Production Of Transforming Growth Factor Research Articles

The comparison of serum TGF-beta levels and associated polymorphisms in patients with coronary artery ectasia and normal coronary artery

BackgroundCoronary artery ectasia (CAE) is described as the enlargement of a coronary artery segment by 1.5 times or more, which is generally associated with the atherosclerotic process. Atherosclerotic changes lead to arterial remodeling result in CAE. In our study, we measured serum transforming growth factor (TGF)-β1 levels, which have a protective role against atherosclerosis. Further, we aimed to assess the TGF-β1 gene variants rs1800469 (–509C>T, c.−1347C>T) and rs1800470 (c.+29T>C, p.Pro10Leu, rs1982073), which might have an effect on TGF production. Overall, 2877 patients were screened including 56 patients with CAE and 44 patients with normal coronary arteries who were included in the study. Serum TGF-β1 levels were measured using ELISA and compared between two groups. Additionally, TGF-β1 rs1800469 and rs1800470 gene variations were determined using TaqMan® SNP Genotyping Assays.ResultsSerum TGF-β1 levels were significantly lower in patients with CAE than in controls (p=0.012). However, there was no difference in terms of the genotype and allele distributions of TGF-β1 rs1800469 and rs1800470 polymorphisms. Serum TGF-β1 levels were higher in individuals carrying the TGF-β1 rs1800470 G allele (GG+AG) than in individuals with normal homozygous AA genotype in the CAE group (p=0.012).ConclusionOur findings suggest that lower serum TGF-β1 levels are associated with an increased risk for CAE development and that TGF-β1 polymorphisms exert a protective effect. Furthermore, TGF-β1 rs1800470 G allele carriers were shown to have higher TGF-β1 levels in the CAE group. This suggests that having the G allele in the TGF-β1 rs1800470 polymorphism could prevent CAE development.

Association between IL-17 and IgA in the joints of patients with inflammatory arthropathies

BackgroundHyperactive secretion and pathogenic effects of interleukin (IL)-17 and IgA have been detected in different arthropathies. Recent evidence has revealed that TH17 cytokines regulate mucosal IgA secretion. However, it is unknown whether and how IL-17 mediates synovial IgA production. Here we aim to investigate the connection of synovial IL-17 with IgA production in the joint.In this study we included synovial fluids (SF) from patients with rheumatoid arthritis (RA; n = 66), spondyloarthritis (SpA; n = 18) and osteoarthritis (OA; n = 36). The levels of IL-17, IL-6, transforming growth factor (TGF)-β1, B-cell-activating factor of the TNF family (BAFF) and anti-lipopolyssacharide (LPS) immunoglobulin (Ig)A were investigated by enzyme-linked immunosorbent assay (ELISA). Total IgA was measured by radial immunodiffusion assay. Synovial fluid-derived mononuclear cells (SFMC) were stimulated with bacterial antigens or SF-conditioned media, and cytokines and IgA were analyzed in the supernatants.ResultsIL-17, IL-6 and TGF-β1 were increased in SF from both RA and SpA compared with OA patients. Concentration of IL-17 correlated with the disease activity score (DAS)-28, IL-6 and anti-LPS IgA levels. Bacterial-stimulated SFMCs from RA and SpA patients secreted higher IL-17 than vehicle-stimulated SFMCs. Conditioned media with SF containing IL-17 induced anti-LPS IgA production by SFMCs which was independent of IL-6 activity. Concentrations of synovial TGF-β1 and BAFF correlated with anti-LPS and total IgA levels, respectively. Blockade of IL-17 decreased the production of TGF-β1 and anti-LPS IgA by SF-stimulated SFMCs.ConclusionsThis study reports a connection between IL-17 and IgA secretion in the joint. In addition, it demonstrates that enterobacterial antigens trigger synovial IL-17 production, and that TGF-β1 and BAFF may mediate the effect of IL-17 on IgA production. This circuit may contribute to the pathogenesis of inflammatory joint diseases.

The Ang-(1–7)/Mas-1 axis attenuates the expression and signalling of TGF-β1 induced by AngII in mouse skeletal muscle

AngII (angiotensin II) induces pathological conditions such as fibrosis in skeletal muscle. In this process, AngII increases ROS (reactive oxygen species) and induces a biphasic phosphorylation of p38 MAPK (mitogen-activated protein kinase). In addition, AngII stimulates the expression and production of TGF (transforming growth factor)-β1 via a mechanism dependent on ROS production mediated by NADPH oxidase (NOX) and p38 MAPK activation. In the present study, we investigated whether Ang-(1-7) [angiotensin-(1-7)], through the Mas-1 receptor, can counteract the signalling induced by AngII in mouse skeletal muscle and cause a decrease in the expression and further activity of TGF-β1in skeletal muscle cells. Our results show that Ang-(1-7) decreased the expression of TGF-β1 induced by AngII in a dose-dependent manner. In addition, we observed that Ang-(1-7) prevented the increase in TGF-β1 expression induced by AngII, ROS production dependent on NOX and the early phase of p38 MAPK phosphorylation. Interestingly, Ang-(1-7) also prevented the late phase of p38 MAPK phosphorylation, Smad-2 phosphorylation and Smad-4 nuclear translocation, an increase in transcriptional activity, as determined using the p3TP-lux reporter, and fibronectin levels, all of which are dependent on the TGF-β1 levels induced by AngII. We also demonstrated that Ang-(1-7) prevented the increase in TGF-β1, fibronectin and collagen content in the diaphragm of mice infused with AngII. All of these effects were reversed by the administration of A779, indicating the participation of Mas-1. In conclusion, our findings support the hypothesis that Ang-(1-7) decreases the expression and further biological activity of TGF-β1 induced by AngII invitro and invivo.

Immunosuppression-induced bronchial epithelial–mesenchymal transition: A potential contributor to obliterative bronchiolitis

Obliterative bronchiolitis is the predominant histopathologic finding in patients with chronic rejection after lung transplant. This fibroproliferative transformation within small airways of lung allograft is poorly understood; however, studies suggest epithelial-mesenchymal transition plays a role. Transplant immunosuppressive therapy has been shown to cause epithelial-mesenchymal transition in renal tubular epithelial cells, with subsequent fibrosis. This study explored whether immunosuppressive therapy contributes to epithelial-mesenchymal transition in airway epithelial cells. Bronchial epithelial cell line RL-65 was treated 3 to 5 days with several immunosuppressive agents, including cyclosporine (INN ciclosporin), tacrolimus, azathioprine, mycophenolic acid, sirolimus, prednisone, and transforming growth factor β1 as control. We then analyzed cells for presence of mesenchymal morphology and protein markers. Treatment with cyclosporine, azathioprine, mycophenolate, and sirolimus resulted in elongated and irregular cell shape, and all but azathioprine showed loss of cell-cell adhesions relative to vehicle-treated cells. Expressions of extracellular matrix proteins, fibronectin and collagen, along with mesenchymal marker, vimentin, were significantly upregulated. Immunofluorescence showed loss of E-cadherin at cell membranes and cytoskeletal rearrangements typical of epithelial-mesenchymal transition. These immunosuppressive agents also increased transforming growth factor produced by cells; however, tacrolimus- and prednisone-treated cells maintained epithelial morphology, baseline levels of matrix protein expression, and transforming growth factor production levels. Overall, we found that certain immunosuppressive agents may contribute to partial epithelial-mesenchymal transition in bronchial epithelial cells, specifically increasing production of excessive extracellular matrix proteins. This may provide novel insights into the pathogenesis of obliterative bronchiolitis after lung transplant.

Effect of Peroxisome Proliferator-Activated Receptor-Gamma on Airway Smooth Muscle Thickening in a Murine Model of Chronic Asthma

Background: Asthma is characterized by airway hyperresponsiveness (AHR), inflammation and remodeling. Peroxisome proliferator-activated receptors (PPARs) were reported to regulate inflammatory responses in many cells. In this study we examined the effect of a PPAR-γ agonist on the airway smooth muscle and the production of transforming growth factor (TGF)-β1 and vascular endothelial growth factor (VEGF). Methods: We developed a mouse model of airway remodeling including smooth muscle thickening in which ovalbumin (OVA)-sensitized mice were repeatedly exposed to intranasal OVA administration twice a week for 3 months. Mice were treated intranasally with ciglitazone during OVA challenge. Results: Mice chronically exposed to OVA developed sustained eosinophilic airway inflammation and AHR to methacholine compared with control mice. In addition, the mice chronically exposed to OVA developed features of airway remodeling, including thickening of the peribronchial smooth muscle layer. Administration of ciglitazone intranasally significantly inhibited the development of AHR, eosinophilic inflammation, and importantly, airway smooth muscle remodeling in mice chronically exposed to OVA. However, intranasal ciglitazone treatment did not reduce the level of TGF-β1 and VEGF in bronchoalveolar lavage fluid. Conclusions: These results suggest that intranasal administration of ciglitazone can prevent not only airway inflammation, but also airway remodeling associated with chronic allergen challenge. The mechanism might not be related to VEGF and TGF production. Further study is needed.

The pattern of cytokine gene expression in lymphoid organs and peripheral blood mononuclear cells of mice with experimental allergic encephalomyelitis

We previously observed Th1-dominated response in the central nervous system (CNS) of mice during the course of experimental allergic encephalomyelitis (EAE) with a semiquantitative reverse transcriptase–polymerase chain reaction (RT/PCR) analysis. We report here that mRNA levels for both inflammatory cytokines including interleukin (IL)-l β, IL-2, IL-6, interferon (IFN)- γ, tumor necrosis factor (TNF)- α and TNF- β and immunoregulatory cytokines including IL-4, IL-10 and transforming growth factor (TGF)- β were up-regulated in the preclinical and/or acute phase but down-regulated in the recovery phase of EAE in lymph node (LN) of mice. Similar profiles for cytokine mRNA levels were also observed in spleen and peripheral blood mononuclear cells (PBMC). The present study also showed that a significant down-regulation of the mRNA level for IL-6 in the acute phase as compared with the preclinical phase, and a significant reduction of the mRNA level for TGF- β in the preclinical and acute phase as compared with the corresponding mRNA levels in the control mice treated with complete Freund's adjuvant alone were characteristic in peripheral immune organs of mice with EAE. These results indicate that no particular bias in cytokine production occurred in peripheral immune organs of mice with actively induced relapsing EAE, and that the relative reduction in production of TGF- β or IL-6 in peripheral circulation might participate in the induction or remission of EAE, respectively. Our results using the animal model of multiple sclerosis (MS) suggested that the mRNA levels for IL-6 and TGF- β in PBMC from patients with MS may be a good indicator to assess the disease activity or to predict relapse.

Normalization of epidermal growth factor receptor and transforming growth factor production in drug resistant variants derived from adenovirus transformed cells.

Variants (G2, G5) resistant to the cancer chemotherapeutic drug methylglyoxal bis (guanylhydrazone) (MGBG) were isolated from adenovirus type 2 transformed rat brain cells (F4; Sircar et al., 1987). Although at least one of these variants continued to express the adenovirus Ela and Elb transforming proteins, they both exhibited a detransformed phenotype as witnessed by flat morphology, loss of anchorage independent growth, and tumor forming capacity. Reverse transformation suggested the possibility of changes in growth factor receptors and the production of transforming growth factors. To test this possibility, we investigated the status of epidermal growth factor receptors (EGF-r) and transforming growth factor alpha (TGF-alpha) production in F4, G2 and G5 cells. The level of 125I-labeled EGF binding to intact drug resistant cells increased by 2- to 3-fold compared to the transformed parental cell. Scatchard analysis suggests that increased binding was the result of increased receptor levels rather than altered affinity of receptor for ligand. The production of growth factors which compete with 125I-labeled EGF binding declined in the detransformed G2 and G5 cells to a level intermediate between transformed (F4) and normal cells (FR3T3). EGF-receptor increase and the complementary decrease in growth factor production in the drug resistant variants may be associated with detransformation.

