Acidic fibroblast growth factor decreases α-smooth muscle actin expression and induces apoptosis in human normal lung fibroblasts

2006 ◽  
Vol 291 (5) ◽  
pp. L871-L879 ◽  
Author(s):  
Carlos Ramos ◽  
Martha Montaño ◽  
Carina Becerril ◽  
José Cisneros-Lira ◽  
Lourdes Barrera ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Growth Rate ◽  
Smooth Muscle ◽  
Caspase 3 ◽  
Smooth Muscle Actin ◽  
Myofibroblast Differentiation ◽  
Fibroblast Growth ◽  
Muscle Actin ◽  
Acidic Fibroblast Growth Factor ◽  
Tgf Β1

Fibroblast/myofibroblast expansion is critical in the pathogenesis of pulmonary fibrosis. To date, most research has focused on profibrotic mediators, whereas studies on antifibrotic factors are scanty. In this study, we explored the effects of acidic fibroblast growth factor (FGF-1) and FGF-1 plus heparin (FGF-1+H) on fibroblast growth rate, apoptosis, and myofibroblast differentiation. Heparin was used because it participates in FGF-1 signaling. Growth rate was evaluated by WST-1 colorimetric assay, DNA synthesis by [3H]thymidine incorporation, and apoptosis by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) and cleaved caspase 3. Expression of α-smooth muscle actin (α-SMA) was examined by immunocytochemistry, flow cytometry, real-time PCR, and immunoblotting. Despite the induction of DNA synthesis, FGF-1+H significantly reduced fibroblast growth rate. This correlated with a significant increase in apoptosis, evaluated by TUNEL (41.6 ± 1.4% vs. 12.5 ± 0.6% from controls; P < 0.01) and cleaved caspase 3 (295 ± 32 vs. 200 ± 19 ng/106 cells from controls; P < 0.05). Double immunostaining (α-SMA-TUNEL) revealed that the levels of induced apoptosis were similar in fibroblasts and myofibroblasts. FGF-1+H inhibited the effect of TGF-β1 on myofibroblast differentiation. α-SMA-positive cells were reduced by immunocytochemistry from 44.5 ± 6.5% to 10.9 ± 1.9% and by flow cytometry from 30.6 ± 2.5% to 7.7 ± 0.6% ( P < 0.01). Also, FGF-1+H significantly inhibited the TGF-β1 induction of α-SMA quantified by real-time PCR and Western blot. This decrease was associated with a 35% reduction in TGF-β1-induced collagen gel contraction. The effect of FGF-1+H was mediated by a significant decrease of TGF-β1-induced Smad2 phosphorylation. FGF-1 alone exhibited similar but lower effects. These findings suggest that FGF-1 can have an antifibrogenic role, inducing apoptosis of fibroblasts and inhibiting myofibroblast differentiation.

scholarly journals Neuronal Wiskott-Aldrich syndrome protein (N-WASP) is critical for formation of α-smooth muscle actin filaments during myofibroblast differentiation

2012 ◽  
Vol 303 (8) ◽  
pp. L692-L702 ◽  
Author(s):  
Guo-Qiang Cai ◽  
Chu-Fang Chou ◽  
Meng Hu ◽  
Anni Zheng ◽  
Louis F. Reichardt ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Actin ◽  
Critical Role ◽  
Lung Fibroblasts ◽  
Myofibroblast Differentiation ◽  
Muscle Actin ◽  
Wiskott Aldrich Syndrome ◽  
Cytoplasmic Filaments ◽  
Syndrome Protein ◽  
Tgf Β1

