role in the suppression of allergic airway inflammation through ASCs, suggesting that it is not the major regulator responsible for suppressing allergic airway inflammation.

Topics: Adipose Tissue; Animals; Asthma; Bronchoalveolar Lavage Fluid; Cell- and Tissue-Based Therapy; Cells, Cultured; Eosinophils; Female; Goblet Cells; Hyperplasia; Immunoglobulin E; Immunoglobulin G; Indoleamine-Pyrrole 2,3,-Dioxygenase; Inflammation; Interferon-gamma; Interleukin-10; Interleukin-13; Interleukin-4; Interleukin-5; Lymphocyte Count; Mesenchymal Stem Cell Transplantation; Mesenchymal Stem Cells; Methacholine Chloride; Mice; Mice, Inbred C57BL; Mice, Knockout; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

To determine the mechanism underlying hypertrophic synovium in rheumatoid arthritis (RA).. We examined micromass cultures of fibroblast-like synoviocytes (FLSs) stimulated with tumor necrosis factor α (TNFα), platelet-derived growth factor (PDGF), and/or transforming growth factor β (TGFβ). The hypertrophic architecture of the micromasses, expression of phosphoinositide 3 kinase (PI3K) isoforms, and persistent activation of PI3K-Akt pathways were investigated. FLSs transfected with siRNA were also examined in the micromass cultures.. The combination of TNFα, PDGF, and TGFβ (TPT condition) induced obvious hypertrophic architecture of the intimal lining layer in FLSs in micromass cultures, and was accompanied by upregulated expression of matrix metalloproteinase-3 (MMP3), Cadherin-11, and PI3Kδ. In monolayer FLSs, the TPT condition enhanced the expression of PI3Kδ and persistent activation of the PI3K-Akt pathway. Knockdown of PI3Kδ significantly inhibited the formation of the hypertrophic synovial lining in the TPT condition.. These results collectively indicate that inducible PI3Kδ plays a crucial role in persistent activation of PI3K-Akt in FLSs, and in the formation of a hypertrophic synovial lining. PI3Kδ may be an alternative treatment target for the regulation of proliferative synovium in RA.

Topics: Arthritis, Rheumatoid; Cadherins; Cells, Cultured; Humans; Hyperplasia; Matrix Metalloproteinase 3; Phosphatidylinositol 3-Kinase; Platelet-Derived Growth Factor; RNA, Small Interfering; Signal Transduction; Synovial Membrane; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha; Up-Regulation

The study of kidney cancer pathogenesis and its treatment has been limited by the scarcity of genetically defined animal models. The FLCN gene that codes for the protein folliculin, mutated in Birt-Hogg-Dubé syndrome, presents a new target for mouse modeling of kidney cancer. Here we developed a kidney-specific knockout model by disrupting the mouse Flcn in the proximal tubules, thus avoiding homozygous embryonic lethality or neonatal mortality, and eliminating the requirement of loss of heterozygosity for tumorigenesis. This knockout develops renal cysts and early onset (6 months) of multiple histological subtypes of renal neoplasms featuring high tumor penetrance. Although the majority of the tumors were chromophobe renal cell carcinomas in affected mice under 1 year of age, papillary renal cell carcinomas predominated in the kidneys of older knockout mice. This renal neoplasia from cystic hyperplasia at 4 months to high-grade renal tumors by 16 months represented the progression of tumorigenesis. The mTOR and TGF-β signalings were upregulated in Flcn-deficient tumors, and these two activated pathways may synergetically cause renal tumorigenesis. Treatment of knockout mice with the mTOR inhibitor rapamycin for 10 months led to the suppression of tumor growth. Thus, our model recapitulates human Birt-Hogg-Dubé kidney tumorigenesis, provides a valuable tool for further study of Flcn-deficient renal tumorigenesis, and tests new drugs/approaches to their treatment.

Topics: Animals; Antibiotics, Antineoplastic; Carcinogenesis; Carcinoma, Renal Cell; Cysts; Disease Models, Animal; Hyperplasia; Kidney Neoplasms; Kidney Tubules, Proximal; Mice; Mice, Knockout; Proto-Oncogene Proteins; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases; Transforming Growth Factor beta; Tumor Suppressor Proteins

Mucus hypersecretion is an important pathological feature of chronic airway diseases, such as asthma and pulmonary diseases. MUC5AC is a major component of the mucus matrix forming family of mucins in the airways. The initiation of endoplasmic reticulum (ER)-mediated stress responses contributes to the pathogenesis of airway diseases. The present study investigated that ER stress was responsible for airway mucus production and this effect was blocked by the flavonoid kaempferol. Oral administration of ≥10 mg/kg kaempferol suppressed mucus secretion and goblet cell hyperplasia observed in the bronchial airway and lung of BALB/c mice sensitized with ovalbumin (OVA). TGF-β and tunicamycin promoted MUC5AC induction after 72 h in human bronchial airway epithelial BEAS-2B cells, which was dampened by 20 μM kaempferol. Kaempferol inhibited tunicamycin-induced ER stress of airway epithelial cells through disturbing the activation of the ER transmembrane sensor ATF6 and IRE1α. Additionally, this compound demoted the induction of ER chaperones such as GRP78 and HSP70 and the splicing of XBP-1 mRNA by tunicamycin. The in vivo study further revealed that kaempferol attenuated the induction of XBP-1 and IRE1α in epithelial tissues of OVA-challenged mice. TGF-β and tunicamycin induced TRAF2 with JNK activation and such induction was deterred by kaempferol. The inhibition of JNK activation encumbered the XBP-1 mRNA splicing and MUC5AC induction by tunicamycin and TGF-β. These results demonstrate that kaempferol alleviated asthmatic mucus hypersecretion through blocking bronchial epithelial ER stress via the inhibition of IRE1α-TRAF2-JNK activation. Therefore, kaempferol may be a potential therapeutic agent targeting mucus hypersecretion-associated pulmonary diseases.

Topics: Animals; Cell Line; Endoplasmic Reticulum Chaperone BiP; Endoplasmic Reticulum Stress; Endoribonucleases; Goblet Cells; Humans; Hyperplasia; Immunization; JNK Mitogen-Activated Protein Kinases; Kaempferols; Male; Mice; Mucus; Ovalbumin; Protein Serine-Threonine Kinases; Respiratory Mucosa; Signal Transduction; TNF Receptor-Associated Factor 2; Transforming Growth Factor beta; Unfolded Protein Response

Hemodialysis as an efficient therapy for advanced CKD is the most used treatment modality all over the world. Even though primary AVF is widely accepted as a best permanent vascular access in hemodialysis patients, up to 60% of all fistulas fail to mature. The pathogenesis of early fistula failure is not very well understood. Many general and local factors are involved: patient's age, sex, primary renal disease, small vessel's diameter, presence of accessory veins, prior venipunctures, surgical skill, genetics, etc. Histological investigations have confirmed the neointimal venous hyperplasia as a major pathological finding in stenotic lesions of AVF failure, due to local inflammation, oxidative stress and migration and proliferation of myofibroblasts, fibroblasts and endothelial cells.. A total of 89 patients with stadium 4-5 of CKD are involved in the study. A typical radio-cephalic AVF is created in all patients. Part of the fistula vein was taken for histological, immunohistochemical (Vimentin, TGF β and KI67) and morphometric analysis. Appriopriate statistical method was applied.. Up to 80% of the patients showed some degree of endothelial changes at the time of creation of AVF, among them 19 pts with substantial intimal hyperplasia, 51 with medial hypertrophy and 19 pts with normal histology. Almost two thirds of the patients did not have expression of TGFβ. More than 95% had some expression of Vimentin. None of the patients had expression of the marker KI 67.. Medial hypertrophy is predominant preexisting pathohistological lesion prior the AVF creation, despite the presence of neointimal hyperplasia. The absence of TGFβ expression in majority of our patients could suggest that inflammation and oxidative stress are developing later, after vascular access surgery. The dominant cells within the stenosis in the veins are myofibroblasts. Their increased presence maybe a reason why some patients are prone to developing venous endothelial changes as a results of exaggerated vascular endothelial response to the effect of uremia, hypertension and other insults.

Topics: Adolescent; Adult; Aged; Aged, 80 and over; Arteriovenous Shunt, Surgical; Biomarkers; Endothelial Cells; Female; Graft Occlusion, Vascular; Humans; Hyperplasia; Hypertrophy; Immunohistochemistry; Ki-67 Antigen; Male; Middle Aged; Neointima; Prospective Studies; Radial Artery; Renal Dialysis; Renal Insufficiency, Chronic; Risk Factors; Severity of Illness Index; Transforming Growth Factor beta; Treatment Failure; Veins; Vimentin; Young Adult

The pathogenesis of denture-induced fibrous hyperplasias has not been examined in detail to explain how tissue injury results in fibrous hyperplasia of the oral mucosa.. We examined the presence of mast cells and myofibroblasts in 33 denture-induced fibrous hyperplasias (DIFH) compared with 10 healthy gingival tissues. The parameters examined included mast cell numbers, tissue distribution, degranulation, and cell subtypes using immunohistochemistry. The presence of myofibroblasts and their likely origin was also examined by double immunofluorescense staining. Furthermore, we investigated the synthesis of osteopontin and TGF-β, considered to be involved in the transformation of a fibroblast to a myofibroblast.. The results demonstrated that the mast cell numbers are significantly increased in the DIFH compared with non-disease controls. The mast cell localization in lesions was higher in the superficial areas with inflammatory cell infiltration compared with the deep fibrotic area (P < 0.01). The number of tryptase-positive mast cells was significantly higher compared with chymase-positive ones. The TGF-β- or osteopontin-positive cell infiltration into the lesion was found in high numbers. The presence of myofibroblasts was identified in 14 of 33 cases (42%), and some of these cells showed apoptosis when assessed by the TUNEL assay. On the survey of the origin of myofibroblasts, results showed αSMA and vimentin positivity indicating these transformed from fibroblasts.. These results are the first to show that mast cells and myofibroblasts can be detected in DIFH, indicating important roles of these cells in the pathogenesis of this lesion.

Topics: Actins; Aged; Aged, 80 and over; Apoptosis; Cell Count; Cell Degranulation; Cell Transdifferentiation; Chymases; Dentures; Female; Fibroblasts; Fibrosis; Gingiva; Humans; Hyperplasia; Male; Mast Cells; Middle Aged; Mouth Mucosa; Myofibroblasts; Osteopontin; Transforming Growth Factor beta; Tryptases; Vimentin

The saphenous vein is the conduit of choice in bypass graft procedures. Haemodynamic factors play a major role in the development of intimal hyperplasia (IH), and subsequent bypass failure. To evaluate the potential protective effect of external reinforcement on such a failure, we developed an ex vivo model for the perfusion of segments of human saphenous veins under arterial shear stress. In veins submitted to pulsatile high pressure (mean pressure at 100 mmHg) for 3 or 7 days, the use of an external macroporous polyester mesh 1) prevented the dilatation of the vessel, 2) decreased the development of IH, 3) reduced the apoptosis of smooth muscle cells, and the subsequent fibrosis of the media layer, 4) prevented the remodelling of extracellular matrix through the up-regulation of matrix metalloproteinases (MMP-2, MMP-9) and plasminogen activator type I. The data show that, in an experimental ex vivo setting, an external scaffold decreases IH and maintains the integrity of veins exposed to arterial pressure, via increase in shear stress and decrease wall tension, that likely contribute to trigger selective molecular and cellular changes.

Topics: Aged; Aged, 80 and over; Caspase 3; Down-Regulation; Ephrin-B2; Female; Heme Oxygenase (Decyclizing); Humans; Hyperplasia; In Vitro Techniques; Male; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Middle Aged; Myocytes, Smooth Muscle; Nitric Oxide Synthase Type III; Perfusion; Plasminogen Activator Inhibitor 1; Pressure; Receptor, EphB4; Saphenous Vein; Stress, Mechanical; Tissue Scaffolds; Transforming Growth Factor beta; Tunica Intima

In the present study, we aimed to deterimine the dose-related effects of ticagrelor, the first reversible inhibitor of the P2Y12 receptor, found in smooth muscle cells as well as platelets, during neointimal hyperplasia in a rabbit carotid anastomosis model.. This study was an experimental, prospective, randomized controlled study including 20 New Zealand white female rabbits (6-months old; weighing 2300 ± 300 g). Under general anaesthesia, the rabbits underwent transection of the right carotid artery and subsequent anastomosis of both ends. The study animals were divided into the following 4 groups: T1 (ticagrelor 5 mg/kg, orally, daily), T2 (ticagrelor 10 mg/kg, orally, daily), T3 (ticagrelor 20 mg/kg, orally, daily) and control (no ticagrelor treatment). The single oral doses were administered in phosphate-buffered saline. The control group received sterile phosphate-buffered saline (2 ml/kg/day, orally) for 3 weeks postoperatively. At the end of the study, the animals were killed, and the anastomosed segment of the right carotid artery and part of the left carotid artery were excised from each animal. Antibodies against transforming growth factor-β were used in staining of arterial sections, which was followed by histomorphological and immunohistochemical studies.. The median intimal thickness (2.0 ± 0.14 µm left vs 73.4 ± 35.8 µm anastomosed right arteries; P <0.05), the median medial thickness (70.8 ± 5.6 µm left vs 92.3 ± 4.5 µm anastomosed right arteries; P <0.05) and the index ratio of intimal thickness to medial thickness (0.03 ± 0.00 left vs 0.8 ± 0.35 anastomosed control right arteries; P <0.05) increased significantly in the anastomosed right arteries compared with the left carotid arteries in the control group. In the treatment groups, the intimal thickness (73.4 ± 35.8 µm in control group vs T1 32.7 ± 19;1 µm, T2 1.9 ± 0.09 µm and T3 2.2 ± 0.5 µm; P = 0.047, P = 0.009 and P = 0.009, respectively), carotid artery intima/media ratio (0.8 ± 0.35 in control group vs T1 0.4 ± 0.2, T2 0.03 ± 0.01 and T3 0.03 ± 0.01 in ticagrelor groups; P = 0.028, P = 0.009 and P = 0.009, respectively) and medial thickness (92.3 ± 4.5 µm in control group vs T2 65.6 ± 7.1 and T3 66.1 ± 7.6 µm; P = 0.009 and P = 0.009, respectively) decreased significantly in the anastomosed right arteries.. This study indicates that effective doses (10 and 20 mg/kg, daily) of the antiplatelet agent ticagrelor in a rabbit model may be beneficial in prevention of intimal hyperplasia. Restenosis due to intimal hyperplasia has been high. Ticagrelor has also been linked to inhibition of smooth muscle cell proliferation and, hence, reduced intimal hyperplasia.

Topics: Adenosine; Anastomosis, Surgical; Animals; Biopsy; Carotid Arteries; Carotid Stenosis; Disease Models, Animal; Dose-Response Relationship, Drug; Female; Hyperplasia; Immunohistochemistry; Neointima; Platelet Aggregation Inhibitors; Purinergic P2Y Receptor Antagonists; Rabbits; Receptors, Purinergic P2Y12; Recurrence; Ticagrelor; Transforming Growth Factor beta

A Disintegrin and Metalloproteinase (ADAM)-10 plays critical roles in neuronal migration and distribution. Recently, ADAM10 deletion was shown to disrupt myelopoiesis. We found that inducible deletion of ADAM10 using Mx1-driven Cre recombinase for a period of three weeks resulted in mast cell hyperplasia in the skin, intestine and spleen. Mast cells express surface ADAM10 in vitro and in vivo, at high levels compared to other immune cells tested. ADAM10 is important for mast cell migration, since ADAM10-deficiency reduced c-Kit-mediated migration. As with some mast cell proteases, ADAM10 expression could be altered by the cytokine microenvironment, being inhibited by IL-10 or TGFβ1, but not by several other T cell-derived cytokines. Collectively these data show that the ADAM10 protease is an important factor in mast cell migration and tissue distribution, and can be manipulated by environmental cues.

Topics: ADAM Proteins; ADAM10 Protein; Amyloid Precursor Protein Secretases; Animals; Cell Movement; Cell Proliferation; Cells, Cultured; Hyperplasia; Interleukin-10; Mast Cells; Membrane Proteins; Mice; Mice, Inbred C57BL; Mice, Knockout; Peritoneum; RNA Interference; RNA, Small Interfering; Stem Cell Factor; T-Lymphocytes; Transforming Growth Factor beta

We have previously shown that in the presence of elevated Smad3, transforming growth factor-β (TGF-β) transforms from an inhibitor to a stimulant of vascular smooth muscle cell (SMC) proliferation and intimal hyperplasia (IH). Here we identify a novel mechanism through which TGF-β/Smad3 also exacerbates IH by inhibiting SMC apoptosis. We found that TGF-β treatment led to inhibition of apoptosis in rat SMCs following viral expression of Smad3. Conditioned media from these cells when applied to naive SMCs recapitulated this effect, suggesting an autocrine pathway through a secreted factor. Gene array of TGF-β/Smad3-treated cells revealed enhanced expression of vascular endothelial growth factor (VEGF), a known inhibitor of endothelial cell apoptosis. We then evaluated whether VEGF is the secreted mediator responsible for TGF-β/Smad3 inhibition of SMC apoptosis. In TGF-β/Smad3-treated cells, VEGF mRNA and protein as well as VEGF secretion were increased. Moreover, recombinant VEGF-A inhibited SMC apoptosis and a VEGF-A-neutralizing antibody reversed the inhibitory effect of conditioned media on SMC apoptosis. Stimulation of SMCs with TGF-β led to the formation of a complex of Smad3 and hypoxia-inducible factor-1α (HIF-1α) that in turn activated the VEGF-A promoter and transcription. In rat carotid arteries following arterial injury, Smad3 and VEGF-A expression were upregulated. Moreover, Smad3 gene transfer further enhanced VEGF expression as well as inhibited SMC apoptosis. Finally, blocking either the VEGF receptor or Smad3 signaling in injured carotid arteries abrogated the inhibitory effect of Smad3 on vascular SMC apoptosis. Taken together, our study reveals that following angioplasty, elevation of both TGF-β and Smad3 leads to SMC secretion of VEGF-A that functions as an autocrine inhibitor of SMC apoptosis. This novel pathway provides further insights into the role of TGF-β in the development of IH.

Topics: Animals; Apoptosis; Autocrine Communication; Cells, Cultured; Humans; Hyperplasia; Male; Myocytes, Smooth Muscle; Rats; Rats, Sprague-Dawley; Signal Transduction; Smad3 Protein; Transforming Growth Factor beta; Tunica Intima; Vascular Endothelial Growth Factor A

Previous analysis of the Cerberus like 2 knockout (Cerl2-/-) mouse revealed a significant mortality during the first day after birth, mostly due to cardiac defects apparently associated with randomization of the left-right axis. We have however, identified Cerl2-associated cardiac defects, particularly a large increase in the left ventricular myocardial wall in neonates that cannot be explained by laterality abnormalities. Therefore, in order to access the endogenous role of Cerl2 in cardiogenesis, we analyzed the embryonic and neonatal hearts of Cerl2 null mutants that did not display a laterality phenotype. Neonatal mutants obtained from the compound mouse line Cer2-/-::Mlc1v-nLacZ24+, in which the pulmonary ventricle is genetically marked, revealed a massive enlargement of the ventricular myocardium in animals without laterality defects. Echocardiography analysis in Cerl2-/- neonates showed a left ventricular systolic dysfunction that is incompatible with a long lifespan. We uncovered that the increased ventricular muscle observed in Cerl2-/- mice is caused by a high cardiomyocyte mitotic index in the compact myocardium which is mainly associated with increased Ccnd1 expression levels in the left ventricle at embryonic day (E) 13. Interestingly, at this stage we found augmented left ventricular expression of Cerl2 levels when compared with the right ventricle, which may elucidate the regionalized contribution of Cerl2 to the left ventricular muscle formation. Importantly, we observed an increase of phosphorylated Smad2 (pSmad2) levels in embryonic (E13) and neonatal hearts indicating a prolonged TGFβs/Nodal-signaling activation. Concomitantly, we detected an increase of Baf60c levels, but only in Cerl2-/- embryonic hearts. These results indicate that independently of its well-known role in left-right axis establishment Cerl2 plays an important role during heart development in the mouse, mediating Baf60c levels by exerting an important control of the TGFβs/Nodal-signaling pathway.

Topics: Animals; Animals, Newborn; Cardiomyopathies; Cyclin D1; Female; Gene Expression Regulation, Developmental; Heart Ventricles; Hyperplasia; Intercellular Signaling Peptides and Proteins; Mice; Myocytes, Cardiac; Nodal Protein; Signal Transduction; Smad2 Protein; Transforming Growth Factor beta; Ventricular Dysfunction, Left

The aims of this study were to distinguish between the primary and secondary effects of TGF-β signalling disruption by Dox treatment in NTPDase2+ cells; and to investigate the interactions between TGF-β signalling and Jagged2/Notch1 pathway in regulating the expansion of tongue epithelia stem cells.Transgenic mice expressing rtTA from the mouse NTPDase2 promoter or K14 promoter were used to generate an inducible dominant negative TGF-β receptor type II (Tgfbr2) mutant model.Disruption of TGF-β signalling in NTPDase2+ cells initially inhibited the formation of filiform papillae but led to their regeneration over time. In contrast, disruption of TGF-β signalling induced proliferation of lingual epithelia in the middle tongue. We also observed the proliferation of lingual epithelia in the posterior tongue near the circumvallate papillae. Interactions among the TGF-β signalling pathways, Jagged2/Notch1 signalling pathways and epigenetic modifications regulate the expansion of lingual epithelial stem cells. Different molecular mechanisms are involved in the developmental regulation of lingual epithelia and filiform papillae, dependent on the location along the whole tongue. The fluctuating phenotype of tongue epithelia, over time, may be the combined effects of signalling pathways and epigenetic modifications.

Topics: Adenosine Triphosphatases; Animals; Disease Models, Animal; Doxycycline; Epithelium; Genes, Reporter; Hyperplasia; Jagged-2 Protein; Mice; Mice, Inbred C57BL; Mice, Transgenic; Protein Serine-Threonine Kinases; Receptor, Notch1; Receptor, Transforming Growth Factor-beta Type II; Receptors, Transforming Growth Factor beta; Signal Transduction; Tongue; Transforming Growth Factor beta

Infants born with intrauterine growth retardation (IUGR) are at increased risk of adverse pulmonary outcomes at birth, including meconium aspiration and persistent pulmonary hypertension. Preterm infants with IUGR are at especially high risk of developing bronchopulmonary dysplasia (BPD), a disease hallmarked by alveolar hypoplasia. Although vitamin A supplementation has been shown to decrease the incidence of BPD or death in preterm very low birth weight infants, its potential to reduce BPD or death in preterm infants with IUGR remains unknown. We used a well-characterized rat model of caloric restriction to mimic IUGR and determine the impact of IUGR on lung development. We hypothesized that retinoic acid treatment would preserve alveolar formation through increases in key signaling molecules of the retinoic acid signaling pathway. Our results showed that alveolar hypoplasia caused by caloric restriction can be reversed with refeeding, and that retinoic acid prevents the alveolar hypoplasia coincident with the increased expression of elastin and retinoic acid receptor-α and decreased transforming growth factor-β activity in developing rat lungs. These findings suggest that alveolar hypoplasia attributable to caloric restriction is reversible, and raises the possibility that retinoic acid therapy may prove a useful strategy to prevent adverse pulmonary sequelae such as BPD in preterm infants with IUGR.

Topics: Animals; Caloric Restriction; Elastin; Female; Hyperplasia; Lung; Maternal Exposure; Pregnancy; Pulmonary Alveoli; Rats; Rats, Sprague-Dawley; Receptors, Retinoic Acid; Retinoic Acid Receptor alpha; Signal Transduction; Transforming Growth Factor beta; Tretinoin

The proliferative response of hepatocytes in vivo can be induced by two mechanisms: severe damage to hepatic tissue results in regenerative growth and so-called primary hepatocyte mitogens can initiate liver cell proliferation without preceding loss of parenchyma. The regulation of the two responses is quite different. The decreased regenerative response of cirrhotic/fibrotic liver is well known, and is a severe obstacle to surgery of the diseased liver. In the present experiments we investigated the efficiency of a primary hepatocyte mitogen 1,4-Bis[2-(3,5-dichloropyridyloxy)]benzene (TCPOBOB) on two different liver cirrhosis/fibrosis models in mice induced by chronic administration of CCl(4) and thioacetamide respectively. BrdU incorporation and cyclin A expression established clearly that there is a reduced but still powerful mitogenic response of the fibrotic livers. Therefore, primary hepatocyte mitogens appear to be suitable to be used to rescue the regenerative response of cirrhotic livers.

Topics: Animals; Aryl Hydrocarbon Hydroxylases; Biomarkers; Bromodeoxyuridine; Carbon Tetrachloride; Cell Proliferation; Cyclin A; Cytochrome P450 Family 2; Disease Models, Animal; Gene Expression; Hepatocytes; Hyperplasia; Liver; Liver Cirrhosis; Liver Regeneration; Male; Mice; Mice, Inbred C57BL; Pyridines; Steroid Hydroxylases; Thioacetamide; Transforming Growth Factor beta

Portal vein embolization is a treatment option to achieve a sufficient future remnant liver volume for patients with central liver tumours requiring an extended resection with an extensive parenchymal loss. However, molecular mechanisms of this intervention are up to now poorly understood. The objective of this prospective pilot study was the characterization of molecular events leading to late hypertrophy of the non-embolized liver tissue in the human liver.. Liver tissue of ten patients was collected before and intraoperatively more than one month after embolization. Investigation of molecular features was performed by pangenomic chips, polymerase chain reaction, immunostaining of proliferation marker Ki-67 and immunofluorescence measurements.. Significantly elevated genes hint towards angiogenesis and signalling by insulin-like growth factor and associated binding proteins. Increased transcript levels of activator protein 1 complex members like c-jun were reflecting potential molecular events of liver growth after embolization. Immunofluorescence data confirmed a predominant upregulation of β-catenin and c-jun (p<0.1) supported by Ki-67 (p<0.05) in the non-embolized liver. In silico analysis of transcriptomic dysplasia and hepatocellular carcinoma data showed divergent signatures compared to embolization.. Our findings indicate a sustained regeneration after portal vein embolization reflected in hyperplasia and angiogenesis in the human liver and provide novel molecular mechanisms of interlobe crosstalk.

Topics: Activating Transcription Factor 3; Aged; beta Catenin; Down-Regulation; Embolization, Therapeutic; Gene Expression Profiling; Humans; Hyperplasia; Inhibitor of Differentiation Protein 1; Inhibitor of Differentiation Proteins; Insulin-Like Growth Factor Binding Protein 1; Insulin-Like Growth Factor Binding Protein 2; Ki-67 Antigen; Liver; Liver Neoplasms; Liver Regeneration; Middle Aged; Neoplasm Proteins; Neovascularization, Physiologic; Pilot Projects; Portal Vein; Prospective Studies; Proto-Oncogene Proteins c-fos; Proto-Oncogene Proteins c-jun; RNA, Messenger; Signal Transduction; Transcription Factor AP-1; Transcription, Genetic; Transforming Growth Factor beta; Up-Regulation; Vascular Endothelial Growth Factor A

CD109, a glycosylphosphatidylinositol-anchored glycoprotein, is highly expressed in several types of human cancer tissues, in particular, squamous cell carcinomas. In normal human tissues, human CD109 expression is limited to certain cell types including myoepithelial cells of the mammary, lacrimal, salivary, and bronchial glands and basal cells of the prostate and bronchial epithelium. Although CD109 has been reported to negatively regulate transforming growth factor-β signaling in keratinocytes in vitro, its physiologic role in vivo remains largely unknown. To investigate the function of CD109 in vivo, we generated CD109-deficient (CD109(-/-)) mice. Although CD109(-/-) mice were born normally, transient impairment of hair growth was observed. At histologic analysis, kinked hair shafts, ectatic hair follicles with an accumulation of sebum, and persistent hyperplasia of the epidermis and sebaceous glands were observed in CD109(-/-) mice. Immunohistochemical analysis revealed thickening of the basal and suprabasal layers in the epidermis of CD109(-/-) mice, which is where endogenous CD109 is expressed in wild-type mice. Although CD109 was reported to negatively regulate transforming growth factor-β signaling, no significant difference in levels of Smad2 phosphorylation was observed in the epidermis between wild-type and CD109(-/-) mice. Instead, Stat3 phosphorylation levels were significantly elevated in the epidermis of CD109(-/-) mice compared with wild-type mice. These results suggest that CD109 regulates differentiation of keratinocytes via a signaling pathway involving Stat3.

Topics: Animals; Antigens, CD; beta-Galactosidase; Epidermis; Gene Knock-In Techniques; Gene Targeting; Hair; Humans; Hyperplasia; Male; Mice; Mice, Knockout; Neoplasm Proteins; Phosphorylation; Signal Transduction; STAT3 Transcription Factor; Testis; Transforming Growth Factor beta

Vascular remodeling in response to alterations in blood flow has been shown to modulate the formation of neo-intima. This process results from a proliferative response of vascular smooth muscle cells and is influenced by macrophages, which potentiate the development of the intima. The LDL receptor-related protein 1 (LRP1) is a large endocytic and signaling receptor that recognizes a number of ligands including apoE-containing lipoproteins, proteases and protease-inhibitor complexes. Macrophage LRP1 is known to influence the development of atherosclerosis, but its role in vascular remodeling has not been investigated.. To define the contribution of macrophage LRP1 to vascular remodeling, we generated macrophage specific LRP1-deficient mice (macLRP1-/-) on an LDL receptor (LDLr) knock-out background. Using a carotid ligation model, we detected a 2-fold increase in neointimal thickening and a 2-fold increase in the intima/media ratio in macLRP1-/- mice. Quantitative RT-PCR arrays of the remodeled vessel wall identified increases in mRNA levels of the TGF-β2 gene as well as the Pdgfa gene in macLRP1-/- mice which could account for the alterations in vascular remodeling. Immunohistochemistry analysis revealed increased activation of the TGF-β signaling pathway in macLRP1-/- mice. Further, we observed that LRP1 binds TGF-β2 and macrophages lacking LRP1 accumulate twice as much TGF-β2 in conditioned media. Finally, TNF-α modulation of the TGF-β2 gene in macrophages is attenuated when LRP1 is expressed. Together, the data reveal that LRP1 modulates both the expression and protein levels of TGF-β2 in macrophages.. Our data demonstrate that macrophage LRP1 protects the vasculature by limiting remodeling events associated with flow. This appears to occur by the ability of macrophage LRP1 to reduce TGF-β2 protein levels and to attenuate expression of the TGF-β2 gene resulting in suppression of the TGF-β signaling pathway.

Topics: Animals; Carotid Arteries; Cell Proliferation; Extracellular Matrix; Extracellular Signal-Regulated MAP Kinases; Gene Deletion; Gene Expression Profiling; Gene Expression Regulation; Hyperplasia; Immunohistochemistry; Ligation; Low Density Lipoprotein Receptor-Related Protein-1; Macrophages; Mice; Models, Animal; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; Protein Binding; Receptor, Platelet-Derived Growth Factor alpha; RNA, Messenger; Signal Transduction; Smad Proteins; Time Factors; Transforming Growth Factor beta; Tunica Intima; Ventricular Remodeling

CCN3 belongs to the CCN family, which constitutes multifunctional secreted proteins that act as matrix cellular regulators. We investigated the pathophysiological roles of CCN3 in the vessels.. We examined the effects of CCN3 on the proliferation and migration of rat vascular smooth muscle cells (VSMC). CCN3 knockout mice were created, and vascular phenotypes and neointimal hyperplasia induced by photochemically induced thrombosis were investigated. CCN3 suppressed the VSMC proliferation induced by fetal bovine serum. The neutralizing antibody for transforming growth factor-beta did not affect the growth inhibitory effect of CCN3. Moreover, CCN3 enhanced the mRNA expression of cyclin-dependent kinase inhibitors, p21 and p15. Gamma secretase inhibitor, an inhibitor of Notch signaling, partially inhibited the enhanced expression of p21 induced by CCN3. CCN3 also inhibited the VSMC migration. Finally, the histopathologic evaluation of the arteries 21 days after the endothelial injury revealed a 6-fold enhancement of neointimal thickening in the null mice compared with the wild-type mice.. CCN3 suppresses neointimal thickening through the inhibition of VSMC migration and proliferation. Our findings indicate the involvement of CCN3 in vascular homeostasis, especially on injury, and the potential usefulness of this molecule in the modulation of atherosclerotic vascular disease.

Topics: Amyloid Precursor Protein Secretases; Animals; Aorta; Cell Cycle; Cell Movement; Cell Proliferation; Cells, Cultured; Cyclin-Dependent Kinase Inhibitor p15; Cyclin-Dependent Kinase Inhibitor p21; Diabetes Mellitus, Experimental; Diabetic Angiopathies; Femoral Artery; Genotype; Hyperplasia; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; Nephroblastoma Overexpressed Protein; Phenotype; Protease Inhibitors; Rats; Rats, Wistar; Receptors, Notch; Recombinant Proteins; Signal Transduction; Thrombosis; Time Factors; Transfection; Transforming Growth Factor beta

Diabetic nephropathy (DN) characterized as nephrotic syndrome and diffuse glomerulosclerosis can cause renal failure and end-stage kidney disease. Expansion of mesangial matrix around capillaries in the kidney glomeruli is a prominent feature of DN. This study investigated whether licorice extracts inhibited mesangial cell (MC) proliferation and matrix accumulation induced by high glucose (HG). Human renal MC were cultured in media containing 5.5 mM glucose plus 27.5 mM mannitol as an osmotic control or 33 mM glucose for 3 d in the presence of water or ethanol extracts from raw licorice (LW, LE) or roasted licorice (RLW, RLE). Non-polar components including glycyrrhetic acid were elevated during licorice roasting, whereas polar components soluble in water extracts were diminished. Exposure of cells to HG caused significant increases in collagen IV secretion and connective tissue growth factor (CTGF) expression, which was appeased by RLW and RLE at transcriptional levels. The inhibitory potency was high in the order of RLE > or = RLW > or = LE > > LW. Non-polar glycyrrhetic acid but not glycyrrhizin retarded HG-stimulated mesangial matrix deposition through diminishing CTGF expression. In addition, RLW and RLE but not LW modulated membrane type matrix metalloproteinase-1 (MT-1 MMP) expression, MMP-2 activity and tissue inhibitor of MMP-2 (TIMP-2), which facilitated the degradation of mesangial matrix. Furthermore, the augmented expression of CTGF and TIMP-2 in HG-exposed cells was mediated by Akt activation and TGF-beta/Smad signaling through PKCbeta2-responsive signaling pathways. However, HG-down-regulated MT-1 MMP expression was independent of activation of ERK1/2 and Akt when using their inhibitors of DB98059 (ERK1/2) and LY294002 (Akt) alone or in combination. These results demonstrate that extracts from roasted licorice may be highly potent therapeutic agents for the prevention and treatment of mesangial fibrosis and glomerulosclerosis leading to diabetes nephropathy due to longstanding diabetes mellitus.

Topics: Base Sequence; Blotting, Western; Cells, Cultured; Chromatography, High Pressure Liquid; DNA Primers; Enzyme Activation; Extracellular Matrix; Glomerular Mesangium; Glucose; Glycyrrhiza; Humans; Hyperplasia; Plant Extracts; Proto-Oncogene Proteins c-akt; Reverse Transcriptase Polymerase Chain Reaction; Signal Transduction; Transforming Growth Factor beta

Topics: Animals; Endothelial Cells; Endothelium; Hyperplasia; Muscle, Smooth, Vascular; Rats; Stem Cells; Transforming Growth Factor beta; Tunica Intima

Castration experiments in rodents show that the stromal vasculature is critical to the androgen-mediated prostate growth regulation. However, the role of angiogenesis inhibitors, such as thrombospondin-1 (TSP-1), in this process is unclear. TSP-1 is a multifunctional glycoprotein that can function as a potent angiogenesis inhibitor and an in vivo activator of latent transforming growth factor-beta (TGF-beta) in some tissues. On the basis of these observations, we hypothesized that TSP-1 regulated androgen withdrawal-induced prostate regression and that this process was mediated not only through antiangiogenic activity but also through TGF-beta activation. To test this, we evaluated angiogenic activity in human prostate epithelial and stromal cells treated with androgens and hypoxia in vitro. TSP-1 knockout mice were characterized to investigate the in vivo functions of TSP-1. In vitro, we found that androgens and hypoxia differentially regulated TSP-1 and angiogenic activity. Androgens stimulated normal epithelial cell, but inhibited normal stromal cell, angiogenic activity. Conversely, hypoxia stimulated stromal while inhibiting epithelial activity. Thus, in vivo, net angiogenic activity must reflect cellular interactions. And, we found that media conditioned by epithelial cells grown under normoxic conditions stimulated stromal cell angiogenic activity, and if epithelial cells were grown under hypoxic conditions, stromal activity was further increased. TSP-1 levels, however, were unchanged. In vivo, TSP-1 loss in a mouse model led to prostate epithelial hyperplasia by 3 months of age with only a modest stromal effect. Androgens suppressed TSP-1 as expression increased after castration both in normal mouse prostate and in human prostate cancer tissues. In addition, TSP-1 expression corresponded to increased TGF-beta activation in mouse tissues, specifically in the stromal compartment. These data show a critical role for TSP-1 in prostate epithelial and stromal growth regulation through angiogenic inhibition and activation of latent TGF-beta. Therefore, loss of TSP-1 during tumorigenesis would eliminate two barriers to cancer progression.

Topics: Androgens; Animals; Carcinoma; Cell Line; Dihydrotestosterone; Epithelial Cells; Humans; Hyperplasia; Hypoxia; Male; Mice; Mice, Inbred C57BL; Neovascularization, Physiologic; Orchiectomy; Phenotype; Prostate; Prostatic Neoplasms; Stromal Cells; Thrombospondin 1; Transforming Growth Factor beta

Transforming growth factor-beta (TGF-beta) levels are elevated in the nasal mucosa in allergic rhinitis. However, because TGF-beta is secreted extracellulary in latent complexes, it remains unclear whether the local TGF-beta expression actually drives active signaling and affects the pathophysiology of allergic rhinitis. The objective of this study is to investigate whether TGF-beta signaling is activated in allergic rhinitis and plays a role in the pathophysiology of allergic rhinitis.. An ovabumin (OVA)-sensitized and -nasally challenged mouse model of allergic rhinitis was established and phosphorylation of Smad2 in the nasal mucosa was examined by immunohistochemistry. In addition, the effects of the pharmacological inhibition of endogenous TGF-beta signaling on the allergic rhinitis model were histologically examined. Furthermore, phosphorylation of Smad2 in the nasal mucosa samples obtained from patients with allergic rhinitis was also evaluated.. In the mouse model of allergic rhinitis, OVA challenge induced phosphorylation of Smad2 predominantly in epithelial cells in the nasal mucosa. In addition, the administration of an inhibitor of TGF-beta type I receptor kinase activity during OVA challenge suppressed goblet cell hyperplasia in the nasal mucosa. Furthermore, phosphorylated Smad2 expression increased in nasal epithelial cells in patients with allergic rhinitis.. These results suggest that TGF-beta signaling is activated in epithelial cells in the nasal mucosa in allergic rhinitis and may contribute to the development of goblet cell hyperplasia.

Topics: Animals; Cell Proliferation; Disease Models, Animal; Goblet Cells; Humans; Hyperplasia; Immunization; Mice; Nasal Mucosa; Ovalbumin; Phosphorylation; Protein Serine-Threonine Kinases; Pyrazoles; Quinolines; Receptor, Transforming Growth Factor-beta Type I; Receptors, Transforming Growth Factor beta; Rhinitis, Allergic, Perennial; Rhinitis, Allergic, Seasonal; Signal Transduction; Smad2 Protein; Transforming Growth Factor beta

Diabetic patients experience exaggerated intimal hyperplasia after endovascular procedures. Recently it has been shown that circulating smooth muscle progenitor cells (SPC) contribute to intimal hyperplasia. We hypothesized that SPC differentiation would be increased in diabetes and focused on modulation of TGF-β/BMP-6 signaling as potential underlying mechanism.. We isolated SPC from C57Bl/6 mice with streptozotocin-induced diabetes and controls. SPC differentiation was evaluated by immunofluorescent staining for αSMA and collagen Type I. SPC mRNA expression of TGF-β and BMP-6 was quantified using real-time PCR. Intima formation was assessed in cuffed femoral arteries. Homing of bone marrow derived cells to cuffed arterial segments was evaluated in animals transplanted with bone marrow from GFP-transgenic mice.. We observed that SPC differentiation was accelerated and numeric outgrowth increased in diabetic animals (24.6 ± 8.8 vs 8.3 ± 1.9 per HPF after 10 days, p < 0.05). Quantitative real-time PCR showed increased expression of TGF-β and decreased expression of the BMP-6 in diabetic SPC. SPC were MAC-3 positive, indicative of monocytic lineage. Intima formation in cuffed arterial segments was increased in diabetic mice (intima/media ratio 0.68 ± 0.15 vs 0.29 ± 0.06, p < 0.05). In GFP-chimeric mice, bone marrow derived cells were observed in the neointima (4.4 ± 3.3 cells per section) and particularly in the adventitia (43.6 ± 9.3 cells per section). GFP-positive cells were in part MAC-3 positive, but rarely expressed α-SMA.. In conclusion, in a diabetic mouse model, SPC levels are increased and SPC TGF-β/BMP-6 expression is modulated. Altered TGF-β/BMP-6 expression is known to regulate smooth muscle cell differentiation and may facilitate SPC differentiation. This may contribute to exaggerated intimal hyperplasia in diabetes as bone marrow derived cells home to sites of neointima formation.

Topics: Animals; Bone Marrow Cells; Bone Morphogenetic Protein 6; Cells, Cultured; Diabetes Mellitus, Experimental; Diabetes Mellitus, Type 1; Disease Models, Animal; Femoral Artery; Green Fluorescent Proteins; Hyperplasia; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Muscle, Smooth, Vascular; RNA, Messenger; Signal Transduction; Stem Cells; Transforming Growth Factor beta; Tunica Intima

Increased airway smooth muscle (ASM) mass, a characteristic finding in asthma, may be caused by hyperplasia or hypertrophy. Cell growth requires increased translation of contractile apparatus mRNA, which is controlled, in part, by glycogen synthase kinase (GSK)-3beta, a constitutively active kinase that inhibits eukaryotic initiation factor-2 activity and binding of methionyl tRNA to the ribosome. Phosphorylation of GSK-3beta inactivates it, enhancing translation. We sought to quantify the contributions of hyperplasia and hypertrophy to increased ASM mass in ovalbumin (OVA)-sensitized and -challenged BALB/c mice and the role of GSK-3beta in this process. Immunofluorescent probes, confocal microscopy, and stereological methods were used to analyze the number and volume of cells expressing alpha-smooth muscle actin and phospho-Ser(9) GSK-3beta (pGSK). OVA treatment caused a 3-fold increase in ASM fractional unit volume or volume density (Vv) (PBS, 0.006 +/- 0.0003; OVA, 0.014 +/- 0.001), a 1.5-fold increase in ASM number per unit volume (Nv), and a 59% increase in volume per cell (Vv/Nv) (PBS, 824 +/- 76 microm(3); OVA, 1,310 +/- 183 mum(3)). In OVA-treated mice, there was a 12-fold increase in the Vv of pGSK (+) ASM, a 5-fold increase in the Nv of pGSK (+) ASM, and a 1.6-fold increase in Vv/Nv. Lung homogenates from OVA-treated mice showed increased GSK-3beta phosphorylation and lower GSK-3beta activity. Both hyperplasia and hypertrophy are responsible for increased ASM mass in OVA-treated mice. Phosphorylation and inactivation of GSK-3beta are associated with ASM hypertrophy, suggesting that this kinase may play a role in asthmatic airway remodeling.

Topics: Actins; Animals; Asthma; Cell Size; Flow Cytometry; Fluorescent Antibody Technique; Glycogen Synthase Kinase 3; Glycogen Synthase Kinase 3 beta; Hyperplasia; Hypertrophy; Immunoblotting; Immunoprecipitation; Lung; Mice; Mice, Inbred BALB C; Microscopy, Confocal; Microscopy, Fluorescence; Muscle, Smooth; Ovalbumin; Phosphorylation; Pneumonia; Respiratory System; Transforming Growth Factor beta

Inflammatory cytokine interleukin (IL)-6 is elevated in the serum and lungs of patients with pulmonary artery hypertension (PAH). Several animal models of PAH cite the potential role of inflammatory mediators. We investigated role of IL-6 in the pathogenesis of pulmonary vascular disease. Indices of pulmonary vascular remodeling were measured in lung-specific IL-6-overexpressing transgenic mice (Tg(+)) and compared to wild-type (Tg(-)) controls in both normoxic and chronic hypoxic conditions. The Tg(+) mice exhibited elevated right ventricular systolic pressures and right ventricular hypertrophy with corresponding pulmonary vasculopathic changes, all of which were exacerbated by chronic hypoxia. IL-6 overexpression increased muscularization of the proximal arterial tree, and hypoxia enhanced this effect. It also reproduced the muscularization and proliferative arteriopathy seen in the distal arteriolar vessels of PAH patients. The latter was characterized by the formation of occlusive neointimal angioproliferative lesions that worsened with hypoxia and were composed of endothelial cells and T-lymphocytes. IL-6-induced arteriopathic changes were accompanied by activation of proangiogenic factor, vascular endothelial growth factor, the proproliferative kinase extracellular signal-regulated kinase, proproliferative transcription factors c-MYC and MAX, and the antiapoptotic proteins survivin and Bcl-2 and downregulation of the growth inhibitor transforming growth factor-beta and proapoptotic kinases JNK and p38. These findings suggest that IL-6 promotes the development and progression of pulmonary vascular remodeling and PAH through proproliferative antiapoptotic mechanisms.

Topics: Animals; Apoptosis; Arterioles; Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; Blood Pressure; Cell Proliferation; Chronic Disease; Endothelial Cells; Hyperplasia; Hypertension, Pulmonary; Hypertrophy, Right Ventricular; Hypoxia; Inhibitor of Apoptosis Proteins; Interleukin-6; Mice; Mice, Inbred C57BL; Mice, Transgenic; Microtubule-Associated Proteins; Mitogen-Activated Protein Kinases; Muscle, Smooth, Vascular; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-bcl-2; Proto-Oncogene Proteins c-myc; Pulmonary Artery; Repressor Proteins; Survivin; Time Factors; Transforming Growth Factor beta; Up-Regulation; Vascular Endothelial Growth Factor A; Vascular Resistance; Ventricular Function, Right; Ventricular Pressure

To characterize and compare primary and restenotic lesions of the superficial femoral artery and analyze the contribution of TGF-beta/Smad3 signaling to the pathophysiology of peripheral artery occlusive disease.. Immunohistochemical studies were performed on specimens retrieved from the superficial femoral artery of patients undergoing either atherectomy for primary atherosclerotic or recurrent disease after stenting and/or prior angioplasty. Immunohistochemical analysis revealed a significantly higher smooth muscle cell (SMC) content (alpha-actin+) and expression of Smad3 in restenotic lesions while primary lesions contained significantly more leukocytes (CD45+) and macrophages (CD68+). Further studies demonstrated colocalization of Smad3 with alpha-actin and PCNA, suggesting a role for Smad3 in the proliferation observed in restenotic lesions. To confirm a role for Smad3 in SMC proliferation, we both upregulated Smad3 via adenoviral mediated gene transfer (AdSmad3) and inhibited Smad3 through transfection with siRNA in human aortic SMCs, then assessed cell proliferation with tritiated thymidine. Overexpression of Smad3 enhanced whereas inhibition of Smad3 decreased cell proliferation.. Differences in cellular composition and cell proliferation in conjunction with the finding that Smad3 is expressed exclusively in restenotic disease suggest that TGF-beta, through Smad3 signaling, may play an essential role in SMC proliferation and the pathophysiology of restenosis in humans.

Topics: Angioplasty; Apoptosis; Arterial Occlusive Diseases; Atherectomy; Atherosclerosis; Cell Proliferation; Cells, Cultured; Constriction, Pathologic; Femoral Artery; Humans; Hyperplasia; Myocytes, Smooth Muscle; Recurrence; Signal Transduction; Smad3 Protein; Stents; Transfection; Transforming Growth Factor beta; Treatment Outcome

Bone marrow-derived progenitor cells have recently been shown to be involved in the development of intimal hyperplasia after vascular injury. Transforming growth factor-beta (TGF-beta) has profound stimulatory effects on intimal hyperplasia, but it is unknown whether these effects involve progenitor cell recruitment. In this study we found that although TGF-beta had no direct effect on progenitor cell recruitment, conditioned media derived from vascular smooth muscle cells (VSMC) stimulated with TGF-beta induced migration of both total bone marrow (BM) cells and BM-mesenchymal stem cells (MSC) and also induced MSC differentiation into smooth muscle like cells. Furthermore, overexpression of the signaling molecule Smad3 in VSMC via adenovirus-mediated gene transfer (AdSmad3) enhanced the TGF-beta's chemotactic effect. Microarray analysis of VSMC stimulated by TGF-beta/AdSmad3 revealed monocyte chemoattractant protein-1 (MCP-1) as a likely factor responsible for progenitor cell recruitment. We then demonstrated that TGF-beta through Smad3 phosphorylation induced a robust expression of MCP-1 in VSMC. Recombinant MCP-1 mimicked the stimulatory effect of conditioned media on BM and MSC migration. In the rat carotid injury model, Smad3 overexpression significantly increased MCP-1 expression after vascular injury, consistent with our in vitro results. Interestingly, TGF-beta/Smad3-induced MCP-1 was completely blocked by both Ro-32-0432 and rotterlin, suggesting protein kinase C-delta (PKCdelta) may play a role in TGF-beta/Smad3-induced MCP-1 expression. In summary, our data demonstrate that TGF-beta, through Smad3 and PKCdelta, stimulates VSMC production of MCP-1, which is a chemoattractant for bone marrow-derived cells, specifically MSC. Manipulation of this signaling system may provide a novel approach to inhibition of intimal hyperplasia.

Topics: Angioplasty, Balloon; Animals; Aorta, Thoracic; Biomarkers; Blotting, Western; Bone Marrow Cells; Cell Communication; Cell Differentiation; Cell Movement; Cells, Cultured; Chemokine CCL2; Chemotaxis; Gene Expression Profiling; Hyperplasia; Immunoenzyme Techniques; Male; Mesenchymal Stem Cells; Muscle, Smooth, Vascular; Oligonucleotide Array Sequence Analysis; Protein Kinase C-delta; Rats; Rats, Sprague-Dawley; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Signal Transduction; Smad3 Protein; Stem Cells; Transforming Growth Factor beta; Tunica Intima

Poor long-term graft patency remains a major limitation of coronary artery bypass grafting using saphenous vein aortocoronary grafts. Neointimal hyperplasia (NIH) represents the principal mechanism of graft failure; a substantial body of evidence implicates transforming growth factor-beta1 (TGF-beta1) in the pathogenesis of NIH. The small leucine-rich proteoglycans decorin and fibromodulin possess TGF-beta-antagonist activity to differing extents and with differing avidities for the isoforms of TGF-beta. We compared their ability to inhibit NIH in an ex vivo model of human saphenous vein organ culture following adenovirus-mediated gene transfer. Surgically prepared human saphenous vein segments received adenovirus expressing fibromodulin (Ad5-Fmod), decorin (Ad5-Dcn), beta-galactosidase (Ad5-lacZ) or vehicle-only. Computerized morphometry 14 days after infection revealed significantly reduced neointimal area, neointimal thickness and intima/media ratio in Ad5-Fmod- and Ad5-Dcn-infected veins. Each parameter was significantly smaller in Ad5-Fmod- than in Ad5-Dcn-exposed segments. Fibrillar collagen content and levels of biologically active TGF-beta were lower in vessels receiving Ad5-Fmod or Ad5-Dcn than in those receiving Ad5-lacZ or vehicle-only. Fibromodulin is a more potent inhibitor of NIH in cultured human saphenous vein than decorin and offers potential therapeutic benefits in saphenous vein graft failure (and possibly in other forms of accelerated atherosclerosis) by reduction of associated neointima formation.

Topics: Adenoviridae; Collagen; Decorin; Extracellular Matrix Proteins; Fibromodulin; Gene Transfer Techniques; Genetic Therapy; Genetic Vectors; Humans; Hyperplasia; Organ Culture Techniques; Proteoglycans; Saphenous Vein; Transforming Growth Factor beta; Tunica Intima

Transforming growth factor beta (TGF-beta) is a crucial mediator of breast development, and loss of TGF-beta-induced growth arrest is a hallmark of breast cancer. TGF-beta has been shown to inhibit cyclin-dependent kinase (CDK) activity, which leads to the accumulation of hypophosphorylated pRB. However, unlike other components of TGF-beta cytostatic signaling, pRB is thought to be dispensable for mammary development. Using gene-targeted mice carrying subtle missense changes in pRB (Rb1(DeltaL) and Rb1(NF)), we have discovered that pRB plays a critical role in mammary gland development. In particular, Rb1 mutant female mice have hyperplastic mammary epithelium and defects in nursing due to insensitivity to TGF-beta growth inhibition. In contrast with previous studies that highlighted the inhibition of cyclin/CDK activity by TGF-beta signaling, our experiments revealed that active transcriptional repression of E2F target genes by pRB downstream of CDKs is also a key component of TGF-beta cytostatic signaling. Taken together, our work demonstrates a unique functional connection between pRB and TGF-beta in growth control and mammary gland development.

Topics: Animals; Cells, Cultured; Female; Gene Knock-In Techniques; Genotype; Humans; Hyperplasia; Lactation; Male; Mammary Glands, Animal; Mammary Neoplasms, Animal; Mice; Mice, Inbred C57BL; Mice, Transgenic; Models, Molecular; Phenotype; Protein Conformation; Retinoblastoma Protein; Signal Transduction; Tissue Transplantation; Transforming Growth Factor beta

The objective of this study was to better understand the role of transforming growth factor-beta (TGF-beta) and its primary signaling protein Smad3 in the development of intimal hyperplasia. Male Sprague-Dawley rats underwent left carotid balloon injury followed by intra-arterial infection with adenovirus-expressing Smad3 (AdSmad3). In uninfected injured arteries, endogenous Smad3 was upregulated with the expression peaking at 14 days. Moreover, in arteries infected with AdSmad3, we observed an enhancement of intimal hyperplasia and increased vascular smooth muscle cell (VSMC) proliferation. The novel finding, that TGF-beta/Smad3 stimulated rather than inhibited VSMC proliferation, was confirmed in cultured VSMCs infected with AdSmad3 and treated with TGF-beta. To identify the mechanism underlying TGF-beta/Smad3-mediated VSMC proliferation, we studied the cyclin-dependent kinase inhibitor p27. Although the upregulation of Smad3 in VSMCs had no significant effect on total p27 levels, Smad3 did stimulate the phosphorylation of p27 at serine-10 as well as the nuclear export of p27, events associated with cell proliferation. Furthermore, serine-10-phosphorylated p27 was also increased in AdSmad3-infected injured rat carotid arteries, demonstrating the existence of this same mechanism in vivo. In conclusion, our findings identify a novel mechanism for the effect of TGF-beta on intimal hyperplasia. In the presence of elevated levels of Smad3 that develop in response to injury, TGF-beta stimulates smooth muscle cell proliferation through a mechanism involving the phosphorylation and nuclear export of p27.

Topics: Angioplasty, Balloon; Animals; Carotid Artery Injuries; Cell Division; Cells, Cultured; Cyclin-Dependent Kinase Inhibitor p27; Down-Regulation; Hyperplasia; Male; Muscle, Smooth, Vascular; Phosphorylation; Rats; Rats, Sprague-Dawley; Signal Transduction; Smad3 Protein; Transforming Growth Factor beta; Tunica Intima; Up-Regulation

Increased expression of plasminogen activator inhibitor-1 (PAI-1) in vascular tissues is a potential factor linking diabetes to restenosis after percutaneous coronary intervention. Recent studies have shown that cilostazol, a selective type 3 phosphodiesterase inhibitor, prevents neointimal hyperplasia and in-stent thrombosis in patients with diabetes after coronary angioplasty and stent implantation. However, the molecular mechanism of this drug has not been fully elucidated. We examined whether cilostazol inhibits PAI-1 expression in vascular smooth muscle cells (VSMCs) and neointimal hyperplasia. We found that cilostazol effectively inhibits angiotensin II-, high glucose- and TGF-beta-stimulated PAI-1 expression in vivo and in vitro. Cilostazol attenuated PAI-1 expression in neointimal regions and inhibited neointimal hyperplasia after balloon injury. Cilostazol inhibited PAI-1 expression by multiple mechanisms including downregulation of TGF-beta, JNK and p38 signaling pathways. Cilostazol also inhibited transactivating activity at the PAI-1 promoter by Smad3, leading to a suppression of PAI-1 gene transcription. Taken together with its antiproliferative effect on VSMCs, this may explain how cilostazol exerts its antithrombogenic effects after angioplasty and stent implantation.

Topics: Angioplasty, Balloon; Angiotensin II; Animals; Binding Sites; Blood Glucose; Carotid Arteries; Carotid Artery Injuries; Cell Proliferation; Cells, Cultured; Cilostazol; Diabetes Mellitus, Experimental; Dose-Response Relationship, Drug; Fibrinolytic Agents; Hyperplasia; JNK Mitogen-Activated Protein Kinases; Male; Muscle, Smooth, Vascular; p38 Mitogen-Activated Protein Kinases; Phosphodiesterase Inhibitors; Plasminogen Activator Inhibitor 1; Promoter Regions, Genetic; Rats; Rats, Sprague-Dawley; Signal Transduction; Smad3 Protein; Tetrazoles; Transcriptional Activation; Transforming Growth Factor beta; Tunica Intima

Topics: Animals; Cell Proliferation; Constriction, Pathologic; Fibrosis; Graft Occlusion, Vascular; Humans; Hyperplasia; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Signal Transduction; Transforming Growth Factor beta; Tunica Intima; Vascular Patency; Veins

Topics: Animals; Carotid Artery Injuries; Cell Differentiation; Cell Division; Cyclin-Dependent Kinase Inhibitor p21; Gene Expression Regulation; Humans; Hyperplasia; Inflammation; Kruppel-Like Factor 4; Kruppel-Like Transcription Factors; Ligation; Mice; Models, Biological; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; Promoter Regions, Genetic; Response Elements; Transcription, Genetic; Transforming Growth Factor beta; Tunica Intima

Despite ubiquitous expression of the keratoepithelin (KE) protein encoded by the transforming growth factor beta induced/beta induced gene human clone 3 (TGFBI/BIGH3) gene, corneal dystrophies are restricted to the cornea, and no other tissues are affected. We investigated the role of TGFBI/BIGH3 in Groenouw corneal dystrophies by generating transgenic mice overexpressing TGFBI/BIGH3 containing the R555W mutation.. Transgenic animals expressing the Groenouw mutation of human TGFBI/BIGH3 were generated using lentiviral vectors. The line expressed TGFBI/BIGH3 containing the R555W mutation under the control of the phosphoglycerate kinase (PGK) promoter. Expression of the transgene was monitored by Southern and western blotting and by RT-PCR. Electroretinogram analysis was performed and four mice were subjected to complete necroscopy.. Transgene expression was observed in different organs although without specific expression in the cornea. The overall morphology of the transgenic animals was not severely affected by KE overexpression. However, we observed an age-dependent retinal degeneration both functionally and histologically. Female-specific follicular hyperplasia in the spleen and increased levels of lipofuscin in the adrenal gland were also seen in transgenic animals.. Cellular degeneration in the retina of transgenic animals suggest that perturbation of the transforming growth factor beta (TGFbeta) family regulation may affect photoreceptor survival and may induce possible accelerated aging in several tissues. No corneal phenotype could be observed, probably due to the lack of transgene expression in this tissue.

Topics: Animals; Blotting, Southern; Blotting, Western; Electroretinography; Extracellular Matrix Proteins; Female; Gene Expression Regulation; Humans; Hyperplasia; Lentivirus; Male; Mice; Mice, Transgenic; Mutant Proteins; Organ Size; Organ Specificity; Phosphoglycerate Kinase; Promoter Regions, Genetic; Reproducibility of Results; Retinal Degeneration; RNA, Messenger; Spleen; Transforming Growth Factor beta; Virus Integration

Transforming growth factor (TGF)-beta signaling has been associated with early tumor suppression and late tumor progression; however, many of the mechanisms that mediate these processes are not known. Using Cre/LoxP technology, with the whey acidic protein promoter driving transgenic expression of Cre recombinase (WAP-Cre), we have now ablated the type II TGF-beta receptor (T beta RII) expression specifically within mouse mammary alveolar progenitors. Transgenic expression of the polyoma virus middle T antigen, under control of the mouse mammary tumor virus enhancer/promoter, was used to produce mammary tumors in the absence or presence of Cre (T beta RII((fl/fl);PY) and T beta RII((fl/fl);PY;WC), respectively). The loss of TGF-beta signaling significantly decreased tumor latency and increased the rate of pulmonary metastasis. The loss of TGF-beta signaling was significantly correlated with increased tumor size and enhanced carcinoma cell survival. In addition, we observed significant differences in stromal fibrovascular abundance and composition accompanied by increased recruitment of F4/80(+) cell populations in T beta RII((fl/fl);PY;WC) mice when compared with T beta RII((fl/fl);PY) controls. The recruitment of F4/80(+) cells correlated with increased expression of known inflammatory genes including Cxcl1, Cxcl5, and Ptgs2 (cyclooxygenase-2). Notably, we also identified an enriched K5(+) dNp63(+) cell population in primary T beta RII((fl/fl);PY;WC) tumors and corresponding pulmonary metastases, suggesting that loss of TGF-beta signaling in this subset of carcinoma cells can contribute to metastasis. Together, our current results indicate that loss of TGF-beta signaling in mammary alveolar progenitors may affect tumor initiation, progression, and metastasis through regulation of both intrinsic cell signaling and adjacent stromal-epithelial interactions in vivo.

Topics: Animals; Bone Marrow Cells; Breast Cyst; Cell Differentiation; Cell Survival; Disease Progression; Hyperplasia; Lung Neoplasms; Mammary Neoplasms, Experimental; Mice; Neoplastic Stem Cells; Precancerous Conditions; Protein Serine-Threonine Kinases; Receptor, Transforming Growth Factor-beta Type II; Receptors, Transforming Growth Factor beta; Signal Transduction; Stromal Cells; Transforming Growth Factor beta

Cyclin D1/cyclin-dependent kinase 2 (Cdk2) complexes are present at high frequency in human breast cancer cell lines, but the significance of this observation is unknown. This report shows that expression of a cyclin D1-Cdk2 fusion protein under the control of the mouse mammary tumor virus (MMTV) promoter results in mammary gland hyperplasia and fibrosis, and mammary tumors. Cell lines isolated from MMTV-cyclin D1-Cdk2 (MMTV-D1K2) tumors exhibit Rb and p130 hyperphosphorylation and up-regulation of the protein products of E2F-dependent genes. These results suggest that cyclin D1/Cdk2 complexes may mediate some of the transforming effects that result from cyclin D1 overexpression in human breast cancers. MMTV-D1K2 cancer cells express the hepatocyte growth factor (HGF) receptor, c-Met. MMTV-D1K2 cancer cells also secrete transforming growth factor beta (TGFbeta), but are relatively resistant to TGFbeta antiproliferative effects. Fibroblasts derived from MMTV-D1K2 tumors secrete factors that stimulate the proliferation of MMTV-D1K2 cancer cells, stimulate c-Met tyrosine phosphorylation, and stimulate the phosphorylation of the downstream signaling intermediates p70(s6k) and Akt on activating sites. Together, these results suggest that deregulation of the Cdk/Rb/E2F axis reprograms mammary epithelial cells to initiate a paracrine loop with tumor-associated fibroblasts involving TGFbeta and HGF, resulting in desmoplasia. The MMTV-D1K2 mice should provide a useful model system for the development of therapeutic approaches to block the stromal desmoplastic reaction that likely plays an important role in the progression of multiple types of human tumors.

Topics: Animals; Cyclin D1; Cyclin-Dependent Kinase 2; Disease Progression; Enzyme Activation; Female; Fibroblasts; Hepatocyte Growth Factor; Hyperplasia; Mammary Glands, Animal; Mammary Neoplasms, Experimental; Mammary Tumor Virus, Mouse; Mice; Mice, Transgenic; Promoter Regions, Genetic; Recombinant Fusion Proteins; Retinoblastoma Protein; Retinoblastoma-Like Protein p130; Transforming Growth Factor beta

Enhanced extracellular matrix accumulation rather than cell proliferation contributes to later stages of in-stent restenosis. Aldosterone itself has been shown to increase cardiovascular fibrosis, therefore, we studied the suppressive effects of eplerenone, a new aldosterone receptor antagonist, on neointimal hyperplasia after coronary stent implantation in swine.. Palmatz-Shatz stents were implanted in the left anterior descending artery of 36 pigs. One hundred milligrams of Eplerenone was orally administered from 1 week before, to 4 weeks after stent implantation in Group E (n=18), and vehicle was given to Group C (n=18). Pigs were sacrificed 1 or 4 weeks after stent implantation. The number of infiltrating macrophages was calculated at 1 week. Morphometrical analysis was performed to measure the area of each layer, and %area of fibrosis and mRNA for collagen I, III and TGF-beta was analyzed by RT-PCR at 4 weeks.. The number of infiltrating macrophages was less in Group E than in Group C (p<0.01). The overall size of coronary arteries at 4 weeks was similar in both groups. However, the luminal area was larger in Group E than in Group C (p<0.05), and the intimal area was smaller in Group E than in Group C (p<0.05). The %area of fibrosis was significantly less in Group E than in Group C at 4 weeks (p<0.01). In Group E, the expression of mRNA for collagen I, III and TGF-beta was significantly reduced.. Orally administered eplerenone attenuated collagen accumulation within the neointima, thereby inhibiting neointimal hyperplasia after stent implantation.

Topics: Actins; Administration, Oral; Animals; Blood Vessel Prosthesis Implantation; Collagen Type I; Collagen Type III; Coronary Restenosis; Coronary Vessels; Disease Models, Animal; Eplerenone; Fibrosis; Hyperplasia; Immunohistochemistry; Macrophages; Male; Mineralocorticoid Receptor Antagonists; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Spironolactone; Stents; Swine; Time Factors; Transforming Growth Factor beta; Tunica Intima

Smad4 is the common mediator for TGFbeta signals, which play important functions in many biological processes. To study the role of Smad4 in skin development and epidermal tumorigenesis, we disrupted this gene in skin using the Cre-loxP approach. We showed that absence of Smad4 blocked hair follicle differentiation and cycling, leading to a progressive hair loss of mutant (MT) mice. MT hair follicles exhibited diminished expression of Lef1, and increased proliferative cells in the outer root sheath. Additionally, the skin of MT mice exhibited increased proliferation of basal keratinocytes and epidermal hyperplasia. Furthermore, we provide evidence that the absence of Smad4 resulted in a block of both TGFbeta and bone morphogenetic protein (BMP) signaling pathways, including p21, a well-known cyclin-dependent kinase inhibitor. Consequently, all MT mice developed spontaneous malignant skin tumors from 3 months to 13 months of age. The majority of tumors are malignant squamous cell carcinomas. A most notable finding is that tumorigenesis is accompanied by inactivation of phosphatase and tensin homolog deleted on chromosome 10 (Pten), activation of AKT, fast proliferation and nuclear accumulation of cyclin D1. These observations revealed the essential functions of Smad4-mediated signals in repressing skin tumor formation through the TGFbeta/BMP pathway, which interacts with the Pten signaling pathway.

Topics: Alopecia; Animals; Bone Morphogenetic Proteins; Carcinoma, Squamous Cell; Cell Differentiation; Cell Nucleus; Cell Proliferation; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p21; Enzyme Activation; Epidermis; Female; Hair Follicle; Hyperplasia; In Situ Hybridization; Integrases; Keratinocytes; Male; Mice; Mice, Knockout; Mice, Transgenic; Proto-Oncogene Proteins c-akt; PTEN Phosphohydrolase; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Signal Transduction; Skin; Skin Neoplasms; Smad4 Protein; Transforming Growth Factor beta

The transforming growth factor (TGF)-beta family of cytokines exerts pleiotropic actions on vascular smooth muscle cell phenotype, proliferation, and extracellular matrix synthesis. This in vivo study assessed the use of TGF-beta3 in attenuating the development of postanastomotic smooth muscle cell proliferation.. Under general anesthesia, 10 adult goats underwent transection and reanastomosis of both common carotid arteries. After reanastomosis, one artery was infiltrated with 50 ng of TGF-beta3 in 100 microL of pH buffer around the anastomosis, and the other side was infiltrated with buffer only. After surgery, each animal received 150 mg of aspirin daily. The arteries were explanted after 3 months for histologic examination.. Vessel wall thickness surrounding the anastomosis was reduced by 30% after TGF-beta3 treatment compared with placebo (P = .003), with a 20% (P = .002) reduction in cellular content. Although total collagen content was not significantly different between TGF-beta3 and placebo, collagen type VIII content was reduced around the TGF-beta3 anastomoses (P = .011). A reduction in the total elastin content (P = .003) and number of elastic fiber lamellae (P = .042) was found surrounding TGF-beta3-treated anastomoses, but not placebo-treated anastomosis. A 29% increase in vasa vasorum (P = .044) was present around TGF-beta3-treated anastomoses. No differences in inflammatory cell infiltration were seen between sides.. Direct subadventitial infiltration of TGF-beta3 immediately after creation of an arterial anastomosis attenuates cell proliferation, with a reduction in elastin and collagen type VIII content and vessel wall thickness.

Topics: Anastomosis, Surgical; Animals; Arteries; Female; Goats; Hyperplasia; Injections, Intralesional; Muscle, Smooth, Vascular; Postoperative Complications; Transforming Growth Factor beta; Transforming Growth Factor beta3; Tunica Intima

Airway remodeling is an important feature of chronic asthma that causes irreversible airflow obstruction. Although asthma is considered to be a Th2 disease, the role of T-bet and GATA-3, the key transcription factors for differentiation toward Th1 and Th2 cells, in the pathogenesis of airway remodeling is poorly understood.. We therefore examined the effects of GATA-3 or T-bet induction of Th1/Th2 bias on the development of airway remodeling in mice.. The development of airway remodeling after repeated allergen challenges was analyzed using transgenic mice overexpressing either GATA-3 or T-bet.. The degrees of subepithelial fibrosis and airway smooth muscle hyperplasia after repeated allergen exposure were significantly enhanced in mice overexpressing GATA-3, compared with wild-type mice. Allergen-induced goblet cell hyperplasia and mucus hypersecretion were significantly lower in mice overexpressing T-bet than in wild-type mice. Eosinophilic airway inflammation increased in mice overexpressing GATA-3, but decreased in mice overexpressing T-bet after repeated allergen exposure. Cytokine analysis revealed that the Th1/Th2 cytokine balance shifted to Th2 in lung homogenates and lung T cells of mice overexpressing GATA-3, whereas this balance shifted to Th1 in those of mice overexpressing T-bet after allergen exposure. Lung transforming growth factor-beta and eotaxin levels were associated with the degree of subepithelial fibrosis and eosinophilic airway inflammation, respectively.. Overall, the results indicate that development of airway remodeling is regulated by the lung Th1/Th2 bias induced by GATA-3 and T-bet.

Topics: Animals; Asthma; Chemokine CCL11; Chemokines, CC; Disease Models, Animal; Eosinophils; Fibrosis; GATA3 Transcription Factor; Goblet Cells; Hyperplasia; Hypertrophy; Immunoglobulins; Interferon-gamma; Interleukin-4; Lung; Mice; Mice, Transgenic; Mucins; Muscle, Smooth; T-Box Domain Proteins; Th1 Cells; Th2 Cells; Transcription Factors; Transforming Growth Factor beta

We investigated the therapeutic potential of a newly developed antifibrotic agent, pirfenidone, to regulate airway remodeling and the development of allergic airway inflammation and airway hyperresponsiveness after chronic allergen challenge. Administration of pirfenidone after sensitization but during the period of ovalbumin challenge significantly prevented the development of airway hyperresponsiveness and prevented eosinophil and lymphocyte accumulation in the airways. IL-4, IL-5, and IL-13 levels in bronchoalveolar lavage fluid and ovalbumin-specific serum IgE antibody levels were also significantly reduced. Treatment with pirfenidone significantly reduced transforming growth factor-beta1 and platelet-derived growth factor levels in bronchoalveolar lavage fluid. Pirfenidone reduced the expression of transforming growth factor-beta1, the development of goblet cell hyperplasia and subepithelial collagenization, and the increases in contractile elements in the lung. These data indicate that pirfenidone may play an important role in the treatment of asthma and has the potential reduce or prevent airway remodeling.

Topics: Allergens; Animals; Anti-Inflammatory Agents, Non-Steroidal; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Proliferation; Cytokines; Eosinophils; Goblet Cells; Hyperplasia; Immunoglobulin E; Interleukin-13; Interleukin-4; Interleukin-5; Leukocytes, Mononuclear; Lymphocytes; Mice; Mice, Inbred BALB C; Ovalbumin; Platelet-Derived Growth Factor; Pyridones; Transforming Growth Factor beta; Transforming Growth Factor beta1

In a rat model of endarterectomy we investigated the potential role of the peroxynitrite-poly(ADP-ribose) polymerase (PARP) pathway in neointima formation and the effects of irradiation, pharmacologic inhibition of PARP, or combined pharmacologic inhibition of PARP and irradiation on vascular remodeling.. Carotid endarterectomy was performed by incision of the left carotid artery with removal of intima in Sprague-Dawley rats. Six groups were studied: sham-operated rats (n = 10), control endarterectomized rats (n = 10), or endarterectomized rats irradiated with 15 Gy (n = 10), or treated with PARP inhibitor, INO-1001 (5 mg/kg/day) (n = 10), or with combined treatment with INO-1001 and irradiation with 5 Gy (n = 10) or with 15 Gy (n = 10). After 21 days, neointima formation and vascular remodeling were assessed.. Neointima formation after endarterectomy was inhibited by postoperative irradiation with 15 Gy and was attenuated by PARP inhibition. However, in parallel to inhibition of neointimal hyperplasia, activation of the peroxynitrite-PARP pathway in the outer vessel wall layers was triggered by postoperative irradiation. Combined pharmacologic PARP inhibition and irradiation with 15 Gy significantly reduced both neointimal hyperplasia and activation of the peroxynitrite-PARP pathway in the outer vessel wall layers. Combination of PARP inhibition and irradiation with 5 Gy was less effective than both PARP inhibition or irradiation with 15 Gy alone.. We conclude, that combined PARP inhibition and irradiation with 15 Gy may be a new dual strategy for prevention of restenosis after surgical vessel reconstruction: combining the strong antiproliferative effect of irradiation and ameliorating irradiation-induced side effects caused by excessive PARP activation.

Topics: Animals; Endarterectomy, Carotid; Hyperplasia; Indoles; Male; Peroxynitrous Acid; Poly Adenosine Diphosphate Ribose; Radiation Dosage; Rats; Rats, Sprague-Dawley; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tumor Necrosis Factor-alpha; Tunica Intima

It was hypothesized that chymase may participate in hemodialysis vascular access dysfunction, as chymase has been known to be an effective enzyme in the conversion of angiotensin I (Ang I) to Ang II and in the latent TGF-beta1 to the active form. An arteriovenous (AV) fistula was created between the brachial artery and vein in dogs. In the AV anastomosis, when the walls of the venous and arterial sides were compared, the eccentric neointimal formation was most evident in the venous wall. Compared with the venous side downstream of the AV anastomosis, a severe neointimal hyperplasia was found in the venous side upstream of the AV anastomosis (intima/media, 153 +/- 25%). The chymase- and TGF-beta-positive mast cells were markedly accumulated in the proliferous neointima and media. In association with the reduction of chymase expression, a marked decrease in Ang II-, AT(1) receptor-, and TGF-beta-positive areas was achieved by NK3201 (a chymase inhibitor) treatment, and the neointima formation (intima/media: region A, 53 +/- 9%, P < 0.001; region B, 54 +/- 14%, P < 0.001) was also significantly suppressed in this group. Although lisinopril treatment also provided some beneficial effects with regard to the prevention of neointimal formation, the degree was less than that seen with chymase inhibition. These findings indicate that mast cell-derived chymase plays an essential role in the pathogenesis of the AV fistula access failure and that chymase inhibition may be a therapeutic target for the treatment of hemodialysis vascular access dysfunction in clinic settings.

Topics: Acetamides; Angiotensin II; Angiotensin II Type 1 Receptor Blockers; Animals; Arteriovenous Shunt, Surgical; Brachial Artery; Chymases; Constriction, Pathologic; Dogs; Hyperplasia; Mast Cells; Protease Inhibitors; Pyrimidines; Receptor, Angiotensin, Type 1; Serine Endopeptidases; Transforming Growth Factor beta; Tunica Intima; Tunica Media; Veins

The role of transforming growth factor (TGF)-beta and its signal in atherogenesis is not fully understood. Here, we examined mice lacking Smad3, a major downstream mediator of TGF-beta, to clarify the precise role of Smad3-dependent signaling in vascular response to injury. Femoral arteries were injured in wild-type and Smad3-null (null) male mice on C57Bl/6 background. Histopathological evaluation of the arteries 1 to 3 weeks after the injury revealed significant enhancement of neointimal hyperplasia in null compared with wild-type mice. Transplantation of null bone marrow to wild-type mice did not enhance neointimal thickening, suggesting that vascular cells in situ play a major role in the response. Null intima contained more proliferating smooth muscle cells (SMC) with less amount of collagen compared with wild-type intima. TGF-beta caused significant inhibition of cellular proliferation in wild-type aortic SMC, whereas the growth of null SMC was only weakly inhibited by TGF-beta in vitro, indicating a crucial role of Smad3 in the growth inhibitory function. On the other hand, Smad3-deficiency did not attenuate chemotaxis of SMC toward TGF-beta. TGF-beta increased transcript level of alpha2 type I collagen and tissue inhibitor of metalloproteinases-1, and suppressed expression and activity of matrix metalloproteinases in wild-type SMC. However, these effects of TGF-beta were diminished in null SMC. Our findings altogether show that the loss of Smad3 pathway causes enhanced neointimal hyperplasia on injury through modulation of growth and matrix regulation in vascular SMC. These results indicate a vasculoprotective role of endogenous Smad3 in response to injury.

Topics: Animals; Arteriosclerosis; Bone Marrow Transplantation; Cell Proliferation; Cells, Cultured; Collagen Type I; DNA-Binding Proteins; Hyperplasia; Matrix Metalloproteinases, Membrane-Associated; Metalloendopeptidases; Mice; Muscle, Smooth, Vascular; RNA, Messenger; Signal Transduction; Smad3 Protein; Tissue Inhibitor of Metalloproteinase-1; Trans-Activators; Transforming Growth Factor beta; Tunica Intima

Autogenous vein grafts are commonly used for arterial reconstructive procedures. Their success is limited by the development of intimal hyperplasia (IH), a fibroproliferative disease that predisposes the grafts to occlusive stenosis. Mesenchymal cell proliferation and the deposition of an extracellular matrix characterize neointimal development. Increasing evidence suggests that, regardless of blood vessel type, IH results from complex interactions among vessel wall cells, infiltrating leukocytes, and cytokines. Transforming growth factor-beta1 (TGF-beta1) is a pleiotropic cytokine with powerful effects on inflammatory cell chemotaxis; smooth muscle cell, fibroblast, and endothelial cell proliferation; and extracellular matrix synthesis.. Epigastric vein to common femoral artery interposition grafts were placed in male Lewis rats and harvested at 1, 2, 4, and 12 weeks after surgery. We used replication-defective adenoviruses to deliver a control reporter gene for the enzyme beta-galactosidase (Ad-GAL), empty virus (Ad-CMVpLpA), or the sequence encoding the antisense strand of TGF-beta1 (Ad-AST). The vein graft was transduced passively in medium containing 10 7 plaque-forming units per milliliter of Ad-GAL, Ad-CMVpLpA, or Ad-AST for 20 minutes at room temperature. The adenovirus-treated grafts were compared with grafts treated with medium without virus (sham).. The Ad-GAL control grafts showed beta-galactosidase activity from 3 days to 4 weeks. Twenty percent of cells were positive out to 2 weeks, at which time the number of cells positive for beta-galactosidase activity began to decline. Treatment with Ad-AST resulted in a significant reduction vs sham, Ad-CMVpLpA, and Ad-GAL in TGF-beta1 messenger RNA, total TGF-beta1 protein, and bioactive TGF-beta1 protein. Neointimal area was significantly reduced in the Ad-AST group vs Ad-GAL at 4 weeks, vs Ad-CMVpLpA at 4 and 12 weeks, and vs sham at 2 and 4 weeks. The medial/adventitial layer was significantly thicker in the Ad-AST group than the Ad-GAL group at 12 weeks. In addition, we studied the effect of Ad-AST on monocyte chemotactic protein 1 (MCP-1). Although the reduction in TGF-beta1 resulted in a reduction of MCP-1 messenger RNA in whole-graft homogenates and MCP-1 protein-positive staining in histologic sections from the perianastomotic region, no reduction in the number of ED1-positive cells (monocytes and macrophages) was observed.. Perioperative antisense TGF-beta1 treatment of the vein to be used in arterial reconstructions resulted in a prolonged diminution of IH; this emphasizes the importance of TGF-beta1 in neointimal thickening and indicates that ex vivo gene therapy can reduce the vessel's predisposition to IH.. The main cause of occlusion and graft failure after peripheral and cardiac arterial reconstruction is IH. The study of the mechanisms and mediators of IH, including TGF-beta1, should lead to future gene therapies to prevent or limit IH. The clinical effect of such treatments would be enormous, because they would increase graft longevity, thereby enhancing quality of life and enabling patients to live without the threat of limb loss or recurrent heart attack.

Topics: Adenoviridae; Animals; Chemokine CCL2; Extracellular Matrix; Gene Transfer Techniques; Graft Occlusion, Vascular; Hyperplasia; Immunohistochemistry; Rats; RNA, Antisense; Transforming Growth Factor beta; Tunica Intima; Veins; Wound Healing

Lack of endothelialization and abnormal smooth muscle cell (SMC) growth adversely affect the outcome of vascular synthetic grafts. The aims of our study were to investigate how a coating of extracellular matrix (ECM) and vascular endothelial growth factor (VEGF) might affect the endothelialization rate, smooth muscle cells (SMC) proliferation, and myointimal hyperplasia in experimental arterial ePTFE grafts.. In each of 30 male Lewis rats, a 1-cm-long ePTFE graft was inserted at the level of the abdominal aorta. Animals were randomized in five groups (six animals each): groups A and A1 received ePTFE grafts coated with a synthetic extracellular matrix (growth factor-reduced matrigel) containing VEGF; groups B and B1 received ePTFE grafts coated with synthetic ECM; and group C received ePTFE grafts alone. The grafts were explanted at 30 days from surgery for immunohistochemical analysis.. Both endothelialization rate and myointimal hyperplasia were augmented in group A versus groups B and C, and these findings were statistically significant. SMC density resulted significantly higher in group A versus groups B and C, and this was associated with an altered expression of bFGF and TGFbeta.. Pretreating ePTFE grafts with synthetic ECM and VEGF results in better endothelialization, but also in undesired higher SMC density and myointimal hyperplasia.

Topics: Animals; Aorta, Abdominal; Blood Vessel Prosthesis; Cell Count; Cell Proliferation; Coated Materials, Biocompatible; Endothelium, Vascular; Extracellular Matrix; Fibroblast Growth Factor 2; Hyperplasia; Male; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; Polytetrafluoroethylene; Proliferating Cell Nuclear Antigen; Rats; Rats, Inbred Lew; Transforming Growth Factor beta; Tunica Intima; Vascular Endothelial Growth Factor A

Aromatase transgenic mice exhibit hyperplastic and dysplastic changes, attesting to the importance of local estrogen in breast carcinogenesis. These mice also show increased levels of the estrogen receptor alpha and beta (ERalpha, ERbeta) suggesting that this receptor may play an important role in the initiation of estrogen-mediated mammary hyperplasia observed in these mice. To address the specific role of ERalpha in the mammary development and in the induction of estrogen-mediated hyperplasia in aromatase transgenic mice, we have generated MMTV-aromatase x ERalpha knockout cross (referred as aromatase/ERKO). Even though ERbeta is expressed in aromatase/ERKO mice, lack of ERalpha leads to impaired mammary growth in these mice. The data suggest that ERalpha plays an important role in the mammary gland development as well as in the induction of mammary hyperplasia in aromatase transgenic mice. Lack of ERalpha expression in the aromatase/ERKO mice resulted in a decrease in the expression of Cyclin D1, PCNA and TGFbeta relative to the aromatase parental strain. The studies involving aromatase/ERKO mice show that lack of ERalpha results in impaired mammary development even in the presence of continuous tissue estrogen, suggesting estrogen/ERalpha-mediated actions are critical for mammary development and carcinogenesis.

Topics: Animals; Aromatase; Cyclin D1; Estrogen Receptor alpha; Hyperplasia; Male; Mammary Glands, Animal; Mice; Mice, Knockout; Mice, Transgenic; Proliferating Cell Nuclear Antigen; RNA, Messenger; Transforming Growth Factor beta

The murine myostatin mutation Mstn(Cmpt-dl1Abc) (Compact; C) was introduced into an inbred mouse line with extreme growth (DUHi) by marker-assisted introgression. To study the allelic effects on muscle fibre hyperplasia and hypertrophy, myonuclear proliferation, protein accretion, capillary density, and muscle fibre metabolism, samples from M. rectus femoris (RF) and M. longissimus dorsi (LD) muscles of animals wild-type (+/+), heterozygous (C/+), and homozygous (C/C) for the Mstn(Cmpt-dl1Abc) allele were examined by histological and biochemical analyses. Homozygous C/C mice exhibited lower body (-12%) but higher muscle weights (+38%) than ++ mice. Total muscle fibre number was increased (+24%), whereas fibre size was not significantly affected. Protein and DNA concentrations and DNA:protein ratios as well as specific CK activity remained unchanged for higher mass muscle implying increases in the total contents of DNA and muscle specific protein. Fibre type distribution was markedly shifted to the white glycolytic muscle fibres (+16-17% units) at the expense of red oxidative fibres. Capillary density was substantially lower in C/C than in ++ mice as seen by lower number of capillaries per fibre (-35%) and larger fibre area per capillary (+77%). However, the Mstn(Cmpt-dl1Abc) allele was partially recessive in heterozygous C/+ mice for both fibre type frequencies and capillary density. The results show that hypermuscularity caused by mutations in the myostatin gene results from muscle fibre hyperplasia rather than hypertrophy, and from balanced increases in myonuclear proliferation and protein accretion. However, capillary supply is adversely affected and muscle metabolism shifted towards glycolysis, which could have negative consequences for physical fitness.

Topics: Alleles; Animals; Body Composition; Body Weights and Measures; Capillaries; Crosses, Genetic; Female; Hyperplasia; Hypertrophy; In Vitro Techniques; Male; Mice; Mice, Transgenic; Muscle Fibers, Skeletal; Muscle, Skeletal; Mutation; Myostatin; Organ Size; Quadriceps Muscle; Transforming Growth Factor beta

Myostatin is a member of the transforming growth factor-beta (TGF-beta) family that functions as a negative regulator of skeletal muscle development and growth. Recently, it has been reported that the transgenic zebrafish expressing myostatin prodomain exhibited an increased number of fiber in skeletal muscle. Other novel results suggest that myostatin plays a mayor role during myogenesis, apart from inhibition of proliferation as well as differentiation. We have investigated the ability of double-stranded RNA (dsRNA) to inhibit myostatin function in the zebrafish. By microinjection dsRNA, corresponding to biologically active C-terminal domain from aminoacid 268 to end codon of tilapia myostatin protein, we produced an increased body mass in treated fish. The dsRNA injection in early development stage in zebrafish produced hyperplasia or hypertrophy. In addition, the interference of gene function showed a strong dependence on the amount of dsRNA.

Topics: Animals; Body Weight; Gene Silencing; Hyperplasia; Hypertrophy; Muscle, Skeletal; Myostatin; Organ Size; Phenotype; RNA Interference; Transforming Growth Factor beta; Zebrafish; Zebrafish Proteins

Rapid bladder growth associated, partial urethral obstruction and embryonic bladder development entail stromal-epithelial interactions involving signaling by the cytokine transforming growth factor-beta (TGF-beta). However, to our knowledge the role of TGF-beta in bladder stromal hyperplasia and hypertrophy is not understood.. In an effort to understand the specific role of TGF-beta signaling in bladder stroma a fibroblast specific conditional knockout mouse of the type II TGF-beta receptor gene, Tgfbr2(/spko), was generated using Cre-lox methodology. Bladders from 18, 7 to 8-week-old mice were harvested for histological and immunohistochemical analysis.. Bladders from homozygous Tgfbr2(/spko), male mice showed marked hypertrophy in the lamina propria and smooth muscle layers in the absence of visible or functional bladder obstruction by age 8 weeks. However, age matched female mice of the same genotype maintained bladder architecture similar to that in wild-type littermate male and female controls. Immunohistochemistry for the phosphorylated form of Smad2 indicated a general loss in TGF-beta signaling in the lamina propria of bladders of male and female Tgfbr2(/spko), mice, and yet pronounced alpha-smooth muscle actin expression was noted in male Tgfbr2(/spko), bladders, which is a marker for myofibroblasts.. A sex disparity was observed in the Tgfbr2(/spko), mouse model lacking TGF-beta signaling in fibroblasts. Deletion of TGF-beta in males leads to a hypertrophied lamina propria and muscularis externa with myofibroblast differentiation and proliferation. Female homozygous Tgfbr2(/spko), bladders appeared the same as those of wild-type male and female controls. This model suggests a role for stromal TGF-beta signaling with estrogens and androgens in bladder fibrosis.

Topics: Animals; Female; Fibroblasts; Hyperplasia; Male; Mice; Mice, Knockout; Signal Transduction; Transforming Growth Factor beta; Urinary Bladder Diseases

To investigate the expression of early growth response gene-1 (Egr-1) and its correlative genes: platelet-derived growth factor-B (PDGF-B) gene and transforming growth factor-beta(1) (TGF-beta(1)) gene, in autogenous vein graft and the relationship of the expression of these genes to intimal hyperplasia (IH).. A segment of the right common jugular vein was transplanted to the infra-renal abdominal aorta of the same individual so as to establish an autogenous vein graft model in 90 Wistar rats. Vein samples were harvested 1, 2, 6, and 24 hours, 3 and 7 days, and 2, 4, and 6 weeks after the operation to undergo histological examination. In situ hybridization and RT-OCR were performed to examine the mRNA expression of Egr-1, PDGF-B, and TGF-beta(1). Western blotting and immunohistochemistry were used to detect the protein expression of Egr-1, PDGF-B, and TGF-beta(1). The contralateral common jugular vein was used as control.. Histological examination showed a great amount of vascular smooth muscle cells (VSMCs) in the tunica intima and tunica media of the vein graft since the 7 th day after the transplantation. mRNA expression of Egr-1, PDGF-B, and TGF-beta(1) was not found in the normal veins. The expression of Egr-1 mRNA in the vein graft showed a biphasic manner: increased remarkably 1 h after the transplantation, decreased during the period 6 h approximately 3 d after, re-increased 7 d after, and peaked 4 weeks after. The mRNA expression of PDGF-B appeared 6 h after, and peaked 2 weeks after, and began to decrease since the 4 th week. The mRNA expression of TGF-beta(1) increased 6 h after, peaked 7d after, and began to decrease 2 weeks after. The protein expression of Egr-1 appeared 2 h after, and peaked 4 weeks after; the protein expression of PDGF-B appeared 6 h after and peaked 4 weeks after; and the protein expression of TGF-beta(1). Appeared 24 h after and peaked 2 weeks after. Immunohistochemistry showed that Egr-1 protein expression was mainly in the VSMCs and monocytes/macrophages in the tunica media in the early stage after transplantation, and peaked 4 weeks later with a positive cell rate of 40% +/- 9%; in the early stage after transplantation PDGF-B protein expression was mainly in the VSMCs in the tunica media, and peaked 4 weeks after with a positive cell rate of 45% +/- 4%; the TGF-beta(1) protein expression peaked 2 weeks after with a positive cell rate of 41% +/- 7%; and 4 weeks after transplantation the protein expressions of Egr-1, PDGF-B, and TGF-beta(1) were all mainly in the VSMCs in the newly-grown tunica intima.. The IH of vein graft is affinitive with the expression of Egr-1, PDGF-B, and TGF-beta(1). The activation and expression of Egr-1, PDGF-B, and TGF-beta(1) depend on Egr-1 and may contribute to the high expression of Egr1 via a feedback mechanism.

Topics: Animals; Blotting, Western; Early Growth Response Protein 1; Female; Gene Expression; Graft Occlusion, Vascular; Hyperplasia; Immunohistochemistry; In Situ Hybridization; Male; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; Platelet-Derived Growth Factor; Rats; Rats, Wistar; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Transforming Growth Factor beta; Transplantation, Autologous; Tunica Intima; Veins

Modern immunosuppressive agents such as tacrolimus and rapamycin are claimed to be associated with a reduction in vascular narrowing, a central feature of chronic rejection. This study assesses the effect of cyclosporine, tacrolimus and rapamycin on the development of intimal thickening, fibrosis-associated genes and deposition of extracellular matrix (ECM) proteins in a model of intimal hyperplasia. Male Sprague-Dawley rats received either no treatment or 5 mg/kg cyclosporine, 0.1 mg/kg tacrolimus or 0.05 mg/kg rapamycin. Animals underwent left common carotid balloon angioplasty, and intima medial ratios, pro-fibrotic gene expression and ECM accumulation were calculated at 14 and 28 days. Cyclosporine was associated with increased intimal thickening compared to controls ( P < 0.004). Tacrolimus had no effect on intimal thickening, whilst rapamycin significantly inhibited intimal thickening at both 14 and 28 days ( P < 0.004 and P < 0.026, respectively). All groups significantly inhibited matrix metalloproteinase (MMP)-2, MMP-9, tissue inhibitor of metalloproteinases (TIMP)-1, transforming growth factor (TGF)-beta and collagen III expression at 14 days ( P < 0.001), but increased ECM deposition. However, rapamycin marginally reduced ECM deposition compared to cyclosporine ( P < 0.06). Treatment with cyclosporine was associated with worsening of vascular narrowing, whilst rapamycin showed a beneficial reduction in intimal thickening. Treatment with all immunosuppressive agents resulted in increased ECM deposition. Rapamycin may halt the progression of vascular narrowing compared to both cyclosporine and tacrolimus.

Topics: Angioplasty, Balloon; Animals; Carotid Artery, Common; Collagen Type III; Cyclosporine; Extracellular Matrix Proteins; Fibrosis; Gene Expression; Hyperplasia; Immunosuppressive Agents; Male; Matrix Metalloproteinase Inhibitors; Rats; Rats, Sprague-Dawley; Sirolimus; Tacrolimus; Tissue Inhibitor of Metalloproteinases; Transforming Growth Factor beta; Tunica Intima

Transforming growth factor (TGF)-beta(1) is a pleiotropic growth factor with known inhibitory effects on immune cell activation. However, the specific mechanism(s) and in vivo significance of the effectors of TGF-beta(1) modulation in the context of vascular inflammation are not well characterized. The chemokine monocyte chemoattractant protein (MCP)-1 is critical for the recruitment of macrophages in inflammatory disease states. In this study, we provide definitive evidence that the ability of TGF-beta(1) to inhibit MCP-1 expression is mediated via its effector Smad3. Adenoviral overexpression of Smad3 potently repressed inducible expression of endogenous MCP-1. Conversely, TGF-beta(1) inhibition of cytokine-mediated induction of MCP-1 expression was completely blocked in Smad3-deficient macrophages. Consistent with this impaired response, cardiac allografts in Smad3-deficient mice developed accelerated intimal hyperplasia with increased infiltration of adventitial macrophages expressing MCP-1. Previous studies show that MCP-1 inducibility is regulated by an AP-1 complex composed of c-Jun/c-Fos heterodimers. We demonstrate that the inhibitory effect of Smad3 occurs via a novel antagonistic effect of Smad3 on AP-1 DNA-protein binding and activity. Thus, Smad3 plays an essential role in modulating vascular inflammation characteristic of transplant-associated arteriopathy, is important in regulating MCP-1 expression, and plays a critical role in the ability of TGF-beta(1) to repress stimuli from a major inflammatory signaling pathway.

Topics: Animals; Cell Line; Chemokine CCL2; DNA-Binding Proteins; Gene Expression Regulation; Heart Transplantation; Hyperplasia; Macrophage Activation; Macrophages; Mice; Mice, Inbred C57BL; Mice, Knockout; Postoperative Complications; Protein Biosynthesis; Proteins; Recombinant Fusion Proteins; Smad3 Protein; Trans-Activators; Transcription Factor AP-1; Transcription, Genetic; Transfection; Transforming Growth Factor beta; Transforming Growth Factor beta1; Transplantation, Homologous; Tunica Intima; Vasculitis

Inflammatory infiltrates, airway hyper-responsiveness, goblet cell hyperplasia and subepithelial thickening are characteristic of chronic asthma. Current animal models of allergen-induced airway inflammation generally concentrate on the acute inflammation following allergen exposure and fail to mimic all of these features.. The aim of this study was to use a murine model of prolonged allergen-induced airway inflammation in order to characterize the cells and molecules involved in the ensuing airway remodelling. Moreover, we investigated whether remodelling persists in the absence of continued allergen challenge.. Acute pulmonary eosinophilia and airways hyper-reactivity were induced after six serial allergen challenges in sensitized mice (acute phase). Mice were subsequently challenged three times a week with ovalbumin (OVA) (chronic phase) up to day 55. To investigate the persistence of pathology, one group of mice were left for another 4 weeks without further allergen challenge (day 80).. The extended OVA challenge protocol caused significant airway remodelling, which was absent in the acute phase. Specifically, remodelling was characterized by deposition of collagen as well as airway smooth muscle and goblet cell hyperplasia. Importantly, these airway changes, together with tissue eosinophilia were sustained in the absence of further allergen challenge. Examination of cytokines revealed a dramatic up-regulation of IL-4 and tumour growth factor-beta1 during the chronic phase. Interestingly, while IL-4 levels were significantly increased during the chronic phase, levels of IL-13 fell. Levels of the Th1-associated cytokine IFN-gamma also increased during the chronic phase.. In conclusion, we have demonstrated that prolonged allergen challenge results in persistent airway wall remodelling.

Topics: Acute Disease; Allergens; Animals; Asthma; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Chronic Disease; Collagen; Female; Goblet Cells; Hyperplasia; Interferon-gamma; Interleukin-13; Interleukin-4; Mice; Mice, Inbred BALB C; Models, Animal; Muscle, Smooth; Ovalbumin; Pulmonary Eosinophilia; Respiratory System; Time Factors; Transforming Growth Factor beta

Transforming growth factor-beta1 (TGFbeta1) is a multifunctional signalling molecule with a wide array of roles. Animal experiments suggest that TGFbeta1 plays a biphasic role in carcinogenesis by protecting against the early formation of benign epithelial growths, but promoting malignant transformation of those growths that do develop. A polymorphism in the signal peptide sequence of the TGFbeta1 gene (L10P) has been associated with increased levels of plasma TGFbeta1 in individuals with the P allele.. We investigated whether this polymorphism was associated with the risk of colorectal adenomatous or hyperplastic polyps in a case-control study of individuals from Minnesota. Risk of colorectal polyps was evaluated separately for individuals with adenomatous polyps (n = 513) and hyperplastic polyps (n = 191) relative to polyp-free controls (n = 606) using logistic regression analysis.. No overall association was seen between the L10P polymorphism and risk of colorectal adenomatous polyps. The age- and sex-adjusted odds ratios (OR) of developing colorectal hyperplastic polyps were 1.0 (95% CI: 0.7, 1.4) and 0.7 (95% CI: 0.4, 1.1) for individuals with the LP and PP genotypes, respectively, compared with individuals with the LL genotype. When stratified by smoking, evidence for a decreased risk of hyperplastic polyps associated with the P allele was seen only among ever smokers (P for trend = 0.05).. Whereas adenoma risk did not vary by TGFbeta1 L10P genotype, these results suggest that the L10P variant allele may have a protective role in the development of colorectal hyperplastic polyps, possibly consistent with its role as an inhibitor of epithelial growths.

Topics: Adenomatous Polyps; Adult; Age Distribution; Aged; Case-Control Studies; Colonic Polyps; Colorectal Neoplasms; Female; Genetic Predisposition to Disease; Genotype; Humans; Hyperplasia; Male; Middle Aged; Polymorphism, Genetic; Risk Factors; Smoking; Transforming Growth Factor beta; Transforming Growth Factor beta1

We previously showed that treatment with liposomally encapsulated dichloromethylene bisphosphonate reduces intimal hyperplasia development and macrophage accumulation in a rat epigastric vein to femoral artery model of intimal hyperplasia. Our objective in this study was to determine the effect of liposomally encapsulated dichloromethylene bisphosphonate on the expression of two cytokines essential to neointimal development, monocyte chemotactic protein-1 (MCP-1) and transforming growth factor-beta1 (TGF-beta).. We injected rats both 2 days preoperatively and 2 weeks postoperatively with liposomally encapsulated dichloromethylene bisphosphonate (Lip-Clod), liposomally encapsulated phosphate-buffered saline solution (Vector), or phosphate-buffered saline solution (PBS), and harvested the grafts at 1 and 4 weeks. In the perianastomotic region, MCP-1 and TGF-beta protein expression in the total graft cross-section and in the neointima was determined with immunohistochemistry. In whole-graft lysates, MCP-1 and TGF-beta protein were determined with an enzyme-linked immunosorbent assay, and messenger RNA expression was determined with reverse transcription quantitative polymerase chain reaction.. Lip-Clod treatment reduced intimal hyperplasia when compared with Vector or PBS treatment. These reductions were significant (P<.05) at both time points. When compared with the PBS treatment, at 1 week but not at 4 weeks Lip-Clod reduced both MCP-1 and TGF-beta protein (P< or =.01 and P< or =.006) in the perianastomotic region of vein grafts. In whole-graft lysates, no significant difference was seen in MCP-1 protein at either time point; however, TGF-beta protein expression was significantly reduced at both 1 and 4 weeks (P=.02 and P=.004). Message analysis in whole-graft lysates at 1 week showed that MCP-1 message expression increased in the Lip-Clod group compared with the PBS group (P=.02), but no significant differences among groups for TGF-beta message levels. Results with Vector were often intermediate to results with Lip-Clod and PBS.. The major effect of Lip-Clod treatment on TGF-beta and MCP-1 protein levels in the perianastomotic region is observed at 1 week, and macrophage depletion with Lip-Clod inhibits graft neointimal hyperplasia and TGF-beta protein expression in whole-graft lysates at 1 and 4 weeks. These results support the concept that the infiltrating macrophages contribute a significant portion of the cytokines that facilitate intimal hyperplasia and that reducing these cytokines early after grafting influences the development of intimal hyperplasia at later time points.. All vascular surgeons have patients who have undergone a technically satisfying vein graft, only to have the bypass fail during the first year due to perianastomotic intimal hyperplasia (IH). We hypothesize that vein graft IH is analogous to aberrant wound healing. Central to wound healing is the recruitment of macrophages with their cytokines. This work raises the question whether clinical strategies designed to either decrease macrophages or the cytokines released by macrophages at the time of vein graft placement will be efficacious for limiting the development of IH.

Topics: Animals; Antimetabolites; Blood Vessel Prosthesis; Chemokine CCL2; Clodronic Acid; Hyperplasia; Immunosuppression Therapy; Liposomes; Macrophages; Male; Models, Animal; Rats; Rats, Inbred Lew; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tunica Intima; Wound Healing

Pericytes (PCs) are smooth muscle-like mural cells of capillaries and venules, which can synthesize matrix components and fibroblast-activating cytokines, and are thus potential mediators of pathological changes in scleroderma. In this study, alterations in microvessels were quantitatively imaged, taking PC into account for the first time.. Skin biopsies from systemic (12) and localized (14) scleroderma forms as well as age-, sex-, and body location-matched controls were examined with respect to capillary and venular densities as well as endothelial cell (EC) and PC counts using a newly developed (in respect of PC and EC) indirect collagen IV immunostaining-based method.. Hyperplasia of the PC that doubled the microvascular PC density was the most conspicuous characteristic. In the capillaries of the upper dermal plexus of the periphery of the sclerotic zones, median ratios of PC : EC were 0.23 (controls 0.10) or 0.18 (controls 0.11) in systemic or localized scleroderma, respectively. Furthermore, an increase in capillary density in the upper dermal plexus could be demonstrated in the marginal zones of both types of disease.. The observed PC increase in the peripheral zones of active disease supports the hypothesis of a vascular pathogenesis of scleroderma and directs the focus to microvascular PC.

Topics: Adolescent; Adult; Aged; Capillaries; Cell Count; Collagen Type IV; Endothelial Cells; Female; Humans; Hyperplasia; Immunohistochemistry; Male; Middle Aged; Pericytes; Scleroderma, Localized; Scleroderma, Systemic; Skin; Transforming Growth Factor beta

In the present study, we show that transforming growth factor beta1 (TGF-beta1) was frequently overexpressed in human head and neck squamous cell carcinomas (HNSCCs) and adjacent tissues in comparison with normal head and neck tissues. To determine the role of TGF-beta1 overexpression in HNSCC carcinogenesis, we generated transgenic mice in which TGF-beta1 transgene expression can be induced in head and neck epithelia. TGF-beta1 transgene induction in head and neck epithelia, at levels similar to those in human HNSCCs, caused severe inflammation and angiogenesis. Consequently, TGF-beta1-transgenic epithelia exhibited hyperproliferation. These phenotypes correlated with enhanced Smad signaling in transgenic epithelia and stroma. Our study suggests that TGF-beta1 overexpression at early stages of HNSCC formation provides a tumor promoting microenvironment.

Topics: Animals; Carcinoma, Squamous Cell; Cell Division; Cell Transformation, Neoplastic; Epithelial Cells; Head and Neck Neoplasms; Humans; Hyperplasia; Inflammation; Mice; Mice, Transgenic; Mouth; Mouth Mucosa; Neovascularization, Pathologic; Oropharynx; Transforming Growth Factor beta; Transforming Growth Factor beta1

SPARC (Secreted Protein Acidic and Rich in Cysteine) is a multifunctional glycoprotein belonging to a group of matrix-associated factors that mediate cell-extracellular matrix interactions but have no structural roles. In the present study we investigated the contribution of SPARC to factors that influence the development of skin tumors in response to UV irradiation. A hairless SPARC-null mouse was developed and compared to control SKH1 hairless mice in terms of skin tumor induction and extracellular matrix changes occurring in response to UV-irradiation. Following 23 weeks of exposure to UVB totaling 14.5 J per cm(2), tumor development in the wild-type mice was severe, with an average of over 20 tumors per mouse, many of which were squamous cell carcinomas. Conversely, the SPARC-null mice were strikingly tumor-resistant, developing no squamous cell carcinomas and averaging less than one small papilloma per mouse. SPARC was undetectable immunohistochemically in skin from the non-irradiated control group yet was present in relatively high quantities in the basal and superficial areas of the tumor mass. The SPARC-null mice also exhibited a limited contact hypersensitivity response and were refractory to UV induced immune suppression. In conclusion, SPARC appears to have a crucial role in mediating tumor formation in response to UV irradiation.

Topics: Animals; Carcinoma, Squamous Cell; Dermatitis, Contact; Extracellular Matrix Proteins; Female; Hyperplasia; Male; Mice; Mice, Hairless; Mice, Inbred C57BL; Mice, Mutant Strains; Neoplasms, Radiation-Induced; Neovascularization, Pathologic; Osteonectin; Skin; Skin Neoplasms; Transforming Growth Factor beta; Transforming Growth Factor beta1; Ultraviolet Rays

The development of fibromuscular intimal hyperplasia and subsequent graft failure remains an urgent problem in cardiac surgery. Transforming growth factor-beta1 (TGF-beta1) is involved in the pathogenesis of arteriosclerosis through induction of extracellular matrix proteins. We tested the hypothesis that intimal hyperplasia is already present in human saphenous veins and left internal mammary arteries before coronary artery bypass surgery and is associated with an increased expression of TGF-beta1.. Forty-six segments of saphenous veins and 27 of left internal mammary arteries were collected from 50 patients undergoing coronary artery bypass surgery. Morphometric analysis was performed by microscopic computer analysis. Immunohistochemistry was performed with antibodies directed against TGF-beta1, its latent binding protein (LTBP-1) and its type 2 receptor (RII).. The incidence of intimal hyperplasia was significantly higher in saphenous veins (67.4%) than in mammary arteries (29.6%; p < 0.05). Saphenous veins and mammary arteries with intimal hyperplasia expressed more TGF-beta1 (endothelial and intimal layers) and LTBP-1 (intimal and medial layers) when compared with corresponding vessels without hyperplasia (both groups p < 0.05). Endothelial and intimal RII expression was significantly higher in saphenous veins with intimal hyperplasia as compared with saphenous veins without hyperplasia (p < 0.05). Transforming growth factor-beta1 staining in the intima correlated with the presence of an intimal hyperplasia in saphenous veins (rho = 0.317) and mammary arteries (rho = 0.428).. Local TGF-beta1 expression is associated with the presence of intimal hyperplasia in the examined vessels. Preexisting intimal hyperplasia is more prevalent and serious in saphenous veins than in left internal mammary arteries, giving further explanation to the superior long-term results of left internal mammary grafts.

Topics: Aged; Arteriosclerosis; Coronary Artery Bypass; Female; Humans; Hyperplasia; Internal Mammary-Coronary Artery Anastomosis; Intracellular Signaling Peptides and Proteins; Latent TGF-beta Binding Proteins; Male; Mammary Arteries; Middle Aged; Organ Specificity; Prospective Studies; Protein Serine-Threonine Kinases; Receptor, Transforming Growth Factor-beta Type II; Receptors, Transforming Growth Factor beta; Saphenous Vein; Time Factors; Tissue and Organ Harvesting; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tunica Intima; Tunica Media

Pulmonary hypertension (PH) is a progressive disease characterized by raised pulmonary vascular resistance, thought to be curable only through lung transplantation. Pathophysiologically, proliferation of pulmonary artery smooth muscle cells triggers pulmonary arterial stenosis and/or regurgitation, especially in advanced PH.. Using a rat model of advanced pulmonary vascular disease produced by injecting monocrotaline, we show that hepatocyte growth factor (HGF) targets pulmonary arterioles and blocks the progression of PH. In these rats, endogenous HGF production was dramatically downregulated during developing experimental PH, but c-Met/HGF receptor was abundant in the medial layers of pulmonary arterioles. HGF gene transfection 2 weeks after the monocrotaline injection resulted in milder medial hyperplasia in lung arterioles and inhibited overgrowth of pulmonary artery smooth muscle cells. Notably, exogenous HGF reduced lung expression levels of endothelin-1 and transforming growth factor-beta, which are critically involved in PH-linked fibrogenic events. Overall, medial wall thickening of pulmonary arteries was almost completely prevented by HGF, and the total collagen deposition in the lung decreased; both effects contributed to the suppression of pulmonary artery hypertension.. Our results suggest that the loss of endogenous HGF may be a feature of the pathogenesis of PH and that HGF supplementation may minimize pathological lung conditions, even advanced PH.

Topics: Animals; Arterioles; Collagen; Endothelin-1; Extracellular Matrix; Fibrosis; Genetic Therapy; Hepatocyte Growth Factor; Humans; Hyperplasia; Hypertension, Pulmonary; Male; Monocrotaline; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; Proto-Oncogene Proteins c-met; Rats; Rats, Wistar; Transfection; Transforming Growth Factor beta; Tunica Media

Asthma is associated with recruitment of eosinophils, accumulation of chronic inflammatory cells in the airway walls, subepithelial fibrosis and other structural changes of airway wall remodelling. The role of ongoing exposure to allergens in their pathogenesis remains unclear.. To examine whether changes of inflammation and remodelling were reversible following cessation of antigenic challenge in a mouse model of chronic asthma.. BALB/c mice sensitized to ovalbumin (OVA) were chronically challenged by inhalation of a low mass concentration of antigen for 8 weeks, leading to development of acute-on-chronic airway inflammation, subepithelial fibrosis and other changes of airway wall remodelling. Epithelial injury was assessed by immunohistochemistry, while inflammation and remodelling were quantified by appropriate histomorphometric techniques. Regression of lesions was assessed in animals examined at 1, 2 and 4 weeks after exposure to OVA ceased.. We did not find evidence of airway epithelial injury in this model of low-level chronic inhalational exposure to antigen. Persistence of the recruitment of eosinophils and chronic inflammatory cells in the airway walls was dependent on continuing antigenic challenge, as was persistence of mucous cell hyperplasia/metaplasia. Subepithelial fibrosis and epithelial hypertrophy exhibited delayed reversibility following cessation of exposure to antigen, possibly related to matrix-associated accumulation of transforming growth factor-beta(1).. In chronic asthma, low-level antigenic challenge may be required to maintain the inflammatory response in the airway wall, but airway remodelling may persist in its absence.

Topics: Administration, Inhalation; Allergens; Animals; Asthma; Chronic Disease; Disease Models, Animal; Eosinophils; Epithelial Cells; Female; Fibrosis; Hyperplasia; Mice; Mice, Inbred BALB C; Ovalbumin; Remission, Spontaneous; Respiratory Mucosa; Trachea; Transforming Growth Factor beta; Transforming Growth Factor beta1

We have shown that pituitary vasoactive intestinal peptide (VIP) mediates the effects of estrogen on lactotrope hyperplasia, angiogenesis and hyperprolactinemia, and reduces the pituitary content of transforming growth factor beta beta1 (TGF-beta1, an inhibitor of lactotrope proliferation). Dopamine agonists reverse lactotrope hyperplasia and hyperprolactinemia and also reduce the pituitary VIP content in hyperestrogenized rats. To elucidate the interaction of bromocriptine (BC) and pituitary VIP, a VIP receptor antagonist (VA), BC, or both drugs were administered for 5 days to F344 rats treated with diethylstilbestrol (DES). Both BC and VA similarly blocked the effects of DES on pituitary weight and pituitary content of prolactin (PRL), proliferating cell nuclear antigen, and vascular endothelial growth factor, without evidence of synergism. The estrogen effect on pituitary TGF-beta1 was completely inhibited by VA, but only partially by BC. On the contrary, serum PRL was close to the normal levels in the BC group 2 h after the first dose, while VA only reduced serum PRL after 5 days. DES increased VIP and VIP mRNA levels specifically at the pituitary, this effect being partially blocked by BC. These data suggest that the dopamine agonists inhibit lactotrope proliferation and angiogenesis by blocking the autocrine/paracrine action of VIP. On the other hand, the dopamine agonists inhibit the estrogen-induced hyperprolactinemia by acting through different pathways than those implicated in the proliferative process.

Topics: Animals; Blotting, Northern; Blotting, Western; Brain; Bromocriptine; Diethylstilbestrol; Dopamine; Dopamine Agonists; Estrogens, Non-Steroidal; Female; Hyperplasia; Pituitary Gland; Prolactin; Rats; Rats, Inbred F344; Receptors, Vasoactive Intestinal Peptide; RNA, Messenger; Transforming Growth Factor beta; Vascular Endothelial Growth Factor A; Vasoactive Intestinal Peptide

The purpose of this study was to evaluate the relationship between intimal hyperplasia and transforming growth factor-beta1 (TGF-beta1) mRNA expression in synthetic arterial grafts and also to clarify the effect of angiotensin-converting enzyme inhibitor (ACEI) on perianastomotic intimal hyperplasia and TGF-beta1 mRNA expression. Thirty New Zealand White rabbits were randomly divided into two groups (15 each); one group was administered Captopril 10 mg/kg/day per os as an ACE inhibitor, and the other group received on saline as a vehicle from 7 days prior to operation until the graft was harvest (1, 8, or 14 weeks). A 10-mm segment of an expanded polytetrafluoroethylene graft (3 mm in diameter) was implanted in the right common carotid artery of the rabbits; 15 rabbits had by-pass grafting alone (Graft Alone group) and the other 15 rabbits had by-pass graft along with the ACEI (Graft plus ACEI group). The artery grafts were harvested. The intima to media height ratio (IMHR) and the TGF-beta1 mRNA expression level in perianastomotic graft tissue by reverse transcription-polymerase chain reaction (RT-PCR). The IMHRs gradually increased from 1 to 14 weeks in both groups (vs. 1 wk in each group, p<0.05). The IMHRs of the Graft plus ACEI group were comparable to those of Graft Alone group at 1 week, but significantly lower at 8 and 14 weeks (vs. Graft Alone group, p<0.05). The TGF-beta1 mRNA expression levels of the Graft plus ACEI group were clearly lower than those of the Graft Alone group at 1 and 8 weeks (vs. Graft Alone group, p<0.05), but similar at 14 weeks. TGF-beta1 in the synthetic artery graft of the Graft Alone group was up-regulated as early as 1 week after the operation, when no definitive development of a quantifiable neointima was observed. The TGF-beta1 mRNA expression of the Graft Alone group was highest at 8 weeks and lowest at 14 weeks (vs. 1 week, *p<0.05), but such time-dependent changes were not observed in the Graft plus ACEI group. The results indicated that ACEI reduced intimal hyperplasia in the grafts of the Graft plus ACEI group and also suppressed TGF-beta1 mRNA expression in perianastomotic intimal hyperplasia tissues to the normal artery level. Perianastomotic intimal hyperplasia in synthetic arterial graft is considered to be related to TGF-beta1, the expression of which is locally mediated by angiotensin II and, therefore, suppressed by ACEI.

Topics: Anastomosis, Surgical; Angiotensin II; Angiotensin-Converting Enzyme Inhibitors; Animals; Blood Vessel Prosthesis Implantation; Captopril; Carotid Artery, Common; Hyperplasia; Polytetrafluoroethylene; Rabbits; RNA, Messenger; Time Factors; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tunica Intima

Serrated adenomas, hyperplastic polyps, and admixed hyperplastic/adenomatous polyps form a distinct group of colorectal tumors, the molecular genetic basis of which is still poorly understood. We describe a novel mouse model for serrated adenomas and mixed polyposis, here referred to as Sad (serrated adenomas), caused by a spontaneously risen splice site mutation in the murine Smad4 gene. The Sad chromosomal region was identified by genetic linkage and loss of heterozygosity (LOH) analysis. Subsequently, several candidate genes were investigated by expression and mutation analysis. By use of genetic linkage and LOH analysis, we mapped the Sad candidate to mouse chromosome 18, 44-48 cM, syntenic to human chromosome band 18q21. Within this chromosomal interval, the Smad2, Smad4, and Smad7 genes were analyzed for the presence of a disease-causing mutation in affected animals. A single nucleotide (nt) deletion was identified in the intron 5/exon 6 splice acceptor site of the Smad4 gene. The single base deletion results in a frameshift and an early termination codon through activation of a cryptic splice site 4 nt downstream in exon 6. The resulting mRNA is unstable, and the Sad mutation is thus likely to represent a null allele. Identification of a Smad4 mutation in the Sad mouse model provides further support for the involvement of the Smad genes, and thus the TGFB pathway, in the serrated/hyperplastic route to colorectal cancer.

Topics: Adenomatous Polyps; Animals; Cell Line; Colonic Polyps; Colorectal Neoplasms; Disease Models, Animal; DNA-Binding Proteins; Female; Fetal Death; Gene Expression Profiling; Genes, Lethal; Homozygote; Hyperplasia; Loss of Heterozygosity; Male; Mice; Mice, Inbred C57BL; Mice, Inbred Strains; RNA Splice Sites; Sequence Deletion; Signal Transduction; Smad4 Protein; Trans-Activators; Transforming Growth Factor beta

We investigated the comparative roles of mitogen-activated protein (MAP) kinases, including c-Jun NH2-terminal kinase (JNK), extracellular signal-regulated kinase (ERK), and p38, in vascular smooth muscle cell (VSMC) proliferation, migration, and gene expression.. VSMCs were infected with recombinant adenovirus containing dominant-negative mutants of ERK, p38, and JNK (Ad-DN-ERK, Ad-DN-p38, and Ad-DN-JNK, respectively) to specifically inhibit the respective MAP kinases and then stimulated with platelet-derived growth factor (PDGF)-BB. Ad-DN-ERK attenuated PDGF-BB-induced VSMC proliferation more potently than Ad-DN-p38 or Ad-DN-JNK, indicating the dominant role of ERK in VSMC proliferation. Ad-DN-ERK, Ad-DN-p38, and Ad-DN-JNK similarly inhibited PDGF-induced VSMC migration. Ad-DN-ERK and Ad-DN-JNK suppressed PDGF-BB-induced downregulation of cyclin-dependent kinase inhibitor p27Kip1, whereas Ad-DN-p38 decreased PDGF-BB-induced upregulation of p21Cip1. Ad-DN-ERK inhibited PDGF-BB-induced plasminogen activator inhibitor type-1 (PAI-1), monocyte chemoattractant protein-1, and transforming growth factor-beta1 expressions, Ad-DN-p38 blocked monocyte chemoattractant protein-1 and transforming growth factor-beta1 expression but not PAI-1, whereas Ad-DN-JNK suppressed only PAI-1 expression. Moreover, in vivo gene transfer of Ad-DN-p38 to rat carotid artery caused the inhibition of intimal hyperplasia by balloon injury, indicating the involvement of p38 in vascular remodeling in vivo.. We propose that these 3 MAP kinases participate in vascular diseases via differential molecular mechanisms and are new therapeutic targets for treatment of vascular diseases.

Topics: Animals; Becaplermin; Carotid Artery Injuries; Cell Cycle Proteins; Cell Division; Cell Movement; Chemokine CCL2; Cyclin-Dependent Kinase Inhibitor p21; Cyclin-Dependent Kinase Inhibitor p27; Cyclins; Gene Expression Regulation; Hyperplasia; JNK Mitogen-Activated Protein Kinases; Male; MAP Kinase Kinase 4; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Mitogen-Activated Protein Kinase Kinases; Mitogen-Activated Protein Kinases; Muscle, Smooth, Vascular; p38 Mitogen-Activated Protein Kinases; Plasminogen Activator Inhibitor 1; Platelet-Derived Growth Factor; Proto-Oncogene Proteins c-sis; Rats; Rats, Sprague-Dawley; Recombinant Fusion Proteins; Transduction, Genetic; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tumor Suppressor Proteins; Tunica Intima

To evaluate the ionizing radiation for intimal hyperplasia prevention and to assess the production of growth factors.. An oversized injury using an embolectomy catheter was performed on a rabbit distal aorta (N=23), associated (test group; N=12) or not (control group; N=11) with a post-operative external radiation (25 Gy). At t=45 days, histological studies and morphometric studies were performed on the aorta. Smooth muscular cells and endothelial cells were stained using immuno-histologic revelation. Immuno-histological analysis was performed on arteries for growth factors PDGFbb, bFGF and TGFb1.. Twenty-one animals survived the procedure, 11 were in the test group and 10 in the control group. Intimal thickness and ratio intima/media were significantly lower after radiation (respectively p=0.008, p=0.008). There was no difference for the medial thickness (p=0.155). Immuno-histochemical positive staining for PDGF and TGFb1 was lower after radiation (respectively 18.44 +/- 2.963% versus 47.64 +/- 6.86%, p<0.001 and 10.11 +/- 3.18% versus 29.45 +/- 4.156%, p<0.001). There was no difference for the expression of bFGF growth factor. After radiation, the media was found to be reduced and replaced by interstitial fibrosis.. After external radiation the thickness parameter of the intima and the ratio intima/media decreased significantly in comparison with the control group. PDGF and TGFb1 were also less expressed in the artery irradiated. Fibrosis recasting needs to be confirmed by further investigation.

Topics: Animals; Aorta, Thoracic; Becaplermin; Extracellular Matrix; Female; Fibroblast Growth Factor 2; Fibrosis; Gamma Rays; Graft Occlusion, Vascular; Hyperplasia; Platelet-Derived Growth Factor; Proto-Oncogene Proteins c-sis; Rabbits; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tunica Intima; Tunica Media

We hypothesized that the venous limb of an arteriovenous (AV) fistula would evince up-regulation of genes relevant to vascular remodeling along with neointimal hyperplasia and relevant histological changes. Using the aorto-caval model of an AV fistula model in the rat, we demonstrate marked up-regulation in such proinflammatory genes as monocyte chemoattractant protein-1, plasminogen activator inhibitor-1, and endothelin-1, 2 weeks after the creation of the fistula. Neointimal hyperplasia occurred in variable degrees by 5 weeks after establishing the fistula, and by 16 weeks, such neointimal hyperplasia was progressive and pronounced; at this time point, abundant extracellular matrix was also observed. Smooth muscle cells were present in the hyperplastic neointima as evidenced by staining for alpha-smooth muscle actin; ultrastructurally, smooth muscle cells with a synthetic as well as a contractile phenotype were readily observed. Accumulation of extracellular matrix in the model at 16 weeks was accompanied by increased expression of transforming growth factor-beta1 mRNA, the latter finding contrasting with the suppression of transforming growth factor-beta1 mRNA observed in this model at 2 weeks. In summary, we describe marked up-regulation in proinflammatory genes and progressive neointimal formation in the venous vasculature in an AV fistula model in the rat. We suggest that such alteration in gene expression and histological injury, in conjunction with the relative simplicity of this model, offer a new approach in the study of such timely biological and clinically relevant phenomena as differential gene expression in response to hemodynamic forces, processes involved in vascular remodeling, mechanisms of injury in venous bypass grafts, and mechanisms of dysfunction of AV fistulae used in hemodialysis.

Topics: Animals; Arteriovenous Fistula; Blotting, Northern; Chemokine CCL2; Disease Models, Animal; Endothelin-1; Gene Expression; Hyperplasia; Inflammation; Inflammation Mediators; Microscopy, Electron; Plasminogen Activator Inhibitor 1; Rats; RNA, Messenger; Time Factors; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tunica Intima; Vena Cava, Inferior

Vasoactive intestinal polypeptide (VIP) content is increased in the hyperplastic pituitaries of estrogen (E)-treated rats, thus suggesting that this neuropeptide could mediate the E effect on lactotrophs. E also decreases pituitary TGF-beta1 content, an autocrine/paracrine inhibitor of lactotroph proliferation, and induces pituitary angiogenesis. To elucidate the role of VIP in this context, lactotroph hyperplasia was induced in female Fisher 344 rats by implanting sc pellets of diethylstilbestrol (DES). Twenty-five days later, the rats were treated with three different increasing doses of a VIP receptor antagonist or the vehicle for 5 d. DES treatment resulted in a marked increase of serum prolactin (PRL), pituitary PRL content, PRL mRNA expression, pituitary weight, and pituitary proliferating cell nuclear antigen. DES treatment also increased pituitary VIP content and VIP mRNA levels, but not in the hypothalamus and cerebral cortex. Simultaneously, DES treatment decreased the pituitary TGF-beta1 content and increased the pituitary content of vascular endothelial growth factor. VIP receptor antagonist partially reverted the effect of DES on serum PRL and pituitary PRL, proliferating cell nuclear antigen, TGF-beta1, and vascular endothelial growth factor contents, as well as on pituitary weight, in a dose-dependent relation. These data suggest that pituitary VIP mediates the effect of E on lactotroph hyperplasia, pituitary TGF-beta1, and angiogenesis.

Topics: Animals; Autocrine Communication; Cerebral Cortex; Diethylstilbestrol; Endothelial Growth Factors; Estrogens, Non-Steroidal; Female; Hyperplasia; Hypothalamus; Intercellular Signaling Peptides and Proteins; Lymphokines; Organ Size; Paracrine Communication; Pituitary Gland; Pituitary Gland, Anterior; Prolactin; Proliferating Cell Nuclear Antigen; Rats; Rats, Inbred F344; RNA, Messenger; Transforming Growth Factor beta; Transforming Growth Factor beta1; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factors; Vasoactive Intestinal Peptide

Topics: Antigens, CD; Antigens, Differentiation, B-Lymphocyte; Cell Adhesion Molecules; Child; DNA-Binding Proteins; Humans; Hyperplasia; Immunohistochemistry; Keratins; Lectins; Myasthenia Gravis; Nerve Tissue Proteins; Nuclear Proteins; Receptors, Steroid; Receptors, Thyroid Hormone; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Sialic Acid Binding Ig-like Lectin 2; Thymoma; Thymus Gland; Transforming Growth Factor beta; Transforming Growth Factor beta1

To demonstrate nephromegaly in children with biliary atresia and children with compensatory renal hypertrophy and to examine their plasma hepatocyte growth factor (HGF), transforming growth factor beta1 (TGF-beta1), and the difference of total kidney volume, 11 children with biliary atresia (age range 5 months to 10 years), 11 with compensatory renal hypertrophy, and 11 age-matched healthy controls were investigated. Kidney volume was measured by renal ultrasonography and plasma HGF and TGF-beta1 levels were studied. To clarify the significance of nephromegaly in biliary atresia, creatinine clearance was also measured in 9 children with biliary atresia and 9 healthy children. The unilateral kidney in biliary atresia and the solitary kidney in compensatory renal hypertrophy had significantly higher kidney volumes compared with those of healthy children (P<0.001 by analysis of covariance). However, a significant increase in total kidney volume was noted only in children with biliary atresia (P<0.001 by analysis of covariance). Although this was actually associated with increased creatinine clearance (117.3+/-22.0 ml/min per 1.73 m(2) vs. 98.3+/-13.6 ml/min per 1.73 m(2) in controls, P<0.05), corrected creatinine clearance was not correlated with total kidney volume (r=0.199, P=0.61) in biliary atresia. Plasma HGF levels and HGF/TGF-beta1 ratios were elevated in children with biliary atresia (2,648+/-1,215 pg/ml and 233.8+/-139.1 pg/ng vs. 493+/-131 pg/ml and 35.9+/-15.7 pg/ng in compensatory renal hypertrophy and 468+/-194 pg/ml and 24.0+/-19.6 pg/ng in controls, P<0.001) and had a positive correlation with total kidney volume by multiple regression analysis (P=0.006 and P=0.002, respectively). These results show that nephromegaly in biliary atresia is associated with increased total kidney volume and a higher glomerular filtration rate, and is positively correlated with plasma HGF and plasma HGF/TGF-beta1 ratio, implying a role of HGF in this situation. However, nephromegaly in compensatory renal hypertrophy may have different mechanisms in terms of normal total kidney volume, transient elevation of plasma HGF followed by normal plasma HGF, and normal plasma HGF/TGF-beta1 ratio. These data also suggest a common mechanism (HGF) for initial renal hypertrophy (as in compensatory renal growth), with dysregulation of control of this process later in the course (as in biliary atresia). The detailed mechanisms for nephromegaly in these two conditions should be fu

Topics: Adaptation, Physiological; Biliary Atresia; Child; Child, Preschool; Creatinine; Hepatocyte Growth Factor; Humans; Hyperplasia; Hypertrophy; Infant; Kidney; Nephrectomy; Transforming Growth Factor beta; Transforming Growth Factor beta1

Diabetes and renal failure have been associated with extremely high restenosis rates following successful angioplasty, resulting in increased morbidity and mortality. Advanced glycosylation end products (AGEs) accumulate in vascular tissues with aging and at an accelerated rate in diabetes and renal failure. AGEs are particularly abundant at sites of atherosclerotic lesions. AGEs interact with specific receptors (RAGE) present on all cells relevant to the restenosis process including inflammatory cells and smooth muscle cells. AGEs-RAGE interaction in vessel wall may lead to inflammation, smooth muscle cell proliferation, and extracellular matrix production, culminating in exaggerated intimal hyperplasia and restenosis. Following arterial injury, the interaction of AGEs with monocytes expressing RAGE can promote migration of inflammatory cells into the lesion and subsequent release of growth factors and cytokines. Binding of AGEs-RAGE on smooth muscle cells increases chemotactic migration and cellular proliferation. AGEs trigger the generation of reactive oxygen species, and upregulate the multifunctional transcription factor NF-kappa B. Finally, AGEs can augment extracellular matrix production by upregulating transforming growth factor-beta. Thus, accumulation of AGEs in vessel wall provides a common mechanism for the high restenosis rates of patients with diabetes and renal failure.

Topics: Angioplasty, Balloon, Coronary; Coronary Disease; Coronary Restenosis; Cytokines; Diabetic Angiopathies; Diabetic Nephropathies; Extracellular Matrix; Glycation End Products, Advanced; Growth Substances; Humans; Hyperplasia; Kidney Failure, Chronic; Models, Biological; Muscle, Smooth, Vascular; NF-kappa B; Receptor for Advanced Glycation End Products; Receptors, Immunologic; Stents; Transforming Growth Factor beta; Tunica Intima; Tunica Media

The immunoregulatory cytokine transforming growth factor (TGF)-beta(1) is secreted as a biologically inactive complex with latency-associated peptide, which must be modified by local factors to expose the functionally active cytokine. The epithelial integrin alpha(v)beta(6) mediates local activation of TGF-beta(1) in the lung and beta(6)(-/-) mice exhibit exaggerated pulmonary inflammation, but their response to inflammatory stimuli in the gut has not been investigated. We found that both beta(6) and TGF-beta(1) are constitutively expressed in the jejunal epithelial compartment in uninfected mice and during infection with the intestinal nematode Nippostrongylus brasiliensis. We also present data showing that beta(6)(-/-) mice are seriously compromised in their ability to mount a mucosal mast cell response after infection, and there is a significant reduction in the expression and systemic release of the granule chymase, mouse mast cell protease-1. Because in vitro expression of this chymase is regulated by TGF-beta(1), these data indicate that in the absence of alpha(v)beta(6) epithelially expressed TGF-beta(1) may not be activated, with a consequent absence of expression of mouse mast cell protease-1 and down-regulation of the mucosal mast cell response.

Topics: Animals; Antigens, Neoplasm; Chymases; Down-Regulation; Humans; Hyperplasia; Integrins; Intestinal Mucosa; Mast Cells; Mice; Mice, Transgenic; Nippostrongylus; Serine Endopeptidases; Strongylida Infections; Transforming Growth Factor beta

Analyses of mutant mice with a deletion for the transforming growth factor beta 2 (Tgfbeta2) gene revealed cysts in the perineal/scrotal region of male mice. We present evidence from in situ, light and electron microscopy that the cysts observed in Tgfbeta2+/- heterozygous mice males derive from Cowper's gland tissue. The Cowper's glands of Tgfbeta2+/- heterozygous mutant mice display all steps of glandular hyperplasia and cystic dilation. TGF-beta isoforms and TGF-beta receptor (TbetaR-II) were localized immunocytochemically in sections of Cowper's glands. TGF-beta2 and TGF-beta3 were located predominantly in myoepithelial cells of the Cowper's gland whereas the TbetaRII was found in the plasma membrane of the acinar cells. TUNEL-assays revealed that apoptotic cell death is significantly reduced in Cowper's glands of TgfbetaB2+/- heterozygous mutant mice. The fact that Tgfbeta2+/- heterozygous mutant mice exhibit hyperplasia of Cowper's gland epithelium and Cowper's gland cysts suggests a disturbance of epithelial-stromal interaction most likely due to reduced TGF-beta2 level, accompanied by a significant decrease in apoptosis.

Topics: Animals; Apoptosis; Bulbourethral Glands; Cysts; Genital Diseases, Male; Heterozygote; Hyperplasia; Immunohistochemistry; In Situ Nick-End Labeling; Male; Mice; Mice, Mutant Strains; Mice, Transgenic; Microscopy, Electron; Receptors, Transforming Growth Factor beta; Transforming Growth Factor beta; Transforming Growth Factor beta2

To study the expression of transforming growth factor alpha, beta 1 in hyperplastic tissue after endoscopic polypectomy and the effect of corticosteroid.. Forty patients with nasal polyps were divided into two groups randomly: corticosteroid group (n = 20) with topical application of Budesonide (BUD, 400 micrograms/d) after endoscopic polypectomy and control group (n = 20) without corticosteroid after surgery. The hyperplastic tissues in operative cavity obtained in the 1st and 8th weeks after operation were studied with HE staining and immunohistochemistry technique respectively.. Morphological changes of hyperplastic tissue after endoscopic polypectomy included pseudostratified epithelium, highly edematous lamina proper and inflammatory cells infiltration, in which the main infiltrative cells were eosinophils (67.5%). Transforming growth factor alpha(TGF alpha) protein was highly expressed in epithelial cells, grand cells and inflammatory cells in the hyperplastic tissue. Transforming growth factor beta 1 (TGF beta 1) protein was highly expressed in inflammatory cells in the hyperplastic tissue. The expression of TGF alpha and beta 1 was significantly decreased after topical BUD spray (P < 0.01, 0.05).. Transforming growth factor alpha and beta 1 may play an important role in the formation and recurrence of nasal polyps.

Topics: Adult; Anti-Inflammatory Agents; Budesonide; Female; Humans; Hyperplasia; Male; Nasal Mucosa; Nasal Polyps; Transforming Growth Factor alpha; Transforming Growth Factor beta

Topics: Apoptosis; Caspase 3; Caspases; Humans; Hyperplasia; Immunohistochemistry; Precancerous Conditions; Stomach; Stomach Neoplasms; Transforming Growth Factor beta; Tumor Cells, Cultured

To study the mechanism of intimal hyperplasia after coronary artery bypass grafting (CABG) and to find an effective way for preventing intimal hyperplasia.. Twenty-four male New Zealand rabbits were randomly divided into two groups of 12 rabbits: operation group and sham-operation (control) group. The external jugular vein was harvested and anastomosed end-to-side to the ipsilateral carotid artery in operation group or grafted in situ in the control group. Six rabbits in each group were killed and their grafted veins were taken 2 weeks and 4 weeks after operation respectively. The mRNA expressions of transforming growth factor beta (TGF-beta), collagen I, collagen III, and angiotension 1 receptor (AT1R) were measured by RT-PCR and electrophoresis.. The intimal hyperplasia was much more remarkable in the operation group than in the control group either 2 weeks or 4 weeks after operation. The mRNA expressions of TGF-beta, AT1R, collagen I, and collagen III were significantly higher in the operation group than in the control group, especially 2 weeks after (P < 0.01). Four weeks after the operation, the expressions of TGF-beta, AT1R, collagen I and collagen III were 4.05 +/- 0.49 vs 2.05 +/- 0.26, 18.23 +/- 1.32 vs 4.61 +/- 0.53, 80 +/- 0.17 vs 0.90 +/- 0.18, and 7.05 +/- 0.68 vs 2.80 +/- 0.17 respectively (all P < 0.05).. TGF-beta and AT1R may have an important role in the intimal hyperplasia of venous graft in CABG. Continuous arterial pressure may be the main factor of increased expression of TGF-beta and AT1R that cause the enormous synthesis and deposit of collagen.

Topics: Animals; Collagen Type I; Collagen Type III; Coronary Artery Bypass; Female; Gene Expression; Hyperplasia; Jugular Veins; Male; Rabbits; Receptor, Angiotensin, Type 1; Receptors, Angiotensin; RNA, Messenger; Time Factors; Transforming Growth Factor beta; Tunica Intima

The vascular hallmark of chronic rejection (CR), as well as of atherosclerosis, is initial hyperplasia. It results from migration and proliferation of vascular smooth muscle cell and increased deposition of extracellular matrix proteins. A possible mechanism responsible for formation of neointima is the release of growth factors and cytokines, such as: transforming growth factor beta (TGF-beta), tumour necrosis factor alfa (TNF-alpha), interleukin 1 (IL-1) and interleukin 6 (IL-6). The expression of these factors in the renal artery wall of chronically rejected allografts was quantified. The renal artery samples were obtained from patients with chronic renal allograft rejection, undergoing graftectomy (n = 11) and patients with autosomal dominant polycystic kidney disease (ADPKD), undergoing nephrectomy (n = 4). Total RNA was isolated and the expression of mRNA for TGF-beta, TNF-alpha, IL-1 and IL-6 was measured using a real time PCR. In patients with CR the expression levels of TGF-beta, TNF-alpha and IL-1 mRNA were higher than in control group. No difference between groups was detected for IL-6. In both groups a correlation was detected between age and TGF-beta expression. The increased expression of TGF-beta, TNF-alpha and IL-1 may be a key factor in the neointimal formation and pathogenesis of CR. The increase in the TGF-b expression with age might be a protective mechanism in atherosclerosis.

Topics: Adult; Aged; Case-Control Studies; Cytokines; Female; Gene Expression; Graft Rejection; Growth Substances; Humans; Hyperplasia; Interleukin-1; Interleukin-6; Kidney Transplantation; Male; Middle Aged; Polycystic Kidney, Autosomal Dominant; Polymerase Chain Reaction; Renal Artery; RNA, Messenger; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

Bone morphogenetic protein (BMP) signaling is known to be involved in multiple inductive events during embryogenesis including the development of amniote skin. Here, we demonstrate that early application of BMP-2 to the lateral trunk of chick embryos induces the formation of dense dermis, which is competent to participate in feather development. We show that BMPs induce the dermis markers Msx-1 and cDermo-1 and lead to dermal proliferation, to expression of beta-catenin, and eventually to the formation of ectopic feather tracts in originally featherless regions of chick skin. Moreover, we present a detailed analysis of cDermo-1 expression during early feather development. The data implicate that cDermo-1 is located downstream of BMP in a signaling pathway that leads to condensation of dermal cells. The roles of BMP and cDermo-1 during development of dermis and feather primordia are discussed.

Topics: Animals; beta Catenin; Biomarkers; Bone Morphogenetic Protein 2; Bone Morphogenetic Proteins; Carrier Proteins; Chick Embryo; Cytoskeletal Proteins; DNA, Complementary; Feathers; Gene Expression Regulation, Developmental; Homeodomain Proteins; Hyperplasia; In Situ Hybridization; Molecular Sequence Data; MSX1 Transcription Factor; Proliferating Cell Nuclear Antigen; Proteins; Signal Transduction; Skin; Trans-Activators; Transcription Factors; Transforming Growth Factor beta

The c-Met tyrosine kinase receptor and its ligand, Hepatocyte Growth Factor/ Scatter Factor, have been implicated in human cancer. We have previously described that the transgenic expression of a truncated form of human c-Met (cyto-Met) in the liver confers resistance to several apoptotic stimuli. Here we show the impact of cyto-Met expression on liver proliferation and transformation. Despite a sixfold increase of hepatocyte proliferation, adult transgenic livers displayed normal size and architecture. We present evidence showing that activation of TGF-beta1 signalling controls the liver mass in cyto-Met mice. The oncogenic potential of cyto-Met was further assessed in the context of c-Myc-induced hepatocarcinogenesis, using WHV/c-Myc transgenic mice. Co-expression of cyto-Met and c-Myc further enhanced hepatocyte proliferation and caused a dramatic acceleration of the Myc-induced tumorigenesis, leading to the emergence of hepatocarcinomas in 3-4-month-old animals. Importantly, the TGF-beta receptor type II expression was strongly downregulated in most tumours, indicating that impairment of TGF-beta1-mediated growth inhibition plays a major role in accelerated neoplastic development. The strong potential of cyto-Met for oncogenic cooperation without direct transforming activity designates cyto-Met mice as an ideal tool for studying the early steps of multistage hepatocarcinogenesis and for identification of prognostic markers of transformation.

Topics: Animals; Apoptosis; Blotting, Western; Cell Division; Cell Transformation, Neoplastic; Down-Regulation; Gene Expression Regulation, Neoplastic; Hepatitis B Virus, Woodchuck; Hepatocytes; Homeostasis; Humans; Hyperplasia; Liver Neoplasms; Mice; Mice, Transgenic; Organ Size; Promoter Regions, Genetic; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins c-met; Proto-Oncogene Proteins c-myc; Receptor, Transforming Growth Factor-beta Type II; Receptors, Transforming Growth Factor beta; RNA, Messenger; Signal Transduction; Transforming Growth Factor beta; Transforming Growth Factor beta1; Transgenes

Halofuginone is a drug that has been shown to have an antifibrotic property in vitro and in vivo. Whereas halofuginone shows promise as a therapeutic agent for a variety of diseases including scleroderma, liver cirrhosis, cystic fibrosis, and certain types of cancer, the mechanism of action remains unknown. Using the tight skin mouse (TSK) model for scleroderma, we evaluated the ability of halofuginone to inhibit spontaneous development of dermal fibrosis. We found that administration of a low dose of halofuginone both in adult and newborn animals for 60 d prevented the development of cutaneous hyperplasia (dermal fibrosis). In vitro halofuginone was found to reduce the amount of collagen synthesized by fibroblasts. This effect was due to a reduction in the promoter activity of the type-I collagen genes as treatment of fibroblast cultures with 10(-8) M halofuginone reduced the level of alpha2(I) collagen message detectible by northern blot and greatly reduced the activity of a reporter construct under control of the -3200 to +54 bp alpha2(I) collagen promoter. In addition, analysis of transforming growth factor beta signaling pathways in fibroblasts revealed that halofuginone inhibited transforming-growth-factor-beta-induced upregulation of collagen protein and activity of the alpha2(I) collagen promoter. Further we found that halofuginone blocked the phosphorylation and subsequent activation of Smad3 after transforming growth factor beta stimulation. Apparently the inhibitory property was specific to Smad3 as there was no inhibitory effect on the activation of Smad2 after stimulation with transforming growth factor beta. Our results demonstrate that halofuginone is a specific inhibitor of type-I collagen synthesis and may elicit its effect via interference with the transforming growth factor beta signaling pathway.

Topics: Animals; Cells, Cultured; Collagen; Collagen Type I; DNA-Binding Proteins; Fibroblasts; Gene Expression; Hyperplasia; Male; Mice; Mice, Inbred C57BL; Mice, Inbred Strains; Mice, Mutant Strains; Piperidines; Quinazolines; Quinazolinones; Receptors, Transforming Growth Factor beta; Sclerosis; Signal Transduction; Skin; Skin Diseases; Smad3 Protein; Trans-Activators; Transforming Growth Factor beta

Arterial intimal thickening is consisted of predominately smooth muscle cells (SMC). The source of these SMCs and mechanisms response for their changes have not been well cleared. Using a model of rabbit common carotid artery (CCA) shear induced intimal thickening, we sought to identify and describe the source of SMCs in intima. The enlarged CCA 28 days after arteriovenous fistula (AVF) creation was subjected to subnormal wall shear stress (WSS) for 1, 3, and 7 days by closure of the AVF. To determine SMC proliferation, BrdU pulse labeling of SMCs was performed. BrdU-labeled SMCs were tracked over time to further confirm SMC migration. In response to subnormal WSS intimal thickening developed progressively. BrdU-labeled SMCs localized in the subendothelial area. When the BrdU-labeled medial SMCs were tracked 1 day after AVF closure, progenies of these BrdU-incorporated SMCs increased by 4.8-fold with 75% of them in the intima. They were 12-fold increased with 83% in the intima 7 days after. En face examination showed an accumulation of SMCs in internal elastic lamina gap after AVF closure, which later migrated into subendothelial area. In situ hybridization revealed increased TGF-beta1 mRNA expression in intimal SMCs. This study demonstrates that the medial SMCs are the predominant cells in subnormal WSS-induced intimal thickening. Early expression of TGF-beta1 may play an important role in the process of intimal thickening.

Topics: Adaptation, Physiological; Animals; Arteriovenous Shunt, Surgical; Cell Movement; Disease Models, Animal; Hyperplasia; In Situ Hybridization; Male; Muscle, Smooth, Vascular; Rabbits; Regional Blood Flow; RNA, Messenger; Stress, Mechanical; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tunica Intima

Runx3/Pebp2alphaC null mouse gastric mucosa exhibits hyperplasias due to stimulated proliferation and suppressed apoptosis in epithelial cells, and the cells are resistant to growth-inhibitory and apoptosis-inducing action of TGF-beta, indicating that Runx3 is a major growth regulator of gastric epithelial cells. Between 45% and 60% of human gastric cancer cells do not significantly express RUNX3 due to hemizygous deletion and hypermethylation of the RUNX3 promoter region. Tumorigenicity of human gastric cancer cell lines in nude mice was inversely related to their level of RUNX3 expression, and a mutation (R122C) occurring within the conserved Runt domain abolished the tumor-suppressive effect of RUNX3, suggesting that a lack of RUNX3 function is causally related to the genesis and progression of human gastric cancer.

Topics: Animals; Apoptosis; Cell Division; Cell Transformation, Neoplastic; Core Binding Factor Alpha 3 Subunit; DNA Methylation; DNA-Binding Proteins; Epithelium; Exons; Female; Gastric Mucosa; Gene Deletion; Gene Expression Regulation, Neoplastic; Gene Targeting; Humans; Hyperplasia; Male; Mice; Mice, Knockout; Protein Structure, Tertiary; Stomach; Stomach Neoplasms; Transcription Factors; Transforming Growth Factor beta; Tumor Cells, Cultured; Tumor Suppressor Proteins

To characterize, at the histopathologic and molecular levels, the irradiated epidermis in cases of human skin fibrosis induced by radiotherapy.. Surgical samples were obtained from 6 patients who had developed cutaneous fibronecrotic lesions from 7 months to 27 years after irradiation. The proliferation and differentiation status of the irradiated epidermis was characterized with specific markers using immunohistochemical methods.. All samples presented with hyperplasia of the epidermis associated with local inflammation. The scar epidermis exhibited an increased expression of proliferating cell nuclear antigen, which revealed hyperproliferation of keratinocytes. Furthermore, an abnormal differentiation was found, characterized by the expression of K6 and K16, and by alterations in protein amounts and localization of cytokeratins, involucrin, and transforming growth factor-beta1.. These results demonstrate that late damage of irradiated skin is not only characterized by fibrosis in the dermis but also by hyperplasia in the epidermis. This hyperplasia was due to both hyperproliferation and abnormal differentiation of keratinocytes.

Topics: Adult; Aged; Cell Differentiation; Cell Division; Cicatrix; Female; Humans; Hyperplasia; Integrins; Keratinocytes; Keratins; Male; Middle Aged; Proliferating Cell Nuclear Antigen; Radiation Injuries; Skin; Transforming Growth Factor beta

Myostatin, which is a member of the TGF-beta superfamily, is a negative regulator of skeletal muscle formation. Double-muscled Piedmontese cattle have a C313Y mutation in myostatin and show increased skeletal muscle mass which resulted from an increase of myofiber number (hyperplasia) without that of myofiber size (hypertrophy). To examine whether this mutation in myostatin gene affects muscle development in a dominant negative manner, we generated transgenic mice overexpressing the mutated gene. The transgenic mice exhibited dramatic increases in the skeletal muscle mass resulting from hyperplasia without hypertrophy. In contrast, it has been reported that a myostatin mutated at its cleavage site produces hypertrophy without hyperplasia in the muscle. Thus, these results suggest that (1) the myostatin containing the missense mutation exhibits a dominant negative activity and that (2) there are two types in the dominant negative form of myostatin, causing either hypertrophy or hyperplasia.

Topics: Amino Acid Substitution; Animals; Cattle; Chickens; DNA Primers; Female; Hyperplasia; Hypertrophy; Male; Mice; Mice, Transgenic; Muscle, Skeletal; Mutation, Missense; Myogenin; Myostatin; Polymerase Chain Reaction; Sex Characteristics; Transforming Growth Factor beta

N(3,4-dimethoxycinnamoyl) anthranilic acid (tranilast) prevents the synchronous upregulation of isoforms and receptors of the transforming growth factor (TGF)-beta system after arterial injury and reduces restenosis after human coronary angioplasty. However, the effects of tranilast and the importance of the TGF-beta system in stent restenosis, in which inward remodeling is unimportant but inflammatory cell stimulation of neointima formation is exaggerated, are uncertain. Boston minipigs, treated with tranilast or vehicle, were subjected to endoluminal stenting, and the expression of TGF-beta1 and TGF-beta3, the expression of their signaling receptors ALK-5 and TbetaR-II, leukocyte numbers around the stent struts, and neointima development were assessed over 28 days. Stenting greatly increased early (5-day) mRNA expression of the 2 TGF-beta isoforms and their receptors. Immunohistochemical localization later showed that their concentrations were greatest in regions adjacent to stent struts, where leukocytes and collagen deposition were prevalent. Tranilast suppressed these elevations in TGF-beta mRNAs and reduced their immunoreactive peptides detectable around stent struts. The accumulation of leukocytes and deposition of collagen in these regions was also greatly inhibited by tranilast. These effects were associated with a 48% reduction in maximal neointimal cross-sectional area and 43% reduction in mean neointimal cross-sectional area at 28 days (P<0.05). We conclude that tranilast suppresses neointima development after stenting, effects that can be at least partly attributed to its ability to attenuate the induction of the TGF-beta system and leukocyte accumulation around stent struts.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Coronary Restenosis; Coronary Vessels; Drug Administration Schedule; Hyperplasia; Inflammation; Leukocytosis; Male; ortho-Aminobenzoates; Stents; Swine; Transforming Growth Factor beta; Tunica Intima; Up-Regulation

Thrombus formation is a key component of the pathogenesis of restenosis after arterial balloon injury. The purpose of this study was to determine whether intimal hyperplasia could be attenuated by infusion of recombinant tissue plasminogen activator (tPA). Forty-two Kurosawa and Kusanagi hypercholesterolemic rabbits were divided into tPA (n = 20) and control (n = 22) groups, the former receiving 7 days of continuous tPA infusion (0.6 mg/kg/day) via ear veins. The walls of the common iliac arteries were injured using 2.5-mm balloon catheters and then examined histologically 7, 14, 21, and 28 days later. Cell proliferation was assessed by immunohistochemical analysis of proliferating cell nuclear antigen (PCNA), and transforming growth factor (TGF)-beta immunohistochemistry was carried out to estimate cell proliferation and differentiation. It was observed that 28 days after balloon injury, intimal cross-sectional areas in the tPA group were significantly smaller than in controls (0.11 +/- 0.03 mm2 vs. 0.57 +/- 0.08 mm2, p < 0.01), as were ratios of the cross-sectional areas of the intima and media (0.21 +/- 0.07 vs. 1.06 +/- 0.18, p < 0.05). In addition, the numbers of PCNA-positive medial cells were significantly lower (0.06 +/- 0.01 vs. 0.36 +/- 0.08, p < 0.05) and TGF-beta-positive vessel wall areas were significantly smaller in tPA-treated animals 7 days after balloon injury (0.47 +/- 0.28% vs. 4.55 +/- 1.44%, p < 0.05). Thus infusion of tPA after arterial balloon injury appears to decrease medial cell proliferation and suppress intimal hyperplasia.

Topics: Animals; Cell Division; Female; Hemodynamics; Hypercholesterolemia; Hyperplasia; Immunohistochemistry; Male; Muscle, Smooth, Vascular; Plasminogen Activator Inhibitor 1; Proliferating Cell Nuclear Antigen; Rabbits; Recombinant Proteins; Tissue Plasminogen Activator; Transforming Growth Factor beta

To date established treatment of transplant arteriosclerosis is basically missing and there is a need for new therapeutic approaches. Angiotensin II (Ang II) and Ang II receptor type 1 (AT) are present in the vascular wall. Blocking of the AT1 receptor by pharmacological agents may inhibit damaging effects of Ang II on endothelial and smooth muscle cells. The purpose of the study was to evaluate the effect of the AT1 receptor blocker Candesartan cilexetil on the development of graft arteriosclerosis in a rat aortic transplant model. Two strain combinations were used for aortic transplantation: DA to PVG; and PVG to PVG. The animals received Candesartan cilexetil treatment (9.5 + 1.4 mg/kg/day) for 8 weeks. Candesartan cilexetil treatment reduced neointimal formation both in allografts (Qint 30.2 +/- 8.8% vs. 22.1 +/- 8.7%, P < 0.05) and in isografts (Qint 15.5 +/- 4.4% vs. 6.7 +/- 3.3%, P = 0.0001). Blocking of the AT1 receptor signalling by Candesartan cilexetil was also associated with a reduced expression of TGF-beta1. Macrophage infiltration was not affected by the treatment. Candesartan cilexetil treatment leads to reduced neointimal formation in aortic transplant. The positive effect of the drug might be partly explained by a reduction of TGF-beta1 expression in the grafts. Candesartan treatment may provide another possibility for prevention of transplant arteriosclerosis and chronic rejection.

Topics: Animals; Aorta, Abdominal; Arteriosclerosis; Benzimidazoles; Biphenyl Compounds; Blood Pressure; Drug Evaluation, Preclinical; Graft Rejection; Hyperplasia; Macrophages; Male; Models, Animal; Postoperative Complications; Rats; Rats, Inbred Strains; Receptor, Angiotensin, Type 1; Receptor, Angiotensin, Type 2; Receptors, Angiotensin; Renin-Angiotensin System; Tetrazoles; Transforming Growth Factor beta; Transforming Growth Factor beta1; Transplantation, Homologous; Transplantation, Isogeneic; Tunica Intima; Tunica Media

The pathological role of oxidative stress in patients treated by hemodialysis has gained increasing recognition in recent years. Because complications related to vascular access are a major source of morbidity, immunohistochemical evidence of oxidative stress and activation of growth factors were examined in native arteriovenous (AV) fistulae (n = 11) and expanded polytetrafluoroethylene (ePTFE) grafts (n = 15) recovered from hemodialysis patients at the time of surgical revision or resection. To show the presence of oxidative stress in tissues, three markers were chosen: N(epsilon)(carboxymethyl)lysine, a structurally identified advanced glycation end product; 4-hydroxy-2,3-nonenol, a lipid peroxidation product; and redox-active transition metals bound to proteins, a source of Fenton chemistry-generated free radicals. Markers of cell growth and proliferation were endothelin-1 (ET-1), a potent mitogenic peptide implicated in the formation of intimal hyperplasia; transforming growth factor-beta (TGF-beta), a stimulus to vascular cell growth and matrix production; and platelet-derived growth factor (PDGF), a mediator of intimal hyperplasia. All specimens studied showed significant intimal hyperplasia. In general, the neointima close to the vascular lumen of the AV fistula and the pseudointima close to the lumen of the ePTFE graft were positive for oxidative stress markers. At sites of injury, especially in the presence of histological evidence of inflammation and healing, expression of oxidative markers was particularly intense. Prominent staining of PDGF was shown at sites of anastomotic hyperplasia and in neovasculature. TGF-beta was associated with proliferation or repair in both AV fistulae and ePTFE grafts. ET-1 staining was most intense in the neointima and pseudointima. This study showed histochemical colocalization of markers of oxidative stress with growth factors known to contribute to intimal hyperplasia.

Topics: Adult; Aged; Arteriovenous Anastomosis; Arteriovenous Fistula; Arteriovenous Shunt, Surgical; Biomarkers; Constriction, Pathologic; Endothelin-1; Female; Growth Substances; Humans; Hyperplasia; Iron; Kidney Failure, Chronic; Lipid Peroxidation; Lysine; Male; Middle Aged; Oxidation-Reduction; Oxidative Stress; Platelet-Derived Growth Factor; Polytetrafluoroethylene; Renal Dialysis; Transforming Growth Factor beta; Tunica Intima; Vascular Patency

To study the role of transforming growth factor type beta1 (TGFbeta1) in epidermal growth control and disease, we have generated a conditional expression system by using the bovine keratin 5 promoter to drive expression of the tetracycline-regulated transactivators tTA and rTA, and a constitutively active mutant of TGFbeta1 linked to the tetO target sequence for the transactivator. This model allows for induction or suppression of exogenous TGFbeta1 with oral doxycycline. Maximal expression of TGFbeta1 during gestation caused embryonic lethality, whereas partial suppression allowed full-term development with neonatal lethality characterized by runting, epidermal hypoproliferation, and blocked hair follicle growth. With complete suppression, phenotypically normal double transgenic (DT) mice were born. Acute induction of TGFbeta1 in the epidermis of adult mice inhibited basal and follicular keratinocyte proliferation and reentry of telogen hair follicles into anagen. However, chronic expression of TGFbeta1 in adult DTs caused severe alopecia characterized by epidermal and follicular hyperproliferation, apoptosis, as well as dermal fibrosis and inflammation. Readministration of doxycycline to tTA DT mice caused hair regrowth within 14 days. The mRNA and protein for Smad7, an inhibitor of TGFbeta signaling, were up-regulated in the epidermis and hair follicles of alopecic skin and rapidly induced in rTA mice in parallel with the TGFbeta1 transgene, suggesting that the hyperproliferative phenotype may result in part from development of a sustained negative feedback loop. Thus, this conditional expression system provides an important model for understanding the role of TGFbeta1 during development, in normal skin biology, and in disease.

Topics: Alopecia; Animals; Animals, Newborn; Apoptosis; Base Sequence; Cell Division; DNA Primers; DNA-Binding Proteins; Doxycycline; Gene Expression Regulation, Developmental; Genes, Lethal; Hyperplasia; Keratinocytes; Mice; Smad7 Protein; Trans-Activators; Transforming Growth Factor beta

Accelerated coronary arteriosclerosis remains a major problem for the long-term survival of cardiac transplant recipients. However, the pathogenesis of graft vasculopathy is poorly understood and there is no effective therapy. Tranilast is a promising drug that may prevent post-angioplasty restenosis. Here, we investigated whether orally administered tranilast inhibits the development of intima hyperplasia in a mouse model of cardiac transplantation. Cardiac allografts from BALB/c mice were transplanted heterotopically into C3H/He mice. Mice were administered either vehicle or tranilast everyday by gavage. Morphometrical analysis of the cardiac allografts harvested at 2 months revealed that the administration of tranilast significantly reduced the development of coronary atherosclerosis. In the mice treated with tranilast, up-regulation of the cyclin-dependent kinase inhibitor p21 was observed in the allografts, accompanied by a reduced number of proliferating cells. Tranilast also suppressed transforming growth factor-beta (TGF-beta) expression. Tranilast may be effective in preventing transplant-associated arteriosclerosis through its anti-inflammatory and anti-proliferative effects.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Cell Division; Coronary Artery Disease; Cyclin-Dependent Kinase Inhibitor p21; Cyclins; Heart Transplantation; Hyperplasia; Mice; Mice, Inbred BALB C; Mice, Inbred C3H; Muscle, Smooth, Vascular; ortho-Aminobenzoates; Transforming Growth Factor beta

To investigate the effect of transforming growth factor beta 1 (TGF beta 1) and p27kip1 in gastric mucosa carcinogenesis.. RT-PCR was used to detect TGF beta 1 and p27kip1 mRNAs in normal gastric mucosa, simple hyperplasia, dysplasia and gastric carcinoma tissues respectively.. There were expressions of TGF beta 1 and p27kip1 mRNAs in normal gastric mucosa, simple hyperplasia, dysplasia and gastric carcinoma tissues, the expressive levels of TGF beta 1 and p27kip1 mRNAs decreased gradually among the four groups. The expressive level of TGF beta 1 and p27kip1 mRNAs in gastric carcinoma group were significantly lower than those in other three groups(P < 0.05).. The data indicate that TGF beta 1 and p27kip1 may play an important role in the development and carcinogenesis of gastric carcinoma. Semi-quantification PCR technique is effective for detecting mRNA expressive level in tissues.

Topics: Cell Cycle Proteins; Cyclin-Dependent Kinase Inhibitor p27; Cyclin-Dependent Kinases; Gastric Mucosa; Genes, Tumor Suppressor; Humans; Hyperplasia; Precancerous Conditions; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Stomach Neoplasms; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tumor Suppressor Proteins

The modification of the distal anastomosis of polytetrafluoroethylene (PTFE) bypass grafts with vein interposition cuffs (VCs) has been reported to increase graft patency. However, the mechanisms that are responsible for this improved patency are unclear. Because intimal hyperplasia (IH) is a primary cause of prosthetic graft failure, we hypothesized that VCs affect the distal anastomosis by decreasing the IH response of the outflow artery.. Twenty-three female domestic Yorkshire pigs (mean weight, 35 kg) underwent 42 femoral PTFE bypass grafting procedures. The PTFE bypass grafts were separated into the following three groups according to distal anastomotic configuration: end-to-side anastomoses (ES), VCs, and cuffs constructed with PTFE (PCs). Four femoral arteries from two pigs served as healthy controls. At sacrifice, the grafts were perfusion fixed, and the distal anastomoses harvested at 1 and 4 weeks. The specimens were hemisected and serially sectioned to identify the heel, toe, and mid-anastomotic regions. The sections were cut into 5-microm segments and analyzed for intima and media thickness and area, intima/media area ratio, and the distribution of IH in the vein cuff. The roles of transforming growth factor-beta1 and platelet-derived growth factor-BB in IH development were assessed with immunohistochemistry.. IH development was significantly lower at all areas of the anastomosis, with VCs compared with ES and PCs at 4 weeks (P

Topics: Actins; Anastomosis, Surgical; Animals; Becaplermin; Blood Vessel Prosthesis; Blood Vessel Prosthesis Implantation; Disease Models, Animal; Female; Femoral Artery; Graft Survival; Hemodynamics; Hyperplasia; Immunohistochemistry; Jugular Veins; Materials Testing; Platelet-Derived Growth Factor; Polytetrafluoroethylene; Prosthesis Design; Proto-Oncogene Proteins c-sis; Saphenous Vein; Swine; Transforming Growth Factor beta; Tunica Intima; Vascular Patency

Myostatin, a TGF-beta family member, is a negative regulator of muscle growth. Here, we generated transgenic mice that expressed myostatin mutated at its cleavage site under the control of a muscle specific promoter creating a dominant negative myostatin. These mice exhibited a significant (20-35%) increase in muscle mass that resulted from myofiber hypertrophy and not from myofiber hyperplasia. We also evaluated the role of myostatin in muscle degenerative states, such as muscular dystrophy, and found significant downregulation of myostatin. Thus, further inhibition of myostatin may permit increased muscle growth in muscle degenerative disorders.

Topics: Animals; Blotting, Northern; Gene Expression; Gene Expression Regulation; Hyperplasia; Hypertrophy; Mice; Mice, Transgenic; Muscle Fibers, Skeletal; Muscle, Skeletal; Muscular Dystrophy, Animal; Mutagenesis; Myostatin; RNA, Messenger; Transforming Growth Factor beta

Diabetes mellitus-induced nephromegaly is thought to involve both hyperplastic and hypertrophic proximal tubule cell growth. The temporal relationship between these growth patterns and the mechanisms that mediate them are unknown.. Renal growth was assayed in isolated renal proximal tubules harvested from diabetic rats. Diabetes mellitus was induced by streptozotocin.. Following the induction of a diabetic state, there was a progressive increase in the kidney:body weight ratio. This was associated with an increase in 5-bromo-2-deoxyuridine incorporation (marker for hyperplastic cell growth) at day 2, which returned to baselines levels by day 4, and an increase in the protein:DNA ratio (marker for hypertrophic cell growth), which was clearly evident by day 10. Thus, diabetes-induced proximal tubule growth involved an initial hyperplastic, followed by a hypertrophic, growth period. During the hyperplastic growth period, both cdk4/cyclin D (cyclin D) and cdk2/cyclin E (cyclin E) kinase activities were increased. The switch between the growth periods was associated with continued activation of cyclin D, but inhibition of cyclin E kinase. The reduction in cyclin E kinase activity correlated with a reduction in cdk2/cyclin E complex abundance and an increased abundance of cyclin kinase inhibitors in cdk2/cyclin E complexes that did form. Also associated with the switch in growth patterns was a change in transforming growth factor-beta (TGF-beta) receptor expression. During the hyperplastic growth period, TGF-beta receptor II expression was decreased, while during the hypertrophic growth period, there was both a return of receptor II expression to baseline levels and increased expression of receptor I. Consistent with an increase in TGF-beta signaling during hypertrophy, there was an increase in Smad 2/3 protein expression and an increase in the abundance of Smad 2/4 complexes.. Diabetes-induced proximal tubule growth involves an initial hyperplastic growth period that switches to a hypertrophic growth period within a couple of days. The pattern of G1 kinase activity associated with the growth pattern switch demonstrates that the hypertrophy is mediated by a cell cycle-dependent mechanism. Regulation of TGF-beta receptor expression and signaling activity through the Smad protein cascade possibly plays a role in the growth pattern switch.

Topics: Animals; CDC2-CDC28 Kinases; Cyclin E; Cyclin-Dependent Kinase 2; Cyclin-Dependent Kinase 4; Cyclin-Dependent Kinases; Diabetes Mellitus, Experimental; Diabetic Nephropathies; DNA-Binding Proteins; G1 Phase; Hyperplasia; Hypertrophy; Kidney Tubules, Proximal; Male; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; Rats; Rats, Sprague-Dawley; Receptors, Transforming Growth Factor beta; Signal Transduction; Smad2 Protein; Smad4 Protein; Trans-Activators; Transforming Growth Factor beta

Abstract. Hyperplasia of mesangial cells (MC) is a frequent finding in glomerulonephritis. The control and function of cyclin D1, a regulator of cell cycle progression, in MC proliferation in vivo and in vitro were investigated. In a rat model of mesangioproliferative glomerulonephritis, increases in the number of cyclin D1-positive MC nuclei were prominent on day 5 of the disease, preceding the peak of MC hyperplasia. In growth-arrested rat MC in culture, mitogenic stimulation with serum or platelet-derived growth factor (PDGF) led to rapid increases in cyclin D1 protein expression. Transforming growth factor-beta1 inhibited PDGF induction of cyclin D1 protein at 12 h. In an examination of the subcellular distribution of cyclin D1, it was observed that stimulation of MC with PDGF for 6 h caused translocation of cyclin D1 from the cytoplasm into the nucleus. Coincubation with PDGF and transforming growth factor-beta1 completely inhibited this effect, without altering the cellular cyclin D1 protein abundance at that time point. To test whether reduction of cyclin D1 protein levels was sufficient to inhibit mitogenesis, MC were transfected with antisense oligonucleotides (ODN) complementary to rat cyclin D1 mRNA. Antisense ODN against cyclin D1 reduced the serum- or PDGF-induced protein expression of cyclin D1 to 27 or 10% of control levels, respectively. These inhibitory effects were correlated with diminished cyclin-dependent kinase 4 activity. Antisense ODN against cyclin D1 also decreased the PDGF-induced increase in p21(Waf-1) protein levels. The MC proliferation caused by serum or PDGF was markedly inhibited by antisense ODN against cyclin D1, as measured by [(3)H]thymidine uptake and cell counts. It is concluded that increased cyclin D1 protein expression of MC is required for MC proliferation. Targeting cyclin D1 expression may represent an effective means to inhibit MC proliferation in vitro and in vivo.

Topics: Animals; Biological Transport; Cattle; Cell Nucleus; Cells, Cultured; Cyclin D1; Cyclin-Dependent Kinase 4; Cyclin-Dependent Kinase Inhibitor p21; Cyclin-Dependent Kinases; Cyclins; DNA; Fetal Blood; Glomerular Mesangium; Glomerulonephritis, Membranoproliferative; Hyperplasia; Male; Mitosis; Oligonucleotides, Antisense; Platelet-Derived Growth Factor; Proto-Oncogene Proteins; Rats; Rats, Sprague-Dawley; Transforming Growth Factor beta

GDF-8, also known as myostatin, is a member of the transforming growth factor-beta (TGF-beta) superfamily of secreted growth and differentiation factors that is expressed in vertebrate skeletal muscle. Myostatin functions as a negative regulator of skeletal muscle growth and myostatin null mice show a doubling of muscle mass compared with normal mice. We examined femoral morphology of adult myostatin-deficient mice to assess the effects of muscle fiber hypertrophy and hyperplasia on bone shape and cross-sectional geometry. Femora of age- and weight-matched adult mice homozygous for the disrupted myostatin sequence were compared with those of wild-type controls (n = 8 per group). Results show that, as was the case in previous studies, myostatin null mice have hindlimb muscle masses that are approximately double those of controls. Myostatin-deficient mice exhibit third trochanters that are significantly larger than those of controls, whereas the femoral midshafts of the control and experimental mice do not differ significantly from one another in cortical area, bending moment of inertia, and polar moment of inertia. Our findings indicate that the increased muscle mass of myostatin-deficient mice primarily affects sites of muscle insertion, but does not induce additional cortical bone deposition in the diaphysis relative to controls. We therefore conclude that the expanded third trochanters of myostatin-deficient subjects result from tendon and Sharpey fiber expansion associated with muscle growth rather than cortical bone deposition in response to increased levels of mechanical stress.

Topics: Animals; Biomechanical Phenomena; Femur; Hindlimb; Hyperplasia; Hypertrophy; Mice; Mice, Inbred C57BL; Mice, Knockout; Muscle Development; Muscle, Skeletal; Myostatin; Transforming Growth Factor beta

To investigate time course of TGF beta(1) and VEGF expression and their role in intimal hyperplasia.. In situ hybridization and immunohistochemical technique were used to detect the time course of intimal hyperplasia, time course of TGF beta(1) mRNA and protein expression of TGF beta(1) and VEGF.. After autogenous vein replacement, the obvious neointima was seen at 2 weeks, and peaked at 8 weeks. The expression of TGF beta(1) mRNA peaked at 1 week and decreased gradually, but at 10 weeks, its positive cell percentage was still 10.1%. Both protein expression of TGF beta(1) and VEGF in VSMCs increased from 24 hours after grafting and peaked at 2 weeks. Their positive cell percentages were 40.6% and 36.6% respectively. After 4 weeks, their expression decreased at 8 weeks, the positive cell percentages were 8.9% and 13.8% respectively.. TGF beta(1) plays an important part in ECM accumulation by promoting ECM synthesis and decreasing ECM degradation. VEGF plays the key role in reendothelialization. They may affect each other and cooperated in the formation of intimal hyperplasia.

Topics: Animals; Female; Hyperplasia; Immunohistochemistry; In Situ Hybridization; Jugular Veins; Male; Rats; Rats, Wistar; RNA, Messenger; Transforming Growth Factor beta; Transplantation, Autologous; Vascular Endothelial Growth Factor A

We have examined the role that individual TGF-beta isoforms, and in particular TGF-beta3, play in control of epidermal homeostasis. Mice with a knockout mutation of the TGF-beta3 gene die a few hours after birth. A full-thickness skin grafting approach was used to investigate the postnatal development and homeostatic control of the skin of these mice. Grafted skin of mice with a disruption of the TGF-beta3 gene developed similarly to grafts of wild type and TGF-beta1 knockout animals. However, a strikingly different response was observed after acute treatment with the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA). When exposed to TPA, the grafted skin of wild type and TGF-beta1 knockout mice underwent a hyperplastic response similar to that of normal mouse skin. In marked contrast, TPA treatment of TGF-beta3 knockout grafts induced widespread areas of keratinocyte cell death. Analysis of cultured keratinocytes treated with purified TGF-beta isoforms revealed that TGF-beta3 plays a direct and specific function in protecting keratinocytes against TPA-induced cell death. The protective function of TGF-beta3 on TPA-induced cell death was not because of general suppression of the signaling pathways triggered by this agent, as ERK1/2 activation occurred to a similar if not greater extent in TGF-beta3-treated versus control keratinocytes. Instead, TGF-beta3 treatment led to a significant reduction in TPA-induced c-Jun N-terminal kinase activity, which was associated and possibly explained by specific counteracting effects of TGF-beta3 on TPA-induced disruption of keratinocyte focal adhesions.

Topics: Actins; Animals; Calcium; Calcium-Calmodulin-Dependent Protein Kinases; Cell Death; Cells, Cultured; Hyperplasia; JNK Mitogen-Activated Protein Kinases; Keratinocytes; Mice; Mice, Knockout; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Mitogen-Activated Protein Kinases; Rabbits; Skin; Tetradecanoylphorbol Acetate; Transforming Growth Factor beta

Using the rat balloon catheter denudation model, we examined the role of transforming growth factor-beta (TGF-beta) isoforms in vascular repair processes. By en face in situ hybridization, proliferating and quiescent smooth muscle cells in denuded vessels expressed high levels of mRNA for TGF-beta1, TGF-beta2, TGF-beta3, and lower levels of TGF-beta receptor II (TGF-betaRII) mRNA. Compared with normal endothelium, TGF-beta1 and TGF-beta2, as well as TGF-betaRII, mRNA were upregulated in endothelium at the wound edge. Injected recombinant soluble TGF-betaRII (TGF-betaR:Fc) localized preferentially to the adventitia and developing neointima in the injured carotid artery, causing a reduction in intimal lesion formation (up to 65%) and an increase in lumen area (up to 88%). The gain in lumen area was largely due to inhibition of negative remodeling, which coincided with reduced adventitial fibrosis and collagen deposition. Four days after injury, TGF-betaR:Fc treatment almost completely inhibited the induction of smooth muscle alpha-actin expression in adventitial cells. In the vessel wall, TGF-betaR:Fc caused a marked reduction in mRNA levels for collagens type I and III. TGF-betaR:Fc had no effect on endothelial proliferation as determined by reendothelialization of the denuded rat aorta. Together, these findings identify the TGF-beta isoforms as major factors mediating adventitial fibrosis and negative remodeling after vascular injury, a major cause of restenosis after angioplasty.

Topics: Actins; Angioplasty, Balloon; Animals; Aorta; Carotid Arteries; Carotid Artery Injuries; Cell Differentiation; Cell Division; Collagen; Endothelium, Vascular; Extracellular Matrix; Fibroblasts; Fibrosis; Gene Expression; Hyperplasia; In Situ Hybridization; Ligands; Protein Serine-Threonine Kinases; Rats; Rats, Sprague-Dawley; Receptor, Transforming Growth Factor-beta Type II; Receptors, Transforming Growth Factor beta; RNA, Messenger; Solubility; Transforming Growth Factor beta; Tunica Intima

The vitronectin receptor (alphavbeta3) mediates several biological processes that are critical to the formation of a neointima after coronary interventions. Blockade of alphavbeta3 could reduce neointima formation by inhibiting smooth muscle cell migration, decreasing transforming growth factor-beta1 expression, enhancing apoptosis, or reducing neovasculature. The effects of short-term administration of Vitaxin, a humanized monoclonal antibody to alphavbeta3, on the responses to balloon injury were tested in hyperlipidemic rabbits. Balloon angioplasty was performed on the iliac arteries of male New Zealand White rabbits that were fed an atherogenic diet for 1 week before injury and until euthanization at 4 weeks. Rabbits were given either saline (control) or 1 of 2 dosing regimens of Vitaxin (high dose, 5.0 mg/kg, and low dose, 0.5 mg/kg), which were administered intra-arterially before injury and intramuscularly on days 2 and 3. High-dose and low-dose Vitaxin were equally effective in decreasing neointima formation even in the presence of hypercholesterolemia, a stimulus to alphavbeta3 expression. Vitaxin reduced transforming growth factor-beta1 and enhanced apoptosis in injured arteries. Despite these positive effects, Vitaxin administration was associated with a reduction in artery size, indicating a negative effect on remodeling. Vitaxin has a potential role in preventing intimal hyperplasia, especially if the negative effects on remodeling can be overcome, by dose adjustment or other strategies.

Topics: Angioplasty, Balloon; Animals; Antibodies, Monoclonal; Antibodies, Monoclonal, Humanized; Apoptosis; Cell Movement; Cholesterol; Fluorescent Antibody Technique; Humans; Hypercholesterolemia; Hyperplasia; Iliac Artery; Male; Rabbits; Receptors, Vitronectin; Transforming Growth Factor beta; Tunica Intima

Exposure to certain UDP-glucuronosyltransferase (UDP-GT) inducers leads to follicular cell hyperplasia, and ultimately thyroid gland tumors. These compounds decrease thyroid hormones, which increases serum concentrations of thyroid stimulating hormone (TSH). This induction of TSH enhances thyroid-follicular cell proliferation. In addition, treatment with classical goitrogenic compounds, such as propylthiouracil (PTU) and methimazole (MMI), induces TGF-beta1 in thyroid-follicular cells, presumably through increased TSH. In other tissues, increases in TGF-beta1 induce apoptosis, a particular form of programmed cell death. In this experiment, we sought to determine whether the UDP-GT inducers, phenobarbital (PB) and pregnenolone-16alpha-carbonitrile (PCN) modulate thyroid-follicular cell apoptosis. If so, are the induction of apoptosis and TGF-beta1 possibly linked? An additional group of rats treated with the thyroid goitrogen, PTU was included. Male Sprague-Dawley rats were treated with thyroid hormone disrupting doses of PB, PCN, or PTU for 3, 7, 14, 21, 28, 45, or 90 days. In this study, PTU treatment increased apoptosis and TGF-beta1 immunoreactive thyroid-follicular cells. PTU treatment of rats produced both a large increase number of TGF-beta1-positive cells (detected by immunohistochemistry), and apoptotic thyroid-follicular cells (detected by morphology). In PB- and PCN-treated rats, a moderate increase in apoptosis coincided with similar increases in TGF-beta1 immunoreactive thyroid-follicular cells. In summary, PB and PCN increase apoptosis and the percentage of TGF-beta1 positive thyroid-follicular cells. Thus, treatment with UDP-GT-inducing chemicals may increase the expression of TGF-beta1 and apoptosis in the thyroid to compensate for the thyroid hypertrophy and hyperplasia.

Topics: Animals; Apoptosis; Enzyme Induction; Glucuronosyltransferase; Hyperplasia; Male; Phenobarbital; Pregnenolone Carbonitrile; Rats; Rats, Sprague-Dawley; Thyroid Gland; Transforming Growth Factor beta

Previous attempts to establish transgenic mouse models to study the functions of transforming growth factor beta1 (TGFbeta1) in the skin revealed controversial roles for TGFbeta1 in epidermal growth (inhibition vs. stimulation) and resulted in neonatal lethality in one instance. To establish a viable transgenic model for studying functions of TGFbeta1 in the skin, we have now developed transgenic mice, which allow focal induction of the TGFbeta1 transgene in the epidermis at different expression levels and at different developmental stages. This system, termed "gene-switch," consists of two transgenic lines. The mouse loricrin vector targets the GLVPc transactivator (a fusion molecule of the truncated progesterone receptor and the GAL4 DNA binding domain), and a thymidine kinase promoter drives the TGFbeta1 target gene with GAL4 binding sites upstream of the promoter. These two transgenic lines were mated to generate bigenic mice, and TGFbeta1 transgene expression was controlled by topical application of an antiprogestin. On epidermal-specific induction of the TGFbeta1 transgene, the BrdUrd labeling index in the transgenic epidermis decreased 6-fold compared with controls. Induction of the TGFbeta1 transgene expression also caused epidermal resistance to phorbol 12-myristate 13-acetate-induced hyperplasia, with a reduction in both epidermal thickness and BrdUrd labeling compared with those in controls. In addition, TGFbeta1 transgene expression induced an increase in angiogenesis in the dermis. Given that the TGFbeta1 transgene can affect both the epidermis and dermis, this transgenic model will provide a useful tool for studying roles of TGFbeta1 in wound-healing and skin carcinogenesis in the future.

Topics: Animals; Bromodeoxyuridine; DNA-Binding Proteins; Epidermis; Estrenes; Fungal Proteins; Gene Expression Regulation, Developmental; Hyperplasia; Membrane Proteins; Mice; Mice, Transgenic; Neovascularization, Physiologic; Promoter Regions, Genetic; Receptors, Progesterone; RNA, Messenger; Saccharomyces cerevisiae Proteins; Tetradecanoylphorbol Acetate; Transcription Factors; Transcriptional Activation; Transforming Growth Factor beta; Wound Healing

The transforming growth factor beta (TGF-beta) pathway is known to play an important role in both human and urine colon cancer. However, the staging, ligand specificity, and mechanism underlying the tumor suppressive activity of this pathway are unknown. We developed a mouse model for colon cancer that identifies an early role for TGF-beta1 in tumor suppression and implicates TGF-beta2 or TGF-beta3 in the prevention of metastasis. Analysis of the development of colon cancer in TGF-beta1 knockout mice pinpoints the defect to the hyperplasty/adenoma transition and reveals that the mechanism involves an inability to maintain epithelial tissue organization and not a loss of growth control, increased inflammatory activity, or increased genetic instability. These mice provide a unique opportunity to investigate the specific role of TGF-beta1 at this critical transition in the development of colon cancer.

Topics: Adenocarcinoma; Adenoma; Adenomatous Polyposis Coli Protein; Animals; Apoptosis; beta Catenin; Biomarkers; Cecum; Cell Division; Cell Transformation, Neoplastic; Colon; Colonic Neoplasms; Crosses, Genetic; Cytoskeletal Proteins; Disease Progression; DNA; DNA-Binding Proteins; DNA, Neoplasm; Genes, APC; Genetic Predisposition to Disease; Humans; Hyperplasia; Inflammation; Intestinal Mucosa; Mice; Mice, Knockout; Microsatellite Repeats; Neoplasm Metastasis; Nuclear Proteins; Specific Pathogen-Free Organisms; Trans-Activators; Transforming Growth Factor beta

To investigate how glutathione-S-transferase placental form (GST-P)+ hyperplastic nodules (HNs) are selected and to determine the driving force for progression or regression of HNs, changes in transforming growth factor-beta1 (TGF-beta) and its receptors were examined during hepatocarcinogenesis initiated by N-nitrosodiethylamine (DEN) and promoted by nodularin. The induction of TGF-beta1 expression in the GST-P+ HNs was dependent on nodularin injections for 10 wk, which started the third week after DEN initiation. The kinetics of TGF-beta1 induction during carcinogenesis were quite different from that of simple regeneration after partial hepatectomy (PH): hepatocytes initiated with DEN alone induced TGF-beta1 expression for 24 d, and subsequent stimulation by PH on the fourteenth day after DEN initiation super-induced TGF-beta1 mRNA (50 times that of the control level), as opposed to a transient expression for less than 5 d by PH alone. GST-P+ HNs did not express TGF-beta receptors I (RI) and II (RII) during the early stage of carcinogenesis, whereas the surrounding hepatocytes strongly expressed both of these receptors. On cessation of nodularin injection, however, the expression of RI and RII in the HNs changed significantly: RII+ nodules appeared, and the number and area of RII+/- nodules were significantly increased at 10 wk after the cessation. These findings indicate that induction of TGF-beta expression in GST-P+ HNs might be a strong selection pressure that allows outgrowth of RII- nodules during liver carcinogenesis.

Topics: Alkylating Agents; Animals; Blotting, Northern; Diethylnitrosamine; Gene Expression Regulation, Neoplastic; Glutathione Transferase; Hyperplasia; Immunohistochemistry; Kinetics; Liver; Liver Neoplasms, Experimental; Male; Peptides, Cyclic; Precancerous Conditions; Rats; Rats, Inbred F344; Receptors, Transforming Growth Factor beta; Time Factors; Transforming Growth Factor beta

Transforming growth factors-beta (TGF-betas) regulate mammary epithelial cell division. Loss of expression of TGF-beta receptor II (TGF-beta-RII) is related to cell proliferation and tumor progression. Breast epithelial hyperplastic lesions lacking atypia (EHLA) are associated with a mild elevation in breast cancer risk. We investigated the expression of TGF-beta-RII in EHLA and the risk of subsequent invasive breast cancer.. We conducted a nested case-control study of women with biopsy-confirmed EHLA who did not have a history of breast cancer or atypical hyperplasia of the breast. Case patients (n = 54) who subsequently developed invasive breast cancer were matched with control patients (n = 115) who did not. Formalin-fixed, paraffin-embedded sections of breast biopsy specimens of all 169 patients with EHLA were studied by immunohistochemical analysis with antibodies against TGF-beta-RII. All P values are two-sided.. Women with breast EHLA and 25%-75% TGF-beta-RII-positive cells or less than 25% TGF-beta-RII-positive cells had odds ratios of invasive breast cancer of 1.98 (95% confidence interval [CI] = 0.95-4.1) or 3.41 (95% CI = 1.2-10.0), respectively (P for trend =.008). These risks are calculated with respect to women with EHLA that had greater than 75% TGF-beta-RII expression. Women with a heterogeneous pattern of TGF-beta-RII expression in their normal breast lobular units and either greater than 75%, 25%-75%, or less than 25% positive cells in their EHLA had odds ratios for breast cancer risk of 0.742 (95% CI = 0.3-1.8), 2.85 (95% CI = 1.1-7.1), or 3.55 (95% CI = 1.0-10.0), respectively (P for trend =.003). These risks are relative to women with a homogeneous pattern of expression in their normal lobular units and greater than 75% positive cells in their EHLA.. This study indicates that loss of TGF-beta-RII expression in epithelial cells of EHLA is associated with increased risk of invasive breast cancer.

Topics: Adult; Aged; Breast; Breast Neoplasms; Carcinoma, Ductal, Breast; Case-Control Studies; Cell Division; Disease Progression; Epithelium; Female; Follow-Up Studies; Gene Expression; Humans; Hyperplasia; Immunohistochemistry; Middle Aged; Odds Ratio; Risk; Transforming Growth Factor beta

To investigate the potential role of Transforming Growth Factor beta 1 (TGF beta 1) in the pathogenesis of chronic allograft rejection, we studied TGF beta 1 expression in a rat aortic allograft model. mRNA and protein expression of total and endogenously active TGF beta 1 were analysed in infra-renal orthotopic aortic syngeneic and allogeneic grafts and matched with the histological appearances of the grafts, 2, 4 and 12 weeks post-transplantation. Serum levels of TGF beta 1 were also measured. The level of TGF beta 1 m RNA and protein expression appeared highest 2 and 4 weeks following transplantation in both syngeneic and allogeneic grafts, with significantly elevated levels of mRNA expression in the 2 week allograft specimens. These time-points correlate histologically with maximal inflammatory cell infiltration of the grafts. By 12 weeks post-transplantation, TGF beta 1 mRNA expression is reduced in allogeneic grafts compared to syngeneic grafts. However, detectable levels of total and endogenously active TGF beta 1 protein levels in the allografts exceed those measured in the syngeneic grafts at this time point. These results demonstrate the complex expression pattern of this growth factor during the progression of chronic rejection and suggest an aetiological link between TGF beta 1 and the process of accelerated graft atherosclerosis.

Topics: Animals; Aorta, Abdominal; Chronic Disease; Enzyme-Linked Immunosorbent Assay; Gene Expression Regulation; Graft Rejection; Hyperplasia; Inflammation; Rats; Rats, Inbred ACI; Rats, Inbred Lew; RNA, Messenger; Time Factors; Transforming Growth Factor beta; Transplantation, Homologous; Tunica Intima

We examined the relative contributions of foreign body granulation and de novo lesions to neointimal hyperplasia in atherectomized specimens of restenosis after coronary stenting.. Clinicopathological studies have suggested that smooth muscle cell (SMC) hyperplasia is the most likely cause of restenosis after coronary stenting. It is not yet fully understood how SMC hyperplasia occurs or how SMCs stimulation can lead to intimal hyperplasia. Although inflammation has been postulated to be a major contributor to restenosis after coronary stenting, there is a paucity of data on the relationsip between inflammation and subsequent neointimal formation in humans. Only in a porcine experimental model of stent restenosis, foreign body granulation tissue as a cause of inflammation in stent restenosis was reported.. Tissue specimens were retrieved by directional atherectomy from 11 patients in whom stent restenosis developed after percutaneous revascularization of coronary artery disease. For specimens preserved in 10% buffered formalin, analysis of cellular composition was performed quantitatively after cell-specific immunostaining, i.e. CD68, UCHL-1, HLA-DR, smooth muscle actin, vimentin, desmin, PCNA and TGF-beta.. Five of the 11 patients showed granulation tissues 3-6 months after stent implantation, of whom 3 patients revealed foreign body multinucleated giant cells around the stent struts where PCNA- and vimentin-positive SMCs were demonstrated. Calcification and de novo lesions in medial and adventitial tissues were observed in 3 other patients, and fresh and/or organized thrombi were documented in 3 of the 11 patients.. These findings support the notion that stent restenosis results from SMC hyperplasia and suggest that the foreign body granulation tissue against metals of the stents and de novo lesions could play an important role in chronic inflammation leading to intimal hyperplasia and subsequently to stent restenosis in some patients. Clinicians should thus consider whether a patient may be allergic to stent components with unknown reaction, e.g. haptens.

Topics: Aged; Atherectomy, Coronary; B-Lymphocytes; Calcinosis; Coronary Angiography; Coronary Disease; Female; Giant Cells, Foreign-Body; Graft Occlusion, Vascular; Granuloma, Foreign-Body; HLA-DR Antigens; Humans; Hyperplasia; Immunoenzyme Techniques; Male; Microfilament Proteins; Middle Aged; Muscle, Smooth, Vascular; Proliferating Cell Nuclear Antigen; Recurrence; Reoperation; Stents; T-Lymphocytes; Transforming Growth Factor beta; Tunica Intima

Transforming growth factor (TGF)-beta1 and TGF-beta3 are normally expressed at high levels in the mammary gland during quiescence and at all stages of development, except lactation. Exogenously added TGF-beta1, -beta2, and -beta3 have been shown to regulate growth and differentiation of mammary epithelial cells in vitro and in vivo. TGF-betas signal through a heteromeric complex of type I and type II serine/threonine kinases. The type II receptor is necessary for ligand binding and growth suppression by TGF-betas. Deletions of the cytoplasmic domains of several kinase receptors known to function in multimeric complexes have been shown to act as dominant-negative mutations. To evaluate the role of endogenous TGF-betas in the growth and differentiation of the mammary gland in vivo, we have targeted expression of a truncated, kinase-defective TGF-beta type II receptor to mammary epithelial cells in transgenic mice using the mouse mammary tumor virus promoter/enhancer. Transgene expression was localized to the epithelial cells of terminal ducts and alveolar buds. At approximately 20 weeks of age, virgin female transgenic mice demonstrated varying degrees of mammary epithelial hyperplasia. Mammary glands from transgenic, virgin animals exhibited alveolar development and expression of the milk protein, beta-casein. The data suggest that impaired responsiveness in the epithelium to endogenous TGF-betas results in inappropriate alveolar development and differentiation in the mammary gland. We conclude that endogenous TGF-betas signal to the epithelium to maintain quiescence in the mammary glands of virgin animals.

Topics: Animals; Caseins; Cell Differentiation; Cell Division; Diestrus; Epithelial Cells; Female; Gene Targeting; Genes, Dominant; Humans; Hyperplasia; Mammary Glands, Animal; Matrix Metalloproteinase 3; Mice; Mice, Transgenic; Protein Serine-Threonine Kinases; Receptor, Transforming Growth Factor-beta Type II; Receptors, Transforming Growth Factor beta; Recombinant Fusion Proteins; RNA, Messenger; Sequence Deletion; Transforming Growth Factor beta

The purpose of this study was to determine the correlation between progression and regression of myointimal hyperplasia (MH) and cytokine production in experimental vein grafts. Although the autologous vein is the best suitable bypass conduit for reconstruction of peripheral arteries, at the end of the first year thrombosis in the coronary and lower extremity circulation ranges from 20% to 50%. Many of these failures are caused by MH.. In 76 inbred Lewis rats, a 1 cm long segment of inferior vena cava was inserted at the level of the abdominal aorta. The segments of inferior vena cava were obtained from syngeneic Lewis rats. In 56 animals the arterial vein graft was explanted 3 days (n = 10), 7 days (n = 10), 4 weeks (n = 26), and 12 weeks (n = 10) after operation. In 20 animals the vein graft was explanted 4 weeks after being in the arterial system and reimplanted as iliac venovenous bypass in syngeneic Lewis rats. These grafts were explanted 2 weeks (n = 10) and 8 weeks (n = 10) later. Grafts were analyzed by light and electron microscopy, morphometric study, and histochemical analysis and were put in an organ culture to assess cytokine production.. We observed MH formation in arterial vein grafts and MH regression in reimplanted vein grafts (p < 0.001). MH formation was correlated with production of platelet-derived growth factor, basic fibroblast growth factor, interleukin-1, and tumor necrosis factor-alpha. MH regression was correlated with transforming growth factor-beta 1 production.. On the basis of the results of our study, we conclude that MH formation in experimental vein grafts depends on production of platelet-derived growth factor, basic fibroblast growth factor, interleukin-1, and tumor necrosis factor-alpha, and MH regression depends on transforming growth factor-beta 1 production. Cytokine therapy may represent a valuable new treatment to prevent vein bypass failures caused by MH.

Topics: Animals; Aorta, Abdominal; Cytokines; Hyperplasia; Interleukin-1; Male; Organ Culture Techniques; Platelet-Derived Growth Factor; Rats; Rats, Inbred Lew; Transforming Growth Factor beta; Transplantation, Heterologous; Transplantation, Isogeneic; Tumor Necrosis Factor-alpha; Tunica Intima; Vascular Surgical Procedures; Vena Cava, Inferior

Increased smooth muscle mass due to hyperplasia and hypertrophy of airway smooth muscle (ASM) cells is a common feature in asthma. Angiotensin II (Ang II), a potent vasoconstrictor and mitogen for a wide variety of cells, has recently been implicated in bronchoconstriction in asthmatics. However, a possible mitogenic role as well as underlying molecular mechanisms of this octapeptide in human ASM cells are not yet known. We studied the effects of Ang II on ASM cell proliferation and growth and on the expression of three transcription factors, egr-1, c-fos, and c-jun, as well as a cytokine, transforming growth factor-beta1 (TGF-beta1). Human ASM cells were isolated by enzymatic digestion of bronchial smooth muscle obtained from lung resection tissue. Confluent cells were growth-arrested and subsequently incubated with Ang II (100 nM) for different time periods and processed for the measurement of cell growth and gene expression. Ang II significantly induced DNA and protein synthesis in human ASM cells at 8 h, resulting in a net increase in the accumulation of protein over DNA (i.e., cellular hypertrophy) at 16 h of incubation. Cell counts and MTT-reduction assay, however, showed no increase in cell number as a result of Ang II stimulation. Ang II stimulated the expression of egr-1 and c-fos as early as 15 min, reaching maximum levels at 45 min, whereas the expression of c-jun peaked at 2 h of Ang II exposure. Furthermore, steady-state mRNA levels of TGF-beta1 were upregulated by Ang II after 4 h and reached peak levels at 16 h of incubation. Secretion of biologically active TGF-beta1 from human ASM cells was significantly (P <= 0.02) enhanced by Ang II incubation after 8 h, which remained elevated until 24 h. Our results suggest that the Ang II-induced transient early expression of transcription factors may regulate autocrine genes like TGF-beta1, of which the subsequent late upregulation could contribute to cellular hypertrophy during, for example, airway remodeling in asthma.

Topics: Angiotensin II; Blotting, Northern; Bronchi; Cells, Cultured; DNA-Binding Proteins; Early Growth Response Protein 1; Humans; Hyperplasia; Hypertrophy; Immediate-Early Proteins; Immunohistochemistry; Muscle, Smooth; Proto-Oncogene Proteins c-fos; Proto-Oncogene Proteins c-jun; RNA, Messenger; Time Factors; Transcription Factors; Transforming Growth Factor beta

Topics: Adolescent; Adult; Aged; Cells, Cultured; Child; Female; Humans; Hyperplasia; Interferon-gamma; Interleukin-4; Leukocytes, Mononuclear; Male; Middle Aged; Myasthenia Gravis; Reference Values; Thymoma; Thymus Gland; Thymus Neoplasms; Transforming Growth Factor beta

Transforming growth factor beta1 (TGF-beta1) has been implicated as inhibitor of cell proliferation and a potent inducer of apoptosis in vitro and in vivo after the administration of high doses. To assess the role of endogenous TGF-beta1, we quantitated the cytokine and its receptors in rat liver during regenerative and hyperplastic growth, regression by apoptosis, and in hepatocellular carcinoma (HCC). This was accomplished by Northern blot analysis and by RNase protection assay of the messenger RNA (mRNA) of TGF-beta1 and TGF-beta receptors (TbetaR) types I to III and by an activity bioassay of the TGF-beta proteins. Untreated rat livers were found to contain 15.6 +/- 4.8 ng TGF-beta1 protein/g tissue; TGF-beta2 protein was not detected. To induce toxic cell death and subsequent regenerative DNA synthesis in the liver, rats were treated with a necrogenic dose of carbon tetrachloride (CCl4). After 24 and 48 hours, there was an upregulation of TGF-beta1 (mRNA, up to tenfold; protein, about twofold) and of TbetaRs (mRNA: two- to fourfold); that indicates an overall enhanced production of and sensitivity to TGF-beta1, which may serve to confine the regenerative response. Hyperplastic liver growth and regression of the hyperplasia were induced by treatment with cyproterone acetate (CPA) or nafenopin (NAF) followed by withdrawal; neither mRNAs of TGF-beta1 and TbetaR types I to III nor TGF-beta1 protein exhibited significant changes during the growth phase or during regression by apoptosis. We also studied neoplastic growth. HCC, obtained after long-term treatment with NAF, exhibited high rates of cell replication and apoptosis. The majority of lesions contained mRNA and protein of TGF-beta1 and mRNA of TbetaR types I to III at concentrations similar to those of the surrounding tissue. In conclusion, during liver regeneration there is a pronounced upregulation of expression of both TGF-beta1 and TbetaRs I to III, but not during mitogen-induced liver growth or regression. It appears that apoptosis is induced via altered local concentration of TGF-beta1, in a paracrine and/or autocrine way. By this mechanism the lethal effects of TGF-beta1 may be locally confined, and overshoots of apoptosis in the liver may be prevented.

Topics: Animals; Apoptosis; Carbon Tetrachloride; Female; Hyperplasia; Liver; Liver Neoplasms, Experimental; Lymphotoxin beta Receptor; Male; Rats; Rats, Wistar; Receptors, Tumor Necrosis Factor; RNA, Messenger; Transforming Growth Factor beta

Hepatocyte apoptosis occurs during involution of hyperplastic liver induced by administration of xenobiotic compounds in rats. With this hyperplasia and involution, hepatic transforming growth factor (TGF)-beta1 is reported to be expressed to stimulate hepatocyte apoptosis. In regenerating liver after partial resection showing no hyperplasia, such expression of TGF-beta1 is also seen. However, no hepatocyte apoptosis develops despite the high levels of TGF-beta1. When rats received an intravenous injection of human hepatocyte growth factor at 12 h intervals for 14 days, the hepatic DNA content was increased 12 h after the last injection to 140% of control. This DNA content was significantly decreased at 108 and 180 h after discontinuation of treatment. At 60 h after the last injection, the number of apoptotic bodies positive for nick end-labelling of DNA in hepatocytes was significantly greater in treated rats than in control rats. Hepatocyte apoptosis was also identified electron micrographically. Hepatic TGF-beta1 mRNA levels in treated rats were significantly lower than in control rats at 12 h and then gradually increased towards control levels. We conclude that hyperplastic liver induced in normal rats by hepatocyte growth factor regresses with hepatocyte apoptosis and suppressed hepatic TGF-beta1 mRNA levels.

Topics: Animals; Apoptosis; DNA; Hepatocyte Growth Factor; Hyperplasia; Liver; Male; Rats; Rats, Sprague-Dawley; RNA, Messenger; Transforming Growth Factor beta

To examine whether synoviocytes from patients with rheumatoid arthritis (RA) have a stronger growth ability than those from patients with osteoarthritis (OA), and to determine whether these synoviocytes clonally expand in situ.. Synovial tissues from 13 RA patients and 4 OA patients were cultured, and their ability to form colonies in soft agarose was examined. RA and OA synoviocytes were also examined in varying concentrations of fetal calf serum (FCS)-containing medium to test the effects of FCS on colony formation. DNA was extracted from clones with colony-forming ability in nonpannus lesions and from synoviocytes in pannus lesions. Restriction fragment length polymorphism (RFLP) analysis was used to examine phosphoglycerate kinase 1 (PGK-1) gene patterns. Production of cytokines by these cells was also assessed.. All 13 RA synoviocytes exhibited colony formation, whereas none of the 4 OA synoviocytes did. This tendency was also seen with all of the concentrations of FCS examined, although growth varied in a dose-dependent manner. In contrast to OA synovial clones, cloned RA synoviocytes obtained from colonies exhibited a partial RFLP PGK-1 gene pattern, suggesting that the clones originated from monoclonal cells. Of note, 3 of 7 noncloned synoviocytes from pannus lesions exhibited a monoclonal pattern. Pannus cells produced high levels of transforming growth factor beta and platelet-derived growth factor.. These findings suggest that synoviocytes with a strong growth ability are present in the rheumatoid synovium, and that these cells expand monoclonally, particularly in pannus lesions.

Topics: Adult; Aged; Arthritis, Rheumatoid; Cartilage; Cell Division; Clone Cells; Gene Expression Regulation, Enzymologic; Humans; Hyperplasia; Male; Middle Aged; Osteoarthritis; Phosphoglycerate Kinase; Platelet-Derived Growth Factor; Polymorphism, Restriction Fragment Length; Stem Cells; Synovial Membrane; Transforming Growth Factor beta

The role of estrogens in providing atheroprotection has been well documented in both epidemiologic and experimental studies. This phenomenon has traditionally been attributed to the beneficial lipid-modifying effects of estrogens. Previous studies have used models of either diet- or injury-induced atherosclerosis. As such, the interrelationship between estrogens, lipids, and atherosclerosis remains unclear. We hypothesized that estrogens are atheroprotective independent of changes in serum lipids by directly influencing the accumulation and effects of the peptide growth factor transforming growth factor beta1 (TGF-beta1).. Thirteen female sheep (8 years old) were randomized to sham, ovariectomy, or ovariectomy with 17beta-estradiol replacement. Serum lipid levels were serially measured. At 9 months, necropsy was performed with histologic morphometric analysis of the aortoiliac bifurcation. Levels of TGF-beta1 were determined in serum and aortic tissue. Human aortic smooth muscle cells were isolated and cultured.. Serum triglyceride, lipoprotein a, and total, low-density lipoprotein, and high-density lipoprotein cholesterol levels were similar and normal between groups. Ovariectomy resulted in aortoiliac intimal hyperplasia compared with sham (P < 0.001) and hormone replacement (P < 0.001) groups. Compared with ovariectomy, estrogen replacement attenuated aortic accumulation of TGF-beta1 (P < 0.02). In vitro, estradiol potentiated TGF-beta1 inhibition of human vascular smooth muscle cell (VSMC) proliferation and increased TGF-beta1 release in stimulated VSMCs (P < 0.001).. Without dietary manipulation, ovarian ablation induces aortic intimal hyperplasia in the ewe. Estradiol abrogates this response independently of its effects on serum lipids. Hormone replacement decreases the accumulation of TGF-beta1, suggesting that estrogens may provide atheroprotection both by modifying local production and by modulating the influence of TGF-beta1 on VSMC growth.

Topics: Animals; Aorta; Arteriosclerosis; Cell Division; Estradiol; Estrogen Replacement Therapy; Female; Humans; Hyperplasia; Lipids; Luteinizing Hormone; Muscle, Smooth, Vascular; Ovariectomy; Sheep; Transforming Growth Factor beta

The distribution of the three isoforms of transforming growth factor (TGF)-beta (TGF-beta 1, -beta 2, and -beta 3) as well as their signaling receptor, TGF-beta type I receptor (T beta R-I), in bronchiolo-alveolar cell hyperplasia (BAH) and bronchiolo-alveolar carcinoma (BAC) was examined by immunohistochemistry using specific antibodies. The results showed that the expression of one or more TGF-beta subtypes occurred in a large proportion of BAC and BAH tissues. The expression of TGF-beta 1 and TGF-beta 3 was increased in tumor cells compared with hyperplastic cells. BAC with expression of two or three ligands was associated with a higher incidence of lymph node metastasis. These results suggest that hyperplastic cells and cancer cells derived from bronchiolo-alveolar cell can synthesize and secrete TGF-beta isoforms and its receptor T beta R-I which may contribute to the progression of BAC in part.

Topics: Adenocarcinoma, Bronchiolo-Alveolar; Adult; Aged; Bronchi; Female; Humans; Hyperplasia; Lung Neoplasms; Male; Middle Aged; Pulmonary Alveoli; Receptors, Transforming Growth Factor beta; Transforming Growth Factor beta

Despite extensive research, the mechanisms of prostate carcinogenesis are not well understood. The slow progress in this area is due, at least in part, to lack of a suitable animal model for prostate carcinogenesis. We have developed an animal model, based on the existing sex hormone-induced prostate carcinogenesis in the Noble rat, by substantially increasing the dosage of testosterone while keeping the level of estrogen unchanged. Using the modified method of combination of testosterone and estradiol-17beta (T+E2), it has been shown in Noble rats that prostate carcinogenesis followed a multi-step process involving hyperplasia, dysplasia, and carcinoma. We have demonstrated the importance of TGF-alpha, TGF-beta1 and bFGF in the development of prostate carcinogenesis. This study also established the roles of VEGF and IGF-1, initially as paracrine factors in epithelial-stromal interactions during the process of carcinogenesis and subsequently switching over to an autocrine mode during the establishment of carcinoma.

Topics: Adenocarcinoma; Animals; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Therapy, Combination; Endothelial Growth Factors; Estradiol; Fibroblast Growth Factor 2; Hyperplasia; Immunohistochemistry; Lymphokines; Male; Precancerous Conditions; Prostate; Prostatic Neoplasms; Rats; Rats, Inbred Strains; Receptor Protein-Tyrosine Kinases; Receptors, Growth Factor; Receptors, Vascular Endothelial Growth Factor; Testosterone; Transforming Growth Factor alpha; Transforming Growth Factor beta; Transforming Growth Factor beta1; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factors

The transforming growth factor-beta (TGF-beta) superfamily encompasses a large group of growth and differentiation factors playing important roles in regulating embryonic development and in maintaining tissue homeostasis in adult animals. Using degenerate polymerase chain reaction, we have identified a new murine TGF-beta family member, growth/differentiation factor-8 (GDF-8), which is expressed specifically in developing and adult skeletal muscle. During early stages of embryogenesis, GDF-8 expression is restricted to the myotome compartment of developing somites. At later stages and in adult animals, GDF-8 is expressed in many different muscles throughout the body. To determine the biological function of GDF-8, we disrupted the GDF-8 gene by gene targeting in mice. GDF-8 null animals are significantly larger than wild-type animals and show a large and widespread increase in skeletal muscle mass. Individual muscles of mutant animals weigh 2-3 times more than those of wild-type animals, and the increase in mass appears to result from a combination of muscle cell hyperplasia and hypertrophy. These results suggest that GDF-8 functions specifically as a negative regulator of skeletal muscle growth.

Topics: Aging; Amino Acid Sequence; Animals; Body Weight; CHO Cells; Cloning, Molecular; Cricetinae; Embryo, Mammalian; Gene Targeting; Homozygote; Humans; Hyperplasia; Hypertrophy; In Situ Hybridization; Mice; Mice, Inbred C57BL; Molecular Sequence Data; Muscle, Skeletal; Myostatin; Polymerase Chain Reaction; Protein Sorting Signals; Stem Cells; Transforming Growth Factor beta

Barrier-raised transforming growth factor beta 1 (TGF beta 1)-deficient mice consistently die before 35 days of age of a severe multiorgan inflammatory disease that can affect the skeletal muscle, heart, liver, pancreas, salivary gland, lung, oesophagus and stomach. The underlying cause of this disease is not known. To determine whether abnormal responsiveness of the immune system to the presence of enteric flora plays a causative role, a colony of TGF beta 1-deficient and wild-type mice were raised in a sterile environment. Seven germ-free TGF beta 1-deficient and 5 germ-free TGF beta 1 wild-type mice were examined. Lesion development was analysed and compared with historical data on 50 barrier-raised TGF beta 1 mutant mice and 32 barrier-raised wild-type mice. All germ-free TGF beta 1-deficient mice died shortly after weaning, as do their barrier-raised counterparts. There was a significant delay in death in germ-free TGF beta 1-deficient mice compared with barrier-raised mutant mice. However, there was no difference in the type, severity or incidence of lesions between TGF beta 1 mutant mice raised under germ-free or barrier conditions. Germ-free wild-type mice had no lesions. It is concluded that microorganisms play a minimal role in disease induction in TGF beta 1-deficient mice.

Topics: Animals; Germ-Free Life; Hyperplasia; Inflammation; Longevity; Mice; Mice, Inbred Strains; Mice, Mutant Strains; Stomach; Stomach Ulcer; Transforming Growth Factor beta; Ulcer

The multifunctional cytokine, transforming growth factor beta 1 (TGF-beta), plays an important role in the development of injury-associated intimal hyperplasia (IH). Strategies to suppress local TGF-beta activity may have a clinical potential to prevent restenosis caused by IH. Photodynamic therapy (PDT) involves the local generation of cytotoxic free radicals by light activation of photosensitizer dyes and has been shown to inhibit experimental IH. This study investigated whether PDT-generated free radicals can affect TGF-beta activity in a biologic system using vascular smooth muscle cells (SMCs).. The release and activation of TGF-beta by injured SMCs in culture was compared between mechanical injury and PDT. Mechanical injury was induced with a rubber policeman, and PDT was performed with the photosensitizer chloroaluminum sulfonated phthalocyanine (5 micrograms/ml) and 675 nm laser light at subtherapeutic 10 J/cm2 and the in vivo therapeutic dose of 100 J/cm2. Cell viability was assessed by the tetrazolium salt conversion assay, and active and total (active + latent) TGF-beta was determined by enzyme-linked immunosorbent assay in the conditioned media of SMCs 24 hours after treatment. Functional TGF-beta activity was assessed by inhibition of endothelial cell mitogenesis.. Both forms of injury severely reduced (p < 0.0005) SMC viability to less than 15%. In untreated SMC conditioned media, only 14.5% of the total TGF-beta was active (27.7 +/- 8.7 pg per 1 x 10(5) cells). However, after mechanical injury and PDT with 10 J/cm2, there was a significant increase (p < 0.02) in active TGF-beta (60.1 +/- 10.1 pg and 48.6 +/- 21.0 pg, respectively), despite a total reduction of approximately 50%. In contrast to this result, PDT with 100 J/cm2 did not result in increased levels of active TGF-beta (8.1 +/- 3.5 pg), despite having similar levels of total TGF-beta. Consequently, the conditioned media of SMCs that had 100 J/cm2 PDT did not inhibit endothelial cell mitogenesis as compared with the conditioned media of SMCs with mechanical injury and 10 J/cm2 PDT (p < 0.0002).. This report describes two novel findings: (1) injury to SMCs in vitro induces the conversion of biologically latent TGF-beta to active TGF-beta; and (2) the therapeutic PDT dose interferes with this injury activation process. This study substantiates the concept of local cytokine inhibition by PDT in a biologic system and provides new insights into the mechanisms of PDT-mediated inhibition of experimental IH.

Topics: Aluminum; Animals; Cattle; Cell Division; Cells, Cultured; Endothelium, Vascular; Free Radicals; Hyperplasia; In Vitro Techniques; Indoles; Muscle, Smooth, Vascular; Organometallic Compounds; Photochemotherapy; Photosensitizing Agents; Transforming Growth Factor beta; Tunica Intima

The success of coronary reconstructive procedures is limited by the high incidence of restenosis secondary to intimal hyperplasia (IH). Transforming growth factor-beta 1 (TGF-beta 1) is a growth factor which has been shown to be important in the early development of IH in arteries and peripheral vein grafts. To date, there is little information concerning the early remodeling in aortocoronary vein grafts (ACVG). The purpose of this study was to characterize the expression of TGF-beta 1 expression in early aortocoronary vein grafts. Eighteen mongrel dogs underwent aortocoronary vein bypass grafting. Vein grafts were excised at 2 hr, 4 hr, and 7 days after implantation, snap frozen, and processed for ribonuclease protection assays (RPA) using 32P-labeled riboprobes for TGF-beta 1 and 18 S rRNA. TGF-beta 1 expression was quantified by densitometric analysis of autoradiographs which were expressed as a ratio TGF-beta 1/rRNA. Representative vessel rings were also collected for histology. There was a significant rise in TGF-beta 1 expression in the 2-hr vein grafts (0.42 +/- 0.04 compared to control saphenous vein (0.21 +/- 0.05, P < 0.02). In addition, there was significant downregulation of TGF-beta 1 at 4 hr (0.28 +/- 0.05) and at 7 days (0.18 +/- 0.01) when compared to 2 hr (P < 0.05). Histological specimens showed minimal intimal hyperplasia at 7 days. These results show for the first time an acute rise in TGF-beta 1 expression in ACVG. This upregulation quickly subsides by 4 hr and gene expression approaches control values by 7 days. By understanding this temporal relationship of expression one could better target potential therapeutic modalities to attenuate IH.

Topics: Animals; Coronary Artery Bypass; Coronary Circulation; Dogs; Hyperplasia; Saphenous Vein; Time Factors; Transforming Growth Factor beta; Tunica Intima; Veins

Synovial chondromatosis (SC) is an uncommon, benign condition of unknown etiology. A case of SC of the temporomandibular joint (TMJ) with the immunohistochemical findings of transforming growth factor-beta (TGF) and tenascin (TN) is reported. The roles of TGF and TN in SC of TMJ are discussed.

Topics: Adult; Arthroscopy; Cartilage, Articular; Chondromatosis, Synovial; Collagen; Coloring Agents; Endoscopy; Extracellular Matrix; Female; Humans; Hyperplasia; Immunoenzyme Techniques; Joint Loose Bodies; Synovial Membrane; Temporomandibular Joint Disorders; Tenascin; Transforming Growth Factor beta

Radiation enteropathy is characterized by sustained increase in transforming growth factor beta (TGF-beta) immunoreactivity and connective tissue mast cell (CTMC) hyperplasia that may be responsible for progressive fibrosis and lead to clinical complications. We examined to what extent these chronic molecular and cellular phenomena are associated with acute mucosal breakdown (consequential injury) and/or direct (primary) radiation injury in late-responding compartments.. Rat small intestine was exposed to 50.4 Gy x-irradiation given either over 18 days (2.8 Gy daily or 5.6 Gy every other day) or 9 days (2.8 Gy twice daily or 5.6 Gy daily). Intestinal complications were recorded and groups of animals were euthanized at 2 and 26 weeks to assess subacute and chronic injury. Histopathologic changes were assessed with a radiation injury scoring system (RIS), total TGF-beta immunoreactivity was quantified with computerized image analysis, and CTMC hyperplasia was assessed in toluidine blue-stained sections.. TGF-beta immunoreactivity and CTMC hyperplasia colocalized in areas of injury and were highly significantly correlated. Increased fraction size and decreased overall treatment time were associated with increased RIS (p < 0.01 and p < 0.00001), increased TGF-beta immunoreactivity (p = 0.01 andp < 0.001), and degree of CTMC hyperplasia (p = 0.01 and p < 0.001). Postradiation CTMC numbers increased across treatment groups from 2 to 26 weeks (p < 0.01). TGF-beta immunoreactivity was independently associated with chronic intestinal wall fibrosis (p = 0.003).. This in vivo study supports in vitro evidence linking increased TGF-beta immunoreactivity and mast cell hyperplasia and strongly suggests their involvement in the molecular pathogenesis of both primary and consequential radiation enteropathy.

Topics: Animals; Biomarkers; Connective Tissue; Fibrosis; Hyperplasia; Intestine, Small; Male; Mast Cells; Radiation Dosage; Radiation Injuries, Experimental; Rats; Rats, Sprague-Dawley; Transforming Growth Factor beta

Transforming growth factor-beta 1 (TGF-beta 1) has been variably associated with the regulation of cellular proliferation and extracellular matrix expansion after arterial injury. We tested these associations in vivo in the rat carotid injury model. At 0, 3, 7, 14 and 28 days following arterial balloon injury, regional expression of TGF-beta 1 mRNA was assessed using in situ hybridization and the results compared to measures of cellular proliferation and extracellular matrix expansion. Both the TGF-beta 1 concentration measured in culture media of explanted carotid arteries and the quantitative in situ hybridization signal for TGF-beta 1 arterial media and neointima were maximal at 14 days after balloon injury. However, medial cellular proliferation was maximal at 3 days whereas neointimal proliferation was maximal at 14 days and significantly greater than medial proliferation. Neointimal cell density declined significantly between 7 and 14 days, indicating the expansion of extracellular matrix; however, medial cell density was unchanged between 3 and 28 days after balloon injury. Thus, differences in the regional arterial wall relationships between the time course of cellular proliferation, extracellular matrix expansion and the level of TGF-beta 1 expression demonstrate in vivo variability in the response to TGF-beta 1.

Topics: Angioplasty, Balloon, Coronary; Animals; Carotid Arteries; Cell Division; Extracellular Matrix; Gene Expression; Hyperplasia; Male; Muscle, Smooth, Vascular; Organ Culture Techniques; Rats; Rats, Sprague-Dawley; RNA, Messenger; Time Factors; Transforming Growth Factor beta; Wound Healing

Chronic metabolic acidosis induces both hyperplastic and hypertrophic renal growth and is associated with progressive loss of renal function. These studies examine the direct effect of media acidification on the growth of rabbit proximal tubule cells in primary culture. The results demonstrate that media acidification has a direct antiproliferative (hypoplastic) effect on both quiescent and mitogen-stimulated [epidermal growth factor (EGF)-stimulated] cells and does not induce hypertrophy. This direct antiproliferative effect of acid is associated with inhibition of EGF-induced phosphorylation of the retinoblastoma protein (pRB), which maintains pRB activity and inhibits cell cycle progression from G1 to S phase. Transforming growth factor-beta (TGF-beta) alone has an antiproliferative effect in these cells. TGF-beta converts EGF-induced hyperplasia to hypertrophy and inhibits EGF-induced pRB phosphorylation. Media acidification inhibits both the antiproliferative effect of TGF-beta and the ability of TGF-beta to convert EGF-induced hyperplasia to hypertrophy. This activity is associated with inhibition of TGF-beta-mediated retention of pRB in the active, hypophosphorylated state. These results demonstrate that metabolic acidosis has a direct growth-suppressive effect on renal epithelial cells but inhibits the growth-suppressive effects of TGF-beta. Inhibition of the antiproliferative effect of cytokines, such as TGF-beta, may be responsible for acidosis-induced hyperplasia in vivo.

Topics: Animals; Cell Division; Cells, Cultured; Culture Media; Epidermal Growth Factor; Growth Inhibitors; Hydrogen-Ion Concentration; Hyperplasia; Hypertrophy; Kidney Tubules, Proximal; Phosphorylation; Rabbits; Retinoblastoma Protein; Transforming Growth Factor beta

The renin-angiotensin system seems to play an important role in the pathogenesis of renal interstitial fibrosis. However, the potential direct effects of angiotensin II (Ang II) on cultured renal fibroblasts have been little studied. We have observed that rat renal interstitial fibroblasts (NRK 49F cell line) possess AT1 receptors coupled to intracellular calcium mobilization. Exposure of these cells to Ang II induced several short and long growth-related metabolic events mediated by the AT1 receptor, including c-fos gene expression, changes in cell cycle and cell proliferation. Activation of interstitial fibroblasts by Ang II could also contribute to extracellular matrix accumulation. Stimulation with Ang II increased mRNA expression of TGF-beta 1, fibronectin and type I collagen. In fact, Ang II enhanced fibronectin production via AT1 receptors by a process depending on autocrine TGF-beta secretion. The mechanism of some Ang II actions (calcium mobilization and fibronectin production) depended on protein kinase C and tyrosine kinase activation. We further investigated whether renal fibroblasts could express some components of the renin-angiotensin system. These cells constitutively expressed the angiotensinogen gene that was up-regulated by Ang II. Collectively, these results indicate that in renal interstitial fibroblasts Ang II causes hyperplasia and extracellular matrix production via the AT1 receptor. Ang II may initiate a positive feedback regulation of fibroblasts growth, inducing the expression of TGF-beta 1 and angiotensinogen genes. Ang II, acting directly on interstitial fibroblasts, may be implicated in the pathogenesis of renal fibrosis.

Topics: Angiotensin II; Animals; Calcium; Cell Communication; Cell Division; Cells, Cultured; Dose-Response Relationship, Drug; Extracellular Matrix Proteins; Fibroblasts; Fibronectins; Fibrosis; Flow Cytometry; Gene Expression Regulation; Hyperplasia; Hypertrophy; Imidazoles; Kidney; Protein Kinase C; Protein-Tyrosine Kinases; Proto-Oncogene Proteins c-fos; Pyridines; Rats; Receptors, Angiotensin; RNA, Messenger; Transforming Growth Factor beta; Up-Regulation

To determine whether tenidap regulates extracellular matrix metabolism in chronic arthritis.. Antigen arthritis was induced in the knees of 30 rabbits. Animals were distributed into 3 groups: untreated, tenidap-treated, and diclofenac-treated rabbits. Three weeks after disease induction, synovial membranes were extracted and processed for histopathologic examination and detection of type I collagen (CI) and fibronectin (FN) by immunoperoxidase. Simultaneously, we analyzed the in vitro effect of tenidap on healthy synovial cell (SC) proliferation, FN expression and synthesis, and expression of transforming growth factor beta1 (TGFbeta1) messenger RNA.. Untreated animals showed synovial lining hyperplasia, cellular infiltration at the sublining, and increased deposition of matrix proteins. These findings were not apparent in tenidap-treated rabbits, where CI and FN had the same distribution as in healthy synovial membranes. In vitro, tenidap inhibited SC proliferation (> or =25 microM) and down-regulated the expression and synthesis of FN in a dose-dependent manner (> or =1 microM). This antifibrotic effect was associated with a reduction of TGFbeta1 message.. Tenidap down-regulates the fibroproliferative changes typical of chronic arthritis, an effect that fits the profile of a disease-modifying agent for rheumatoid arthritis.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Arthritis, Experimental; Cell Division; Cells, Cultured; Chronic Disease; Collagen; Diclofenac; DNA Primers; Dose-Response Relationship, Drug; Down-Regulation; Extracellular Matrix; Fibronectins; Hyperplasia; Indoles; Oxindoles; Rabbits; RNA, Messenger; Synovial Membrane; Transforming Growth Factor beta

We investigated the effect of zinc sulfate on the proliferation of cultured bovine aortic smooth muscle cells stimulated with or without growth factors. Zinc had no effect on the incorporation of [3H]thymidine into the acid-insoluble fraction of the cells stimulated with or without platelet-derived growth factor or transforming growth factor beta 1. However, it was shown that stimulation of the [3H]thymidine incorporation by either basic or acidic fibroblast growth factor was significantly potentiated by zinc. Other cations including copper, manganese and nickel did not exhibit such an activity. The present data suggest that zinc is a particular heavy metal which potentiates vascular smooth muscle cell proliferation stimulated by basic and acidic fibroblast growth factors as well as thrombospondin. Zinc may be involved in the intimal hyperplasia of atherosclerosis.

Topics: Animals; Cattle; Cell Division; Cells, Cultured; Fibroblast Growth Factor 1; Fibroblast Growth Factor 2; Hyperplasia; Muscle, Smooth, Vascular; Platelet-Derived Growth Factor; Transforming Growth Factor beta; Zinc

Recently, cases of liver damage and liver tumors have been reported after treatment of prostate cancer patients with the antiandrogen cyproterone acetate (CPA). In rat liver, CPA initiates a wave of DNA synthesis that is accompanied by apoptosis. In apoptotic hepatocytes, a latent form of transforming growth factor beta 1 (TGF-beta 1) is detectable by immunohistochemistry. Injection of a single dose of TGF-beta 1 induces apoptosis in the liver of animals pretreated with CPA but has an insignificant effect in untreated animals. In this study, we show by Northern analysis that there is increased expression of TGF-beta 1 in the liver after CPA treatment. Detection of TGF-beta 1 with in situ hybridization showed that TGF-beta 1 was synthesized in the parenchymal cells. Time course and dose-response experiments performed 48 hours after the last application of CPA showed that apoptotic nuclei with chromatin condensed at the nuclear periphery (AN) were already visible 2 hours after injection (0.13%), and apoptotic bodies (ABs) increased 2 to 9 hours after the injection (from 1.28% to 6.67%) after 25 micrograms TGF-beta 1/kg. At 4.5 hours after injection, an induction of apoptosis could be detected with 0.25 microgram TGF-beta 1/kg and after the maximum dose (250 micrograms TGF-beta 1/kg) ANs (0.24%) and ABs (16.74%) were homogeneously distributed throughout the liver lobe. Irrespective of the dose or time after injection of TGF-beta 1, 82% of the ABs were localized within hepatocytes. Liver enzymes were detected in high amounts in the serum (eightfold elevation of glutamate dehydrogenase, fivefold elevation of alanine transaminase [ALT]) 7 hours after the first visible sign of apoptosis. After an additional 20 hours, the liver contained many necrotic figures. These results suggest that the combination of TGF-beta 1 expression coupled with a strikingly enhanced sensitivity to the induction of apoptosis could be responsible both for the liver damage and the development of liver tumors observed after treatment with CPA.

Topics: Androgen Antagonists; Animals; Apoptosis; Body Weight; Cyproterone Acetate; Dose-Response Relationship, Drug; Enzymes; Female; Hyperplasia; Liver; Necrosis; Phagocytosis; Rats; Rats, Wistar; Time Factors; Transforming Growth Factor beta

Forty-three 8-week-old male Wistar rats were studied to evaluate temporal changes of transforming growth factor beta1, (TGF-beta1) mRNA levels in thyroid tissue during pharmacologically induced goiter. Four rats were treated with purified bovine thyrotropin (TSH; Ambinon, 2 mU/day sc) for 7 days before being sacrificed. Thirty-one were treated with propylthiouracil (PTU), added to their drinking water at a concentration of 0.2 g%, and subsequently were sacrificed as follows: five after 1 week (PTU-1): five after 2 weeks (PTU-2); five after 4 weeks (PTU-4); five after 8 weeks (PTU-8); five after 12 weeks (PTU-12). In six rats, after 12 weeks of treatment. PTU was withdrawn for 2 months and subsequently started again in three rats which were sacrificed after 2 weeks (PTU-R); the remaining three rats were sacrificed without any further treatment (PTU-R control). Eight rats (control rats) were never treated and served as controls. After sacrifice, blood was drawn for determination of total thyroxine and the thyroid was excised and subdivided into two lobes. Northern analysis for TGF-beta1 was performed in one lobe. while histological and immunohistochemical studies were performed in the other lobe. Gene expression of TGF-beta1 was induced in TSH- and PTU-treated rats. In TSH-treated rats TGF-beta1 gene expression was less detectable than in PTU-treated rats, where it became evident after 2 weeks and remained through weeks 4-8. Gene expression of TGF-beta1 wits also seen in PTU-R rats, but not in the control and in the PTU-R control. Immunohistochemical analysis showed a different presence and location for the TGF-beta1 protein, which appears to be dependent on the time of exposure to mitogenic stimulus. In conclusion, TGF-beta1 is produced in response to both a direct (TSH by itself) and indirect (TSH induced by PTU-induced hypothyroidism) cellular proliferative stimulus and is not linked to an adaptative phenomenon secondary to hypothyroidism. The immunohistochemical location of TGF-beta1 within the thyrocytes is influenced by mitogen exposure time. A TGF-beta1 immunohistochemical evaluation may be important to define exposure time and activity of goitrogenic stimuli.

Topics: Animals; Blotting, Northern; Cattle; Goiter; Hyperplasia; Male; Propylthiouracil; Rats; Rats, Wistar; RNA, Messenger; Thyroid Gland; Thyrotropin; Time Factors; Transforming Growth Factor beta

Administration of a goitrogen (methimazole) and a low iodine diet to rats over a two-week period resulted in hypothyroidism and thyroid hyperplasia compared with controls (control: total serum thyroxine (T4) 66 +/- 4 nmol/l, thyroid weight 5 +/- 1 mg/100 g body weight; experimental: T4 undetectable, thyroid weight 27 +/- 4 mg/100 g body weight after 2 weeks of treatment; mean +/- S.D., n = 10). Immunohistochemistry carried out using a specific endothelial cell marker, CD31, and morphometric analysis (point counting of immunopositive cells) revealed that the progression of goitre in the rat thyroid is accompanied by an increase in capillary endothelial cell growth (neovascularisation). Fibroblast growth factor-2 (FGF-2) immunohistochemistry revealed widespread staining for the protein in the follicular cells of control glands. Less intense staining was found in the stroma and follicular cell nuclei. During hyperplasia and subsequent neovascularisation there was a progressive increase in the FGF-2 immunoreactivity at all locations during the two-week treatment period. Thrombospondin-1 (TSP1) immunoreactivity in the control rat thyroid was found in the stroma and in the endothelial cells, while weak follicular cell staining was also present. In the goitrous rat thyroid the TSP immunoreactivity was present after 1 week of treatment in the endothelial cells and most follicular cells, whilst stroma localisation was weak. After week 2 of treatment the endothelial cell and stromal localisation was no longer apparent, although a follicular localisation was still present. Transforming growth factor-beta 1 (TGF beta 1) immunoreactivity was present in the cytoplasm of a minority of the follicular cells in control rat thyroids, while their nuclei were unstained. In the goitrous rat thyroid an increase intensity of staining for TGF beta 1 was seen in all follicular cells, many of which now also demonstrated immuno-positive nuclei, within one week of goitrogen administration. These results show that in the hyperplastic thyroid increases in FGF-2 and TGF beta 1, and decreases in TSP1 accompany angiogenesis. These factors may interact in an autocrine/paracrine relationship to stimulate the neovascularisation that occurs during goitre formation.

Topics: Animals; Cell Adhesion Molecules; Cytoplasm; Endothelium; Fibroblast Growth Factor 2; Goiter; Growth Substances; Hyperplasia; Immunohistochemistry; Male; Membrane Glycoproteins; Methimazole; Neovascularization, Pathologic; Rats; Rats, Inbred Strains; Thrombospondins; Thyroid Gland; Transforming Growth Factor beta

Transforming growth factor beta-1 (TGF beta 1) is known to inhibit the growth of many epithelial cell types in culture. Consequently, it is important to determine whether it has any tumor suppressor activity in vitro. Fifteen heterozygous and eight wild type TGF beta 1-deficient mice were examined to determine if there was a difference in lifespan or lesion development due to the loss of one TGF beta 1 allele. Mice were killed when there was evidence of neoplasia or severe illness. There was no significant difference in the lifespan of the two groups. Hyperplastic lesions in the glandular mucosa were seen in 10 TGF beta 1 (+/-) mice. These lesions were localized to the lesser curvature of the stomach, extending from the limiting ridge to the pylorus. Seven of the 10 glandular hyperplastic lesions in the TGF beta 1 (+/-) mice had features similar to human gastritis cystica profunda. Associated with the glandular invasion of the muscularis were a mixed inflammatory infiltration of the surrounding muscular wall and mucosa with chronic vasculitis in the tissues adjacent to these lesions. In contrast to the distinct genotypic differences in lesion incidence observed in the glandular stomach, there was no significant difference in lesion incidence in other organs. The increased incidence of the hyperplastic lesions in the TGF beta 1 (+/-) mice is highly suggestive that allelic loss of TGF beta 1 plays an important role in the genesis of these lesions. However, allelic loss of TGF beta 1 does not cause alterations in the incidence of neoplasia.

Topics: Alleles; Animals; Gastric Mucosa; Gastritis; Genotype; Heterozygote; Hybridization, Genetic; Hyperplasia; Mice; Mice, Inbred Strains; Stomach; Transforming Growth Factor beta

Tenascin is an extracellular matrix glycoprotein which plays a role in cell attachment, proliferation and migration. To elucidate the function of tenascin in the proliferation of endometrial carcinoma, we studied tenascin expression in the endometrial carcinoma of 36 cases. In 22 of the carcinomas, tenascin expression was intense in the entire extracellular space, especially at the front of muscle invasion. Furthermore, in cases with metastases, deep invasion into muscles and vascular invasion, the rate of tenascin expression was significantly high. Immunoelectron microscopy revealed the tenascin reaction product in the stroma around fibroblasts located some distance from the basal lamina of cancer cells. On the other hand, tenascin expression was found in a high proportion of cases showing weak or no expression of estrogen receptor, and intense expression of transforming growth factor-beta 1. These results suggest that tenascin not only promotes cell proliferation and invasion but also inhibits further proliferation of carcinoma.

Topics: Adenocarcinoma; Endometrial Neoplasms; Endometrium; Extracellular Space; Female; Humans; Hyperplasia; Immunohistochemistry; Microscopy, Immunoelectron; Receptors, Estrogen; Tenascin; Transforming Growth Factor beta

The expression of transforming growth factor beta 1 and beta 2 (TGF-beta 1 and beta 2) in aortic and venous tissue from male New Zealand White rabbits, at selected time intervals after birth, was examined by the reverse transcriptase polymerase chain reaction. Stable levels of TGF-beta 1 were found in all segments derived from the aortic arch and descending aorta at each time interval. However, increasing amounts of TGF-beta 2 transcripts were observed for the aortic arch from day 4, with peaks occurring between 1 and 6 months of age, followed by progressively decreasing levels thereafter. TGF-beta 2 transcripts in the descending aorta generally did not change significantly over time. TGF-beta transcripts manifested a significantly lower expression in the vena cava than in aortic segments. Histological analysis of the vascular tissue showed cellular hyperplasia (2.5-fold greater prevalence of nuclei per field) in the aortic arch media at 1 month of age as compared with nuclei per field at 12 months and increasing thickness of the aortic arch media with time. No significant differences in relative collagen concentrations were observed among the aortic and vena cava segments. These results suggest that these TGF-beta isoforms may participate in the physiological induction and differentiation of arterial and venous tissue during early normal vascular maturation.

Topics: Actins; Animals; Aorta, Thoracic; Blotting, Northern; Endothelium, Vascular; Hyperplasia; Male; Oligonucleotide Probes; Polymerase Chain Reaction; Rabbits; RNA; Transcription, Genetic; Transforming Growth Factor beta; Venae Cavae

Intimal hyperplasia is a common cause of obstructive stenosis after arterial reconstructive procedures. It has been postulated that growth factors elaborated by vascular wall cells regulate fibroproliferative changes that can cause graft failure. This study characterizes transforming growth factor beta-1 (TGF-beta 1) and platelet-derived growth factor-A chain (PDGF-A) mRNA transcript profiles and their temporal relationship to the development of intimal hyperplasia in vein grafts.. Epigastric vein-to-common femoral artery interposition grafts were performed in male Lewis rats (350 to 450 gm) with standard microsurgical techniques. Grafts were harvested at 1 and 4 hours, 1 and 4 days, and 1 and 2 weeks (n = 5/time). Graft RNA was extracted, reverse-transcribed, and amplified by polymerase chain reaction with sense/antisense primers for TGF-beta 1 and PDGF-A (30 cycles). Polymerase chain reaction fragments were confirmed by Southern hybridization.. Variable induction of TFG-beta 1 gene transcription was evident in vein grafts at 1 and 4 hours, with prominent mRNA expression from 1 day to 2 weeks. PDGF-A mRNA was detected in ungrafted control veins but was downregulated at 1 hour and absent at 4 hours after grafting. PDGF-A transcription was upregulated by 1 day, with prominent expression from 4 days to 1 week. Early loss of PDGF-A mRNA correlated with the early denudation of the endothelium, whereas upregulation by 4 days was preceded by TGF-beta 1 mRNA expression.. Upregulation of TGF-beta 1 and PDGF-A mRNA expression is detected in vein grafts before the development of a quantifiable neointima, which occurs by 2 weeks in our model. This suggests a role for these growth factors in the development of vein graft intimal hyperplasia.

Topics: Animals; Femoral Artery; Gene Expression; Hyperplasia; Male; Nucleic Acid Hybridization; Platelet-Derived Growth Factor; Polymerase Chain Reaction; Rats; Rats, Inbred Lew; RNA, Messenger; Time Factors; Transforming Growth Factor beta; Tunica Intima; Up-Regulation; Veins

Dermal effects of KH 1060, a novel, highly potent 20-epi analogue of 1 alpha,25-dihyroxyvitamin D3, were investigated in a hairless mouse model. During daily topical applications of a 0.4 microM solution of KH 1060 for 4 weeks, epidermal hyperplasia and an increase of dermal thickness and mass were observed. KH 1060 upregulated glycosaminoglycan and collagen synthesis in the skin, and increased glycosaminoglycan deposition in the subepidermal region. Reverse transcription-polymerase chain reaction amplification of the transforming growth factor (TGF) beta 1-specific mRNA revealed that KH 1060 stimulated expression of this growth factor in the epidermis, but not in the dermis. Changes observed after application of 1 alpha,25-dihydroxyvitamin D3 were much less pronounced but qualitatively similar to the effects of KH 1060, whereas structurally related but receptor inactive compounds, vitamin D3 and 1 beta,25-dihydroxyvitamin D3, did not produce any effects. Furthermore, we were unable to demonstrate the involvement of the non-genomic, receptor-independent vitamin D signalling in the skin, using a specific stimulator (Ro 24-2090) and a blocker (1 beta,25-dihydroxyvitamin D3) of this pathway. Our findings provide the first evidence that a strong vitamin D3 analogue triggers synthesis of skin connective tissue, possibly via vitamin D receptor activation and the paracrine action of epidermis-derived TGF-beta 1.

Topics: Animals; Calcitriol; Collagen; Epidermis; Female; Glycosaminoglycans; Hyperplasia; Immunosuppressive Agents; Mice; Mice, Hairless; Mice, Inbred C3H; Skin; Transforming Growth Factor beta

Transforming growth factor-beta 1 (TGF-beta 1) is a modulator of cellular proliferation, differentiation, and extracellular matrix deposition. It is a potent epithelial growth inhibitor and can alter the differentiative properties of keratinocytes, in vitro, but little is known about its normal physiological function in the epidermis in vivo. Transgenic mice were generated using a keratin 10 (K10) gene promoter to drive constitutive expression of TGF-beta 1 in the suprabasal keratinocyte compartment. Surprisingly, these mice showed a two- to threefold increase in epidermal DNA labeling index over control mice, in the absence of hyperplasia. The transgene, however, acted in the expected fashion, as a negative regulator of cell growth, when hyperplasia was induced by treatment by 12-tetradecanoyl-phorbol-13-acetate (TPA). Epidermal TGF-beta type I and II receptor (T beta RI and T beta RII) levels were examined in control and transgenic mice during induction of hyperplasia by TPA. Whereas T beta RI levels remained relatively constant, T beta RII expression was strongly induced in TPA-treated skins, prior to the induction of the growth inhibitory response to TGF-beta 1, and its level of expression correlated with growth sensitivity to TGF-beta 1 in vivo and in vitro. These results suggest that TGF-beta 1 and its type II receptor are part of the endogenous homeostatic regulatory machinery of the epidermis.

Topics: Activin Receptors, Type I; Animals; Base Sequence; Epidermal Cells; Epidermis; Female; Gene Targeting; Homeostasis; Hyperplasia; Keratinocytes; Male; Mice; Mice, Inbred C57BL; Mice, Inbred CBA; Mice, Transgenic; Mitotic Index; Molecular Sequence Data; Protein Serine-Threonine Kinases; Receptor, Transforming Growth Factor-beta Type I; Receptor, Transforming Growth Factor-beta Type II; Receptors, Transforming Growth Factor beta; Tetradecanoylphorbol Acetate; Transforming Growth Factor beta

Transforming growth factor beta 1 (TGF beta 1) is a multifunctional cytokine. The role of TGF beta 1 in hepatocarcinogenesis is still a matter of discussion. To assess the expression of TGF beta 1 in human liver tumors, the following study was conducted.. Formalin fixed, paraffin embedded sections of 50 hepatocellular carcinomas (HCC), 30 cholangiocellular carcinomas (CCC), 15 hepatocellular adenomas (HCA), 15 focal nodular hyperplasias (FNH), and 20 normal livers (NL) were immunostained with a monoclonal anti-TGF beta 1 antibody (clone TB-21). TGF beta 1 mRNA was measured in snap frozen tissue of 2 HCC, 3 HCA, 2 FNH, and 3 NL of the immunohistochemistry group using slot blot hybridizations. Total RNA was extracted, and slot blots were hybridized with 32P endlabeled TGF beta 1 oligonucleotides. TGF beta 1 mRNA was measured in cpm with a scintillation counter.. TGF beta 1 protein was strongly expressed in 14/15 FNH and 13/15 HCA, whereas it was scarcely detectable in 46/50 HCC and 25/30 CCC. All NL were moderately positive. The 2 HCC cases contained far less TGF beta 1 mRNA than the HCA, FNH, and NL examined here.. TGF beta 1 expression is markedly lower in malignant liver tumors than in benign tumors or NL. Therefore, TGF beta 1 down-regulation may be an important step in hepatocarcinogenesis. Additionally, TGF beta 1 immunostaining may be useful in distinguishing well-differentiated HCC from adenomas.

Topics: Adenoma; Carcinoma, Hepatocellular; Cholangiocarcinoma; Humans; Hyperplasia; Immunohistochemistry; In Situ Hybridization; Liver; Liver Neoplasms; Precancerous Conditions; RNA, Messenger; Transforming Growth Factor beta

Hyperplasia of transitional cell epithelium adjacent to human transitional cell carcinomas (TCC) is a common finding in pathology. This hyperplasia may be a precancerous aberration. Alternatively, it has been suggested that the hyperplasia is due to paracrine action of tumour-derived growth factors. In this study we tested the latter hypothesis using the mouse tumorigenic TCC cell line NUC-1. Transplantation of NUC-1 tumour cells into the urinary bladder submucosa of syngeneic mice in vivo induced hyperplasia of normal adjacent urothelium in all tested mice. Implantation of normal mouse bladder mucosa did not induce urothelial hyperplasia. In vitro, conditioned medium of NUC-1 cells induced the proliferation of the mouse urothelial cell line g/G, which closely resembles normal urothelial cells. This induction was inhibited by transforming growth factor beta 1 (TGF beta 1). Similarly, TGF beta 1 inhibited the fibroblast growth factor-1 (FGF-1) and FGF-2 induced proliferation of g/G cells. Chemico-physical examination, bioassays with conditioned media, and RNA analysis of NUC-1 cells revealed that these cells secreted a growth factor with FGF-like properties. These results indicate that epithelial hyperplasia surrounding carcinomas is not necessarily a precancerous aberration, but may result from direct paracrine action of tumour-derived growth factors.

Topics: Animals; Carcinoma, Transitional Cell; Cell Division; Cell Line; Culture Media, Conditioned; Dithiothreitol; DNA Replication; Epidermal Growth Factor; Epithelium; Fibroblast Growth Factor 1; Fibroblast Growth Factor 2; Granulocyte-Macrophage Colony-Stimulating Factor; Growth Substances; Humans; Hyperplasia; Mice; Mucous Membrane; Neoplasm Transplantation; Thymidine; Transforming Growth Factor alpha; Transforming Growth Factor beta; Transplantation, Isogeneic; Tumor Cells, Cultured; Urinary Bladder; Urinary Bladder Neoplasms

Intimal hyperplasia is induced by therapeutic vascular interventions and often results in clinically important narrowing of the vascular lumen. Examination of the role of TGF-beta 1 in a rat carotid artery injury model confirmed the presence of a previously reported increase in TGF-beta 1 mRNA in the media of injured arteries. Administration of neutralizing anti- TGF-beta 1 antibodies significantly (P < 0.05) reduced the size of the intimal lesions that developed after carotid balloon injury. A control antibody had no effect. The intimal/medial area ratio was also reduced in the anti-TGF-beta 1 group relative to controls (P < 0.01). Immunohistochemical staining showed that two TGF-beta 1-induced extracellular matrix components, EDA + fibronectin and versican, were greatly increased in the untreated neointimal lesions, but were almost completely absent from the lesions of the anti-TGF-beta 1-treated animals. We conclude that TGF-beta 1 is causally involved in the development of intimal hyperplasia, and that anti-TGF-beta 1 agents may be useful in achieving at least partial control of this condition.

Topics: Animals; Antibodies; Base Sequence; Cell Division; Extracellular Matrix; Hyperplasia; Male; Molecular Sequence Data; Muscle, Smooth, Vascular; Rats; Rats, Sprague-Dawley; Transforming Growth Factor beta

Partial obstruction of the rabbit urethra induces rapid bladder growth. This growth is characterized by hypertrophy of smooth muscle cells in addition to hyperplasia of cells in the urothelium and serosa. The local synthesis of growth factors has been proposed to be influential in this growth since partial outlet obstruction rapidly increases the bladder's expression of basic fibroblast growth factor, while suppressing the expression of transforming growth factor-beta. Upon release of the outlet obstruction, the hypertrophied bladder regresses to its normal weight. Here, we examined whether regression of the hypertrophied rabbit bladder involves apoptosis (programmed cell death) of specific cellular elements and whether the expression of growth factors is altered concomitant with apoptotic cell deletion.. Regressing rabbit bladders were analyzed for markers of apoptosis, including DNA fragmentation and histology. An in situ enzymatic immuno-histochemical procedure was utilized to localize apoptotic cells in these tissues. Finally, Northern blot analysis was used to identify changes in the expression of basic fibroblast growth factor and transforming growth factor-beta during bladder regression.. Regressing rabbit bladders demonstrated the characteristic electrophoretic "ladder" pattern of DNA fragmentation associated with apoptosis. An in situ technique to distinguish cells with degraded nuclear DNA identified apoptosis only within the urothelium and serosal lamina of the regressing bladders. RNAs extracted from regressing bladders exhibited decreased expression of basic fibroblast growth factor mRNA as well as increased expression of transforming growth factor-beta 1 mRNA when compared with RNAs from hypertrophied bladders.. We conclude that hyperplasia and apoptosis are opposing cellular processes that mediate the bladder's response to short-term obstructive stimuli and that local synthesis of growth-promoting and growth-inhibitory factors may be responsible for initiating both of these responses.

Topics: Animals; Apoptosis; DNA Damage; Fibroblast Growth Factor 2; Gene Expression; Hyperplasia; Male; Muscle, Smooth; Rabbits; RNA, Messenger; Time Factors; Transforming Growth Factor beta; Urinary Bladder Neck Obstruction

These studies investigate the role of transforming growth factor-beta 1, a potent inhibitor of epithelial cell proliferation and stimulator of extracellular matrix biosynthesis, during intrahepatic bile duct hyperplasia and biliary fibrosis. These pathogenic responses were induced in rats by common bile duct ligation. Bile duct cell replication, measured by the bromodeoxyuridine labeling index, was significantly increased 24 hr after common bile duct ligation. This response diminished to baseline by 1 wk. Liver collagen content, determined by quantification of hydroxyproline, was increased significantly after 1 wk of common bile duct ligation, and by 4 wk was increased by a factor of 4. Immunohistochemistry revealed low levels of TGF-beta 1 in normal intrahepatic bile duct epithelium. In contrast, the bile duct epithelium in bile duct-ligated rats stained strongly positive for transforming growth factor-beta 1 at 1 and 4 wk after ligation. These results suggest that transforming growth factor-beta 1 may play a role in both the termination of the bile duct epithelial cell proliferative response and the induction of fibrogenesis after common bile duct ligation. In addition, the mannose 6-phosphate/insulin-like growth factor II receptor was up-regulated in hyperplastic bile duct epithelium 1 and 4 wk after ligation. Because the mannose 6-phosphate/insulin-like growth factor-II receptor has been shown to facilitate the proteolytic activation of transforming growth factor-beta 1, these results suggest that the bile duct epithelium may also be involved in the activation of transforming growth factor-beta 1.

Topics: Animals; Bile Duct Diseases; Bile Ducts, Intrahepatic; Common Bile Duct; DNA; Fibrosis; Hydroxyproline; Hyperplasia; Immunohistochemistry; Ligation; Liver; Male; Rats; Rats, Sprague-Dawley; Receptor, IGF Type 2; Transforming Growth Factor beta

Acute treatment of mouse skin with the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) induces marked epidermal hyperplasia, which is well evident by 24 h and maximal by 48-72 h. These effects are associated with the early induction of transforming growth factor (TGF) beta 1 expression in the epidermis. We show here that, in contrast to TGF-beta 1, TGF-beta 2, and TGF-beta 3, skin expression is significantly down-modulated in response to TPA. TGF-beta 3 RNA levels decreased by 6 h of treatment but returned to normal or even higher levels at later times. The TGF-beta 3 protein could be detected immunohistochemically in both dermis and epidermis in control skins and at early times of TPA treatment. However, at later times, TGF-beta 3 was found only in dermal cells and not in the epidermis. TGF-beta 2 RNA expression was found to be significantly down-modulated by 24 h of TPA treatment and remained low even at later times. Thus, differential control of the 3 TGF-beta isoforms appears to be a likely determinant of normal skin homeostasis and could be at least partially responsible for TPA-induced skin hyperplasia.

Topics: Animals; Down-Regulation; Female; Hyperplasia; Mice; RNA; Skin; Tetradecanoylphorbol Acetate; Transforming Growth Factor beta

Tight skin (TSK/+) mice develop a cutaneous hyperplasia associated with the occurrence of autoantibodies characteristic for scleroderma. In order to study the role of autoimmunity in the production of skin fibrosis, we conducted adoptive transfer experiments in which bone marrow cells of TSK/pa mice were infused into pa/pa mice littermates. (C57BL/6 pa/pa mice are used to produce heterozygous TSK/pa mice). Our results showed that after a prodromal period of several months, the transfer of bone marrow cells led to skin fibrosis, the presence of autoantibodies, and increased transcription of (alpha 1) collagen I and TGF beta genes. Infusion of enriched B or T cells alone did not cause skin fibrosis but of B cells alone increased autoantibody production. By contrast, transfer of T and B lymphocytes led to earlier mild fibrosis, cellular infiltration and autoantibody production as well as increased transcription of the (alpha 1) collagen gene. Our results strongly demonstrate, for the first time, that immunocompetent cells can play a role in the activation of collagen synthesis leading to skin fibrosis.

Topics: Animals; Autoantibodies; Autoimmune Diseases; Bone Marrow Transplantation; Collagen; Connective Tissue Diseases; Disease Models, Animal; Fibrosis; Gene Expression Regulation; Hyperplasia; Immunocompetence; Immunoglobulin G; Immunoglobulin M; Immunotherapy, Adoptive; Lymphocyte Cooperation; Lymphocyte Subsets; Mice; Mice, Inbred C57BL; Mice, Mutant Strains; Scleroderma, Systemic; Skin; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

The arterial wall responds to thrombosis or mechanical injury through the induction of specific gene products that increase cellular proliferation and connective tissue formation. These changes result in intimal hyperplasia that is observed in restenosis and the early phases of atherosclerosis. Transforming growth factor beta 1 (TGF-beta 1) is a secreted multi-functional protein that plays an important role in embryonal development and in repair following tissue injury. However, the function of TGF-beta 1 in vascular cell growth in vivo has not been defined. In this report, we have evaluated the role of TGF-beta 1 in the pathophysiology of intimal and medial hyperplasia by gene transfer of an expression plasmid encoding active TGF-beta 1 into porcine arteries. Expression of TGF-beta 1 in normal arteries resulted in substantial extracellular matrix production accompanied by intimal and medial hyperplasia. Increased procollagen, collagen, and proteoglycan synthesis in the neointima was demonstrated by immunohistochemistry relative to control transfected arteries. Expression of TGF-beta 1 induced a distinctly different program of gene expression and biologic response from the platelet-derived growth factor B (PDGF B) gene: procollagen synthesis induced by TGF-beta 1 was greater, and cellular proliferation was less prominent. These findings show that TGF-beta 1 differentially modulates extracellular matrix production and cellular proliferation in the arterial wall in vivo and could play a reparative role in the response to arterial injury.

Topics: Animals; Arteries; Arteriosclerosis; Base Sequence; Collagen; DNA Primers; Extracellular Matrix; Extracellular Matrix Proteins; Gene Expression; Gene Transfer Techniques; Hyperplasia; Molecular Sequence Data; Platelet-Derived Growth Factor; Polymerase Chain Reaction; Procollagen; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-sis; RNA, Messenger; Swine; Transforming Growth Factor beta; Tunica Intima

Increasing evidence suggests that angiotensin II (Ang II) may act as a growth factor for the heart. However, direct effects of Ang II on mammalian cardiac cells (myocytes and nonmyocytes), independent of secondary hemodynamic and neurohumoral effects, have not been well characterized. Therefore, we analyzed the molecular phenotype of cultured cardiac cells from neonatal rats in response to Ang II. In addition, we examined the effects of selective Ang II receptor subtype antagonists in mediating the biological effects of Ang II. In myocyte culture, Ang II caused an increase in protein synthesis without changing the rate of DNA synthesis. In contrast, Ang II induced increases in protein synthesis, DNA synthesis, and cell number in nonmyocyte cultures (mostly cardiac fibroblasts). The Ang II-induced hypertrophic response of myocytes and mitogenic response of fibroblasts were mediated primarily by the AT1 receptor. Ang II caused a rapid induction of many immediate-early genes (c-fos, c-jun, jun B, Egr-1, and c-myc) in myocyte and nonmyocyte cultures. Ang II induced "late" markers for cardiac hypertrophy, skeletal alpha-actin and atrial natriuretic factor expression, within 6 hours in myocytes. Ang II also caused upregulation of the angiotensinogen gene and transforming growth factor-beta 1 gene within 6 hours. Induction of immediate-early genes, late genes, and growth factor genes by Ang II was fully blocked by an AT1 receptor antagonist but not by an AT2 receptor antagonist. These results indicate that: (1) Ang II causes hypertrophy of cardiac myocytes and mitogenesis of cardiac fibroblasts, (2) the phenotypic changes of cardiac cells in response to Ang II in vitro closely mimic those of growth factor response in vitro and of load-induced hypertrophy in vivo, (3) all biological effects of Ang II examined here are mediated primarily by the AT1 receptor subtype, and (4) Ang II may initiate a positive-feedback regulation of cardiac hypertrophic response by inducing the angiotensinogen gene and transforming growth factor-beta 1 gene.

Topics: Angiotensin II; Angiotensinogen; Animals; Cardiomegaly; Cells, Cultured; Fibroblasts; Gene Expression Regulation; Genes, fos; Hyperplasia; Myocardium; Rats; Receptors, Angiotensin; Transforming Growth Factor beta

Previous studies have indicated that aged animals show an increased intimal hyperplasia after arterial injury. The present studies examined the hypothesis that the increased serum-free proliferation of aged smooth muscle cells (SMC), in vitro, was due to a loss of an antiproliferative signal, such as transforming growth factor-beta 1 (TGF-beta 1). Northern blot analysis of the mRNA derived from old (> 19 mo) or young (3-4 mo) rat aortic SMC indicated that both groups had an equivalent level of the 2.5 kB TGF-beta 1 message. Metabolic labeling with 35S-methionine and immunoprecipitation for TGF-beta 1 confirmed the de novo synthesis of TGF-beta 1 in rat SMC. Old and young SMC supernatants showed equal levels of active or latent (acid-activated) TGF-beta activity. Despite the similarities in the production of TGF-beta 1, old SMC were refractory to inhibition by TGF-beta 1, whereas young SMC were markedly inhibited (80%) by low levels of TGF-beta 1 (IC50 < 5 pg/ml). Binding studies at 4 degrees C indicated that old SMC exhibited reduced binding capacity for 125I-TGF-beta 1. Cross-linking studies confirmed that old SMC showed reduced binding of 125I-TGF-beta 1 to membrane sites corresponding to the high molecular weight type III receptor, as well as the 85-kDa type II and 65-kDa type I receptor. However, at 37 degrees C, old SMC degraded 125I-TGF-beta 1 more rapidly than young SMC. Combined, this data suggests that SMC derived from older animals are capable of normal production of TGF-beta 1 but fail to respond to the autocrine growth inhibitory effects of this agent, thereby leading to enhanced proliferation.

Topics: Aging; Animals; Biological Transport, Active; Cell Division; Hyperplasia; In Vitro Techniques; Male; Muscle, Smooth, Vascular; Proteoglycans; Rats; Rats, Inbred F344; Receptors, Cell Surface; Receptors, Transforming Growth Factor beta; Transforming Growth Factor beta

Immunohistochemical localization of transforming growth factor-beta 1 (TGF-beta 1) was studied in the lungs of rats given crystalline silica or ferric oxide by single intratracheal instillation. Ferric oxide elicited no progressive granulomatous reaction, no epithelial hyperplasia, and no lung tumors; no demonstrable reactivity to TGF-beta 1 was observed. Silica induced a granulomatous reaction with progressive fibrosis, adjacent alveolar type II hyperplasia, and alveolar carcinomas. Rabbit polyclonal antibodies to synthetic peptides corresponding to the first 30 amino acids of mature TGF-beta 1, anti-LC (1-30), and anti-CC (1-30) were used for the localization of intracellular and extracellular TGF-beta 1. An antibody to a peptide corresponding to amino acids 266-278 of the TGF-beta 1 precursor sequence, anti-Pre (266-278), was used to detect the TGF-beta precursor and the latency-associated peptide. Intracellular mature TGF-beta (anti-LC) was demonstrated in fibroblasts and macrophages located at the periphery of silicotic granulomas and in fibroblasts adjacent to hyperplastic type II cells. Extracellular mature TGF-beta 1 was localized in the connective tissue matrix of the granulomas and in the stroma of both hyperplastic type II cells and well-differentiated adenocarcinomas. Immunoreactivity to anti-Pre was localized, intracellularly, in hyperplastic alveolar type II cells and their proliferative lesions adjacent to granulomas, in adenomas, but not in adenocarcinomas. The hyperplastic type II cells appear to be the sites of production and secretion of TGF-beta 1, which may regulate their own growth and differentiation and mediate the production of extracellular TGF-beta 1-associated matrix. The lack of reactivity to TGF-beta 1 precursor in the adenocarcinomas is consistent with the loss of normal cellular differentiation and function. TGF-beta 1 appears to have a pathogenetic role in silica-induced mesenchymal and epithelial lesions. The role of TGF-beta 1 and other cytokines in silica-induced carcinogenesis requires further investigation.

Topics: Adenocarcinoma; Adenoma; Animals; Disease Models, Animal; Female; Ferric Compounds; Hyperplasia; Lung; Lung Neoplasms; Male; Protein Precursors; Pulmonary Alveoli; Rats; Rats, Inbred F344; Silicon Dioxide; Silicosis; Transforming Growth Factor beta

Cell death by apoptosis is a major determinant of growth of normal tissues and tumours. The present study aimed to elucidate signal factors involved in its regulation. Epithelial cells in control liver, during regression of cyproterone acetate induced liver hyperplasia, in liver (pre)neoplasia and in uterus undergoing apoptosis in vivo show immunostaining for transforming growth factor beta 1 (TGF-beta 1) as detected by anti-pre(266-278) TGF-beta 1 antibodies. Positive immunostaining is also seen in a few intact cells of hyperplastic, regressing liver apparently preparing for apoptosis, but is virtually not found in hepatocytes of normal or growing liver nor in cells undergoing death by necrosis. Recombinant latency associated protein (rLAP, dimer of the pro-region non-covalently associated with the mature region) complex and mature TGF-beta 1 induce apoptosis in isolated hepatocytes cultured in vitro. These findings suggest an involvement of TGF-beta 1 in the induction of apoptosis in certain epithelia in vivo.

Topics: Animals; Apoptosis; Biomarkers; Cell Death; Cyproterone Acetate; Female; Hyperplasia; Liver; Necrosis; Rats; Transforming Growth Factor beta

We have shown that angiotensin II (Ang II)-induced hypertrophy of vascular smooth muscle cells is dependent on the balance between proliferative and antiproliferative growth factors, specifically basic fibroblast growth factor and transforming growth factor-beta 1 (TGF-beta 1), respectively. We now present evidence, based on two phenotypically distinct cell cultures, that the ability to secrete the biologically active form of TGF-beta 1 is central to the growth response to Ang II. Two separate cultures were examined, one in which Ang II induces hypertrophy and the other in which Ang II induces hyperplasia. Ang II induces the expression of basic fibroblast growth factor twofold to fivefold in both cultures. Furthermore, both cultures express TGF-beta 1. In the culture that responds with hypertrophy, Ang II induces the expression of the active form of TGF-beta 1 twofold to threefold. However, in the culture that responds with hyperplasia, no active TGF-beta 1 was detected either at baseline or after Ang II exposure. Interestingly, all the TGF-beta 1 present was in the inactive, latent form. In the culture that responded with hyperplasia, Ang II induced a fourfold to fivefold increase in DNA synthesis. This increase could be abolished by the addition of active TGF-beta 1. Thus in these two cultures the ability to activate TGF-beta 1 dictates the cellular response to Ang II. These results support our hypothesis that a balance of proliferative and antiproliferative autocrine signals mediates the growth control of vascular smooth muscle cells.

Topics: Angiotensin II; Animals; Biological Assay; Cell Division; Cells, Cultured; DNA; Fibroblast Growth Factor 2; Hyperplasia; Hypertrophy; Muscle, Smooth, Vascular; Rats; Rats, Inbred WKY; Transforming Growth Factor beta

We previously reported that heparin inhibits the proliferation of fibroblasts and vascular smooth muscle cells (SMC), in part, by binding to and increasing the antiproliferative activity of transforming growth factor-beta 1 (TGF-beta 1). We now report that certain other polyanions which are structurally distinct from heparin, such as fucoidan and polyinosinic acid, are more avid ligands for TGF-beta 1 and more potent antiproliferative agents than heparin. Fucoidan possessed more potent antiproliferative activity than heparin against rat and bovine aortic SMC in vitro, though possessing much lower anticoagulant activity than heparin. Furthermore, fucoidan suppressed in vivo intimal hyperplasia when continuously infused into rats subjected to balloon-catheter injury. Unlike heparin, which also suppressed intimal hyperplasia, fucoidan did not cause systemic anticoagulation. Thus, fucoidan may be useful as a non-anticoagulant inhibitor of post-angioplasty intimal hyperplasia.

Topics: Animals; Anions; Aorta; Cattle; Cell Division; Cells, Cultured; Dose-Response Relationship, Drug; Heparin; Hyperplasia; Muscle, Smooth, Vascular; Polysaccharides; Rats; Rats, Inbred F344; Receptors, Cell Surface; Receptors, Transforming Growth Factor beta; Transforming Growth Factor beta

Transforming growth factor beta (TGF-beta) is a multifunctional homodimeric polypeptide with potent actions upon many target cells, including those of mesenchymal and haemopoietic lineage. The recent reports of high levels of the cytokine in rheumatoid synovium and synovial fluid, prompted this study into the effect of intra-articular injection of TGF beta-2 into rabbit knee-joints. Four daily injections of 1 microgram caused swelling, probably as a consequence of prostaglandin E2 production, synovial fibroblastic hyperplasia and a striking loss of femoral condyle proteoglycan. Using the polymerase chain reaction, no evidence could be obtained for the induction of interleukin-1 alpha gene expression in either synovial tissue or synovial fluid cells. These findings suggest that the TGF-beta present in the rheumatoid joint may contribute directly to the pathogenesis of rheumatoid arthritis.

Topics: Animals; Arthritis; Cartilage, Articular; Dinoprostone; Edema; Hyperplasia; Injections, Intra-Articular; Interleukin-1; Polymerase Chain Reaction; Proteoglycans; Rabbits; Recombinant Proteins; Synovial Membrane; Transforming Growth Factor beta

In previous studies hepatocytes undergoing cell death by apoptosis but not normal hepatocytes in rat liver showed immunostaining for transforming growth factor beta 1 (TGF-beta 1). Staining was much stronger with antibodies recognizing the pro-region of TGF-beta 1 than the mature peptide itself. Therefore we investigated the ability of both forms of TGF-beta 1 to induce apoptosis in primary cultures of rat hepatocytes. Mature TGF-beta 1 induced rounding up of the cells and fragmentation into multiple vesicles. As revealed by the DNA-specific stain H33258, the chromatin of these cells condensed and segregated into masses at the nuclear membrane; this was obviously followed by fragmentation of the nucleus. Ultrastructurally the cytoplasm was well preserved, as demonstrated by the presence of intact cell organelles. These features strongly suggest the occurrence of apoptosis. Quantification of nuclei with condensed chromatin revealed that mature TGF-beta 1 was 30-fold more effective than the TGF-beta 1 latency-associated protein complex. Finally, we administered TGF-beta 1 in vivo using an experimental model in which regression of rat liver was initiated by a short preceding treatment with the hepatomitogen cyproterone acetate. Two doses of TGF-beta 1, each 1 nM/kg, augmented the incidence of apoptotic hepatocytes 5-fold. Equimolar doses of TGF-beta 1 latency-associated protein complex were ineffective. These studies suggest that TGF-beta 1 is involved in the initiation of apoptosis in the liver and that the mature form of TGF-beta 1 is the active principle.

Topics: Animals; Cell Death; Cell Nucleus; Cells, Cultured; Dose-Response Relationship, Drug; Female; Humans; Hyperplasia; Kinetics; Liver; Microscopy, Electron; Rats; Rats, Inbred Strains; Recombinant Proteins; Time Factors; Transforming Growth Factor beta

Recent observations in our laboratory suggest that angiotensin II (Ang II) is a bifunctional vascular smooth muscle cell (VSMC) growth modulator capable of inducing hypertrophy or inhibiting mitogen-stimulated DNA synthesis. Because transforming growth factor-beta 1 (TGF beta 1) has similar bifunctional effects on VSMC growth, we hypothesized that autocrine production of TGF beta 1 may mediate the growth modulatory effects of Ang II. Indeed, this study demonstrates that Ang II induces a severalfold increase in TGF beta 1 mRNA levels within 4 h that is dependent on de novo protein synthesis and appears to be mediated by activation of protein kinase C (PKC). Ang II not only stimulates the synthesis of latent TGF beta 1, but also promotes its conversion to the biologically active form as measured by bioassay. The coincubation of VSMCs with Ang II and control IgG has no significant mitogenic effect. However, the co-administration of Ang II and the anti-TGF beta 1 antibody stimulates significantly DNA synthesis and cell proliferation. We conclude that: (a) Ang II induces increased TGF beta 1 gene expression via a PKC dependent pathway involving de novo protein synthesis; (b) Ang II promotes the conversion of latent TGF beta 1 to its biologically active form; (c) Ang II modulates VSMC growth by activating both proliferative and antiproliferative pathways; and (d) Autocrine active TGF beta 1 appears to be an important determinant of VSMC growth by hypertrophy or hyperplasia.

Topics: Angiotensin II; Animals; Cell Division; Cells, Cultured; DNA Replication; Gene Expression; Hyperplasia; Hypertrophy; In Vitro Techniques; Muscle, Smooth, Vascular; Rats; Rats, Inbred Strains; RNA, Messenger; Tetradecanoylphorbol Acetate; Transforming Growth Factor beta

The transforming growth factors type beta (TGF-beta) are immunoregulatory cytokines with pronounced effects on tissue homeostasis and repair. We observed that TGF-beta 1, TGF-beta 2, and the heterodimeric TGF-beta 1.2 induced astrocyte hyperplasia and strongly affected monolayer formation. After confluent growth, astrocytes are contact inhibited and form a dense monolayer. Addition of TGF-beta induced migration of cells to local centers, the formation of foci, followed by detachment of these aggregates from the cell culture surface. This effect was unique and could neither be induced by BMP2, a member of the decapentaplegic subfamily of type beta transforming growth factors nor by other cytokines or interleukins. Thus TGF-beta secreted by activated leukocytes might be a local regulator of astrocyte function during regenerative processes in inflammatory demyelinating brain disease.

Topics: Animals; Astrocytes; Bone Morphogenetic Proteins; Cell Adhesion; Cell Aggregation; Cell Division; Cell Line; Cell Movement; Cell Size; Hyperplasia; Multigene Family; Proteins; Rats; Recombinant Proteins; Transforming Growth Factor beta

Transforming growth factor-beta (TGF-beta) modulates the growth and differentiation of many cells and often functions in an autocrine or paracrine fashion. The myoepithelial cells of the renal juxtaglomerular apparatus (JGA) synthesize and secrete renin. Under conditions which chronically stimulate renin production, the JGA undergoes hypertrophy and hyperplasia. The molecular factors responsible for these changes in the JGA have not been identified. In the present study, plasma renin activity was stimulated in the mouse by water deprivation. Using immunoperoxidase staining with specific antibodies against TGF-beta 1, beta 2, and beta 3, we found increased TGF-beta 2 accumulation in the JGA and interlobular arteries. Immunostaining with renin antiserum demonstrated colocalization of TGF-beta 2 and renin. TGF-beta 1 and beta 3 expression was not different between control and water-deprived mice. Our results suggest that in the setting of water deprivation, TGF-beta 2 is localized in a manner which would allow it to act either as a growth factor for or as a phenotypic modulator of the JGA and renal arterioles.

Topics: Angiotensin II; Animals; Dehydration; Female; Hyperplasia; Juxtaglomerular Apparatus; Mice; Renin; Transforming Growth Factor beta

We have studied the consequences of introducing human recombinant transforming growth factor beta 1 (hrTGF-beta 1) into synovial tissue of the rat, to begin to better understand the significance of the fact that biologically active TGF-beta is found in human arthritic synovial effusions. Within 4-6 h after the intra-articular injection of 1 microgram of hrTGF-beta 1 into rat knee joints, extensive recruitment of polymorphonuclear leukocytes (PMNs) was observed. Cytochemistry and high resolution histological techniques were used to quantitate the influx of PMNs, which peaked 6 h post-injection. In a Boyden chamber assay, hrTGF-beta 1 at 1-10 fg/ml elicited a chemotactic response from PMNs greater in magnitude than that evoked by FMLP, establishing that TGF-beta 1 is an effective chemotactic agent for PMNs in vitro as well as in vivo. That PMNs may represent an important source of TGF-beta in inflammatory infiltrates was strongly suggested by a demonstration that stored TGF-beta 1 was secreted during phorbol myristate acetate-stimulated degranulation in vitro. Acid/ethanol extracts of human PMNs assayed by ELISA contained an average of 355 ng of TGF/beta 1 per 10(9) cells potentially available for secretion during degranulation of PMNs. [3H]Thymidine incorporation in vivo and autoradiography of tissue sections revealed that widespread cell proliferation was triggered by TGF-beta 1 injection. Synovial lining cells and cells located deep within the subsynovial connective tissue were identified as sources of at least some of the new cells that contribute to TGF-beta 1-induced hyperplasia. Our results demonstrate that TGF-beta is capable of exerting pathogenic effects on synovial tissue and that PMNs may represent a significant source of the TGF-beta present in synovial effusions.

Topics: Animals; Cell Division; Chemotaxis, Leukocyte; DNA; Enzyme-Linked Immunosorbent Assay; Female; Hyperplasia; Injections, Intra-Articular; Microscopy, Electron; N-Formylmethionine Leucyl-Phenylalanine; Neutrophils; Rats; Recombinant Proteins; Synovial Membrane; Synovitis; Thymidine; Transforming Growth Factor beta; Tritium