32] Soft agar growth assays for transforming growth factors and mitogenic peptides

Publisher Summary Many different tumor cells produce and release transforming growth factors (TGFs) that influence the growth and behavior of the cells that produce them. Consequently, the production of TGFs may have important effects on the growth of tumors in vivo . The ability to grow in soft agar is one of the best indicators of a cell's tumorigenic capacity. TGFs can induce the soft agar growth of nontransformed cells. TGFs are usually assayed by their ability to induce the soft agar growth of nontransformed cells. This chapter describes methods to examine TGFs and related growth factors for their ability to influence anchorage-independent growth. The chapter discusses the most commonly used indicator cell lines, the general methodology employed, the selection of appropriate culture media, and a rapid assay that is suitable for testing small numbers of cells for the release of factors that influence anchorage-independent growth.

Flat revertants derived from Kirsten murine sarcoma virus-transformed cells produce transforming growth factors.

Two flat cellular revertant cell lines, F-2 and C-11, which were originally selected from the DT line of Kirsten murine sarcoma virus (Ki-MuSV)-transformed NIH/3T3 cells, were examined for the production of transforming growth factors (TGFs). The revertant cells fail to grow in semisolid medium as colonies and exhibit a markedly reduced level of tumorigenicity in nude mice, although they are known to express high levels of p21ras, the product of the Kirsten sarcoma virus oncogene, ras, and they contain a rescuable transforming virus. TGF activity associated with the transformed, revertant, and non-transformed cell lines was measured by the ability of concentrated conditioned medium (CM) from these cells to induce normal rat kidney (NRK) and NIH/3T3 cells to form colonies in semisolid agar suspension cultures and to inhibit the binding of 125I epidermal growth factor (EGF) to specific cell surface receptors. CM from the transformed DT cells and from both the F-2 and C-11 revertants contains TGF activity, in contrast to CM obtained from normal NIH/3T3 cells. Furthermore, unlike NIH/3T3 cells, neither the DT nor the revertant cells were able to bind 125I EGF. All four cell lines were able to proliferate in serum-free medium supplemented with transferrin, insulin, EGF, and Pedersen fetuin. However, in basal medium lacking these growth factors, only DT cells and, to a lesser extent, the revertant cells were able to grow. These results suggest that the F-2 and C-11 revertants fail to exhibit all of the properties associated with transformation because the series of events leading to the transformed phenotype is blocked at a point(s) distal both to the expression of the p21 ras gene product and also to the production of TGFs and that the production of TGFs may be necessary but not sufficient for maintaining the transformed state.

Transformation induced by Abelson murine leukemia virus involves production of a polypeptide growth factor.

Rat embryo fibroblasts transformed by Abelson murine leukemia virus (MuLV) produce and release a transforming growth factor (TGF). Production of this factor is correlated with a tyrosine-specific protein kinase that is functionally active and is associated with the major Abelson MuLV gene product, P120. Transformation-defective mutants of Abelson MuLV do not transform cells, do not have their virus coded transforming gene product phosphorylated in tyrosine, and do not induce TGF production. Abelson MuLV-induced TGF morphologically transforms cells in culture, competes with 125I-labeled epidermal growth factor (EGF) for binding to cell receptors, and induces phosphorylation of tyrosine acceptor sites in the 160,000-dalton EGF membrane receptor. After purification to homogeneity, Abelson virus-induced TGF migrates as a single polypeptide with an apparent size of 7400 daltons as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.

Transforming growth factors (TGFs): properties and possible mechanisms of action.

Transforming growth factors (TGFs) are growth-promoting polypeptides that cause phenotypic transformation and anchorage-independent growth of normal cells. They have been isolated from several human and animal carcinoma and sarcoma cells. One TGF is sarcoma growth factor (SGF) which is released by murine sarcoma virus-transformed cells. The TGFs interact with epidermal growth factor (EGF) cell membrane receptors. TGFs are not detectable in culture fluids from cells which contain high numbers of free EGF cell membrane receptors. SGF acts as a tumor promoter in cell culture systems and its effect on the transformed phenotype is blocked by retinoids (vitamin A and synthetic analogs). The production of TGFs by transformed cells and the responses of normal cells to the addition of TGFs to the culture medium raise the possibility that cells "autostimulate" their own growth by releasing factors that rebind at the cell surface. The term "autocrine secretion" has been proposed for this type of situation where a cell secretes a hormone-like substance for which it has external cell membrane receptors. The autocrine concept may provide a partial explanation for some aspects of tumor cell progression.