Myofibroblasts are implicated in pathological stromal responses associated with lung fibrosis. One prominent phenotypic marker of fully differentiated myofibroblasts is the polymerized, thick cytoplasmic filaments containing newly synthesized α-smooth muscle actin (α-SMA). These α-SMA-containing cytoplasmic filaments are important for myofibroblast contractility during tissue remodeling. However, the molecular mechanisms regulating the formation and maturation of α-SMA-containing filaments have not been defined. This study demonstrates a critical role for neuronal Wiskott-Aldrich syndrome protein (N-WASP) in regulating the formation of α-SMA-containing cytoplasmic filaments during myofibroblast differentiation and in myofibroblast contractility. Focal adhesion kinase (FAK) is activated by transforming growth factor-β1 (TGF-β1) and is required for phosphorylation of tyrosine residue 256 (Y256) of N-WASP. Phosphorylation of Y256 of N-WASP is essential for TGF-β1-induced formation of α-SMA-containing cytoplasmic filaments in primary human lung fibroblasts. In addition, we demonstrate that actin-related protein (Arp) 2/3 complex is downstream of N-WASP and mediates the maturation of α-SMA-containing cytoplasmic filaments. Together, this study supports a critical role of N-WASP in integrating FAK and Arp2/3 signaling to mediate formation of α-SMA-containing cytoplasmic filaments during myofibroblast differentiation and maturation.

Expression of alpha-smooth muscle actin, TGF-β1 and TGF-β type II receptor during connective tissue contraction

1997 ◽  
Vol 33 (8) ◽  
pp. 622-627 ◽  
Author(s):  
M. Reza Ghassemifar ◽  
Roy W. Tarnuzzer ◽  
Nasser Chegini ◽  
Erkki Tarpila ◽  
Gregory S. Schultz ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Connective Tissue ◽  
Smooth Muscle Actin ◽  
Type Ii ◽  
Muscle Actin ◽  
Alpha Smooth Muscle Actin ◽  
Tissue Contraction ◽  
Tgf Β1

scholarly journals Evaluation of myofibroblasts and its association with TGF-β and IFN-γ in lesions of patients with american tegumentary leishmaniasis

2012 ◽  
Vol 87 (4) ◽  
pp. 545-549 ◽  
Author(s):  
Agostinho Gonçalves Viana ◽  
Carlos Alberto de Carvalho Fraga ◽  
Paulo Rogério Ferreti Bonan
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Actin ◽  
Healing Process ◽  
Sand Fly ◽  
Muscle Actin ◽  
Alpha Smooth Muscle Actin ◽  
Tegumentary Leishmaniasis ◽  
American Tegumentary Leishmaniasis ◽  
Ifn Γ ◽  
Tgf Β1

BACKGROUND: Leishmaniasis is caused by protozoa of Leishmania spp. genus. It is transmitted by the bite of the sand fly insect. It is believed that 12 million people are infected with this disease and that its annual incidence is 2 million; this number is increasing. OBJECTIVES: The present study aimed to evaluate the expression of myofibroblasts through alpha smooth muscle actin labeling, and to analyze their relationship with the expression of the cytokines Interferon gama (IFN-γ) and Transforming growth factor beta (TGF-β1) in lesions of American tegumentary leishmaniasis (ATL). METHODS: For this retrospective study, we gathered 28 patients diagnosed with ATL between 2002 and 2006. We verified α-SMA positivity and performed IFN-γ and TGF-β1 immunolabeling to identify the profile of these cytokines in both positive and negative cases for myofibroblasts, via immunohistochemistry, in order to assess the presence of myofibroblasts,. RESULTS: We observed that out of the 28 cases, 17 (60.71%) were positive for alpha smooth muscle actin, while 11 (39.29%) were negative, and IFN-γ was more expressed than TGF-β1 (p=0.007). The mean percentages of expression of IFN-γ and TGF-β1 in the group negative for alpha smooth muscle actin were different, with an increased expression of IFN-γ (p=0.047). However, in the group positive for alpha smooth muscle actin, there was no difference in cytokine labeling (p>0.05). CONCLUSION: We verified the presence of positive α-SMA stromal cells in the majority of the cases of ATL, indicating the presence of myofibroblasts. In cases negative for alpha smooth muscle actin, an increased expression of IFN-γ compared to TGF-β1 was observed, revealing an inflammatory phase progressing to a healing process.

MKL1 mediates TGF-β1-induced α-smooth muscle actin expression in human renal epithelial cells

2008 ◽  
Vol 294 (5) ◽  
pp. F1116-F1128 ◽  
Author(s):  
Gerard Elberg ◽  
Lijuan Chen ◽  
Dorit Elberg ◽  
Michael D. Chan ◽  
Charlotte J. Logan ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Epithelial Cells ◽  
Transforming Growth Factor ◽  
Smooth Muscle Actin ◽  
Primary Cultures ◽  
Muscle Actin ◽  
Mesenchymal Transition ◽  
Endogenous Expression ◽  
Tgf Β1 ◽  
Stimulation Of

Transforming growth factor-β1 (TGF-β1) is known to induce epithelial-mesenchymal transition in the kidney, a process involved in tubulointerstitial fibrosis. We hypothesized that a coactivator of the serum response factor (SRF), megakaryoblastic leukemia factor-1 (MKL1), stimulates α-smooth muscle actin (α-SMA) transcription in primary cultures of renal tubular epithelial cells (RTC), which convert into myofibroblasts on treatment with TGF-β1. Herein, we study the effect of MKL1 expression on α-SMA in these cells. We demonstrate that TGF-β1 stimulation of α-SMA transcription is mediated through CC(A/T)6-rich GG elements known to bind to SRF. These elements also mediate the MKL1 effect that dramatically activates α-SMA transcription in serum-free media. MKL1 fused to green fluorescent protein localizes to the nucleus and induces α-SMA expression regardless of treatment with TGF-β1. Using proteasome inhibitors, we also demonstrate that the proteolytic ubiquitin pathway regulates MKL1 expression. These data indicate that MKL1 overexpression is sufficient to induce α-SMA expression. Inhibition of endogenous expression of MKL1 by small interfering RNA abolishes TGF-β1 stimulation of α-SMA expression. Therefore, MKL1 is also absolutely required for TGF-β1 stimulation of α-SMA expression. Western blot and immunofluorescence analysis show that overexpressed and endogenous MKL1 are located in the nucleus in non-stimulated RTC. Chromatin immunoprecipitation assay demonstrates that TGF-β1 induces binding of endogenous SRF and MKL1 to the α-SMA promoter in chromatin. Since MKL1 constitutes a potent factor regulating α-SMA expression, modulation of endogenous MKL1 expression or activity may have a profound effect on myofibroblast formation and function in the kidney.

TGF-β1 induces lens cells to accumulate α-smooth muscle actin, a marker for subcapsular cataracts

1994 ◽  
Vol 13 (12) ◽  
pp. 885-890 ◽  
Author(s):  
Angela M. Hales ◽  
Mark W. Schulz ◽  
Coral G. Chamberlain ◽  
John W. McAvoy
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Actin ◽  
Muscle Actin ◽  
Lens Cells ◽  
Tgf Β1

Abstract 18216: Epigenetic Up-regulation of α-smooth Muscle Actin Expression and Cell Aggregation in Human Aortic Valve Interstitial Cells Promotes Calcific Nodule Formation

Circulation ◽  
2014 ◽  
Vol 130 (suppl_2) ◽  
Author(s):  
Rui Song ◽  
David A. Fullerton ◽  
Lihua Ao ◽  
Kesen Zhao ◽  
Xianzhong Meng
Keyword(s):  
Smooth Muscle ◽  
Aortic Valve ◽  
Smooth Muscle Actin ◽  
Cell Aggregation ◽  
Interstitial Cells ◽  
Nodule Formation ◽  
Muscle Actin ◽  
Aortic Valves ◽  
Valve Interstitial Cells ◽  
Tgf Β1

Older people are at risk of calcific aortic valve disease. Aortic valve interstitial cells (AVICs) play an important role in nodular calcification in aortic valve leaflets. AVICs in human aortic valves consist of fibroblasts and myofibroblasts that express α-smooth muscle actin (α-SMA). We have observed that AVICs of diseased aortic valves have greater osteogenic activities. However, molecular mechanism underlying AVIC formation of calcification nodules is not well understood. We hypothesized that an epigenetic mechanism promotes AVIC calcification nodule formation through induction of α-SMA expