Transforming growth factor β1 represses proximal tubular cell microRNA-192 expression through decreased hepatocyte nuclear factor DNA binding

2012 ◽  
Vol 443 (2) ◽  
pp. 407-416 ◽  
Author(s):  
Robert H. Jenkins ◽  
John Martin ◽  
Aled O. Phillips ◽  
Timothy Bowen ◽  
Donald J. Fraser
Keyword(s):  
Growth Factor ◽  
Promoter Activity ◽  
Transforming Growth Factor ◽  
Transforming Growth Factor Β1 ◽  
Luciferase Reporter ◽  
Nuclear Factor ◽  
Constitutive Promoter ◽  
Hepatocyte Nuclear Factor ◽  
Activation Of Transcription ◽  
Tgf Β1

miR (microRNA)-192 plays key roles in renal pathological and physiological responses, by repressing targets including Zeb1, Zeb2 and Wnk1. In the present study, we have assessed the regulation of miR-192 expression. We found that TGF-β1 (transforming growth factor β1) down-regulates miR-192 and miR-194, co-transcribed in the shared precursor pri-miR (primary miR transcript)-192/194. Luciferase reporter analysis showed constitutive promoter activity within nucleotides +21 to −223. We identified HNF (hepatocyte nuclear factor) and p53 binding sites within this region that were required for constitutive promoter activity, which was decreased by TGF-β1 through an Alk5-dependent mechanism. TGF-β1 treatment decreased HNF binding to the miR-194-2/192 promoter, whereas knockdown of HNF-1 inhibited mature miR-192 and miR-194 expression. miR-192, miR-194 and HNF expression were restricted to a defined subset of human tissues including kidney, small intestine, colon and liver. Our results from the present study identify co-ordinated regulation of miR-192 and miR-194, with binding of HNF and p53 transcription factors necessary for activation of transcription, and TGF-β1-mediated repression through decreased HNF binding to its cognate promoter element.

Activation of Transforming Growth Factor β1 (TGF-β1) Released by Platelets Is Enhanced by Shear and Occurs in Platelet-Rich Thrombi In Vivo.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 3632-3632
Author(s):  
Jasimuddin Ahamed ◽  
Nathalie Burg ◽  
Christin Janczak ◽  
Jihong Li ◽  
Barry S. Coller
Keyword(s):  
Growth Factor ◽  
Transforming Growth Factor ◽  
Thrombus Formation ◽  
Transforming Growth Factor Β1 ◽  
Sharp Contrast ◽  
Luciferase Reporter ◽  
Plasminogen Activator Inhibitor 1 ◽  
Total N ◽  
Tgf Β1

Abstract Platelets contain 40–100 times as much transforming growth factor β1 (TGF-β1) as other cells, and release it as an inactive (latent) complex. Although several TGF-β1 activating factors have been identified in various cell types, it is still not known whether and how platelet TGF-β1 becomes activated in vivo. In the present study, cell-free human washed platelet releasates (supernatant after stimulation with thrombin 0.125U/ml for 5 min) or serum samples were subjected to either stirring (1,200 rpm) in an aggregometer or a shear stress of 1800 s−1 in a cone and plate device at 37°C. Activation of TGF-β1 was analyzed by ELISA and selectively confirmed using a cell-based plasminogen activator inhibitor 1 (PAI-1) luciferase reporter assay. TGF-β1 was maximally released within 5 min after stimulation of platelets with thrombin (80 ± 23 ng/ml; n=3), but only 0.2% was active. TGF-β1 activity increased slowly and progressively under shear, reaching a maximum of ∼5% of total TGF-β1 (4.1 ± 1.5 ng/ml) after 1 h. In sharp contrast, in two experiments conducted without shear, active TGF-β1 constituted 0.04 and 0.2% of total at 0 time and remained nearly the same (0.1 and 0.2% of total) after 2 h. Activation of TGF-β1 in serum under shear occurred more slowly, but also reached a maximum of ∼5% of total at 2 h (4.1 ± 1.3 ng/ml active; 76 ± 14 ng/ml total; n=6). Without shear at 0 time, active TGF-β1 constituted 0.02 and 0.06% of total and at 2 h it was still only 0.05 and 0.07% of total. To asses whether TGF-β1 becomes activated in vivo during thrombosis, we induced thrombi in the carotid arteries of C57/BL mice with ferric chloride (8%) for 3 min and then removed the platelet-rich thrombi that formed after 5 or 120 min. The arteries were excised (∼4 mm) and the thrombi were removed and dispersed in buffer (200 μl) on ice for 1 h. Total TGF-β1 recovered from 5 min thrombi was 3.4 and 4.8 ng/ml and total TGF-β1 recovered from 120 min thrombi was 3.7 and 0.7 ng/ml. Active TGF-β1 could be detected in thrombi after 5 min and constituted 1.7 ± 1.1% of the total TGF-β1 recovered (n=3). After 120 min, active TGF-β1 could also be detected in thrombi and it constituted 5.1 ± 3.0% of the total recovered (p=0.14 compared to 5 min value; n=3). In sharp contrast, only 0.05% of TGF-β1 released by thrombin from mouse platelets in vitro was active without stirring or shear (n=3). These data indicate that shear can dramatically enhance TGF-β1 activation after release from platelets. Moreover, TGF-β1 activation occurs in vivo during platelet-rich thrombus formation and thus platelets may be an important source of active TGF-β1 in cardiovascular disease and wound healing.

scholarly journals Transforming growth factor-β1 selectively inhibits hepatocyte growth factor expression via a micro-RNA-199–dependent posttranscriptional mechanism

2013 ◽  
Vol 24 (13) ◽  
pp. 2088-2097 ◽  
Author(s):  
Ognoon Mungunsukh ◽  
Regina M. Day
Keyword(s):  
Growth Factor ◽  
Hepatocyte Growth Factor ◽  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Transforming Growth Factor Β1 ◽  
Micro Rna ◽  
Luciferase Reporter ◽  
And Migration ◽  
Hepatocyte Growth ◽  
Tgf Β1

Hepatocyte growth factor (HGF) is a multipotent endogenous repair factor secreted primarily by mesenchymal cells with effects on cells expressing its receptor, Met. HGF promotes normal tissue regeneration and inhibits fibrotic remodeling in part by promoting proliferation and migration of endothelial and epithelial cells and protecting these cells from apoptosis. HGF also inhibits myofibroblast proliferation. The profibrotic cytokine transforming growth factor beta 1 (TGF-β1) suppresses HGF expression but not the expression of NK2, an HGF splice variant that antagonizes HGF-induced proliferation. We investigated the mechanism for differential regulation of HGF and NK2 by TGF-β1. TGF-β1 down-regulated HGF in primary human adult pulmonary fibroblasts (HLFb) and increased the expression of miR-199a-3p, a microRNA (miRNA) associated with fibrotic remodeling. HGF and NK2 contain completely different 3′ untranslated regions (UTRs), and we determined that miR-199a-3p targeted HGF mRNA for suppression but not NK2. A pre–miR-199 mimic inhibited the expression of a luciferase reporter harboring the HGF 3′ UTR but not a pmirGLO reporter containing the NK2 3′ UTR. In contrast, an anti-miRNA inhibitor specific for miR-199a-3p prevented TGF-β1–induced reduction of both HGF mRNA and HGF protein secretion. Taken together, these findings demonstrate that HGF is distinctly regulated at the posttranscriptional level from its antagonist NK2.

Pyrrolidine dithiocarbamate attenuates paraquat-induced acute pulmonary poisoning in vivo via transforming growth factor β1 and nuclear factor κB pathway interaction

2016 ◽  
Vol 35 (12) ◽  
pp. 1312-1318 ◽  
Author(s):  
M Huang ◽  
D Lou ◽  
H-H Li ◽  
Q Cai ◽  
Y-P Wang ◽  
...  
Keyword(s):  
Growth Factor ◽  
Pulmonary Fibrosis ◽  
Protein Expression ◽  
Transforming Growth Factor ◽  
Serum Levels ◽  
Nuclear Factor Κb ◽  
Transforming Growth Factor Β1 ◽  
Pyrrolidine Dithiocarbamate ◽  
Nuclear Factor ◽  
Tgf Β1

Paraquat (PQ) exposure could cause pulmonary fibrosis. The aim of this study was to investigate the protective effect of pyrrolidine dithiocarbamate (PDTC) in an acute PQ poison model. One hundred and forty-four Sprague Dawley rats were equally divided into three experimental groups: control group, PQ group, and PQ + PDTC group. At days 1, 3, 7, 14, 28, and 56 of treatment, the serum levels of transforming growth factor β1 (TGF-β1), the levels of hydroxyproline, the protein expression of nuclear factor κB (NF-κB) pathway, and histopathological change in lung tissue were assessed. The survival rate of rats treated with PQ + PDTC was increased compared with that of rats treated only with PQ ( p < 0.05), and the occurrence of pathological changes was dramatically attenuated in the PQ + PDTC group. The serum levels of TGF-β1 and the hydroxyproline levels in the PQ group were significantly increased in a time-dependent manner compared with those in the control and PQ + PDTC groups on days 7, 14, 28, and 56 ( p < 0.05). Additionally, the protein levels of NF-κB proteins p65, inhibitor of κB (IκB) kinase (IKKβ, and IκB-α were significantly downregulated in the PQ + PDTC group as determined by array analysis. The present findings suggest that overexpression of TGF-β1 may play an important role in PQ-induced lung injury and that PDTC, a strong NF-κB inhibitor, can rescue PQ-induced pulmonary fibrosis by influencing the protein expression of NF-κB pathway.

scholarly journals JNK and p38 Inhibitors Prevent Transforming Growth Factor-β1-Induced Myofibroblast Transdifferentiation in Human Graves’ Orbital Fibroblasts

2021 ◽  
Vol 22 (6) ◽  
pp. 2952
Author(s):  
Tzu-Yu Hou ◽  
Shi-Bei Wu ◽  
Hui-Chuan Kau ◽  
Chieh-Chih Tsai
Keyword(s):  
Growth Factor ◽  
Transforming Growth Factor ◽  
Mapk Pathway ◽  
Transforming Growth Factor Β1 ◽  
Tissue Growth ◽  
Mitogen Activated Protein Kinase ◽  
Tissue Inhibitors Of Metalloproteinases ◽  
Western Blots ◽  
Orbital Fibroblasts ◽  
Tgf Β1

Transforming growth factor-β1 (TGF-β1)-induced myofibroblast transdifferentiation from orbital fibroblasts is known to dominate tissue remodeling and fibrosis in Graves’ ophthalmopathy (GO). However, the signaling pathways through which TGF-β1 activates Graves’ orbital fibroblasts remain unclear. This study investigated the role of the mitogen-activated protein kinase (MAPK) pathway in TGF-β1-induced myofibroblast transdifferentiation in human Graves’ orbital fibroblasts. The MAPK pathway was assessed by measuring the phosphorylation of p38, c-Jun N-terminal kinase (JNK), and extracellular-signal-regulated kinase (ERK) by Western blots. The expression of connective tissue growth factor (CTGF), α-smooth muscle actin (α-SMA), and fibronectin representing fibrogenesis was estimated. The activities of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) responsible for extracellular matrix (ECM) metabolism were analyzed. Specific pharmacologic kinase inhibitors were used to confirm the involvement of the MAPK pathway. After treatment with TGF-β1, the phosphorylation levels of p38 and JNK, but not ERK, were increased. CTGF, α-SMA, and fibronectin, as well as TIMP-1 and TIMP-3, were upregulated, whereas the activities of MMP-2/-9 were inhibited. The effects of TGF-β1 on the expression of these factors were eliminated by p38 and JNK inhibitors. The results suggested that TGF-β1 could induce myofibroblast transdifferentiation in human Graves’ orbital fibroblasts through the p38 and JNK pathways.

scholarly journals Transforming growth factor-β1-induced N-cadherin drives cell–cell communication through connexin43 in osteoblast lineage

2021 ◽  
Vol 13 (1) ◽  
Author(s):  
Yueyi Yang ◽  
Wenjing Liu ◽  
JieYa Wei ◽  
Yujia Cui ◽  
Demao Zhang ◽  
...  
Keyword(s):  
Growth Factor ◽  
Gap Junction ◽  
Cell Line ◽  
Transforming Growth Factor ◽  
Transforming Growth Factor Β1 ◽  
Cx43 Expression ◽  
Mc3t3 Cell ◽  
Primary Osteoblasts ◽  
Tgf Β1 ◽  
Cell Cell

AbstractGap junction (GJ) has been indicated to have an intimate correlation with adhesion junction. However, the direct interaction between them partially remains elusive. In the current study, we aimed to elucidate the role of N-cadherin, one of the core components in adhesion junction, in mediating connexin 43, one of the functional constituents in gap junction, via transforming growth factor-β1(TGF-β1) induction in osteoblasts. We first elucidated the expressions of N-cadherin induced by TGF-β1 and also confirmed the upregulation of Cx43, and the enhancement of functional gap junctional intercellular communication (GJIC) triggered by TGF-β1 in both primary osteoblasts and MC3T3 cell line. Colocalization analysis and Co-IP experimentation showed that N-cadherin interacts with Cx43 at the site of cell–cell contact. Knockdown of N-cadherin by siRNA interference decreased the Cx43 expression and abolished the promoting effect of TGF-β1 on Cx43. Functional GJICs in living primary osteoblasts and MC3T3 cell line were also reduced. TGF-β1-induced increase in N-cadherin and Cx43 was via Smad3 activation, whereas knockdown of Smad3 signaling by using siRNA decreased the expressions of both N-cadherin and Cx43. Overall, these data indicate the direct interactions between N-cadherin and Cx43, and reveal the intervention of adhesion junction in functional gap junction in living osteoblasts.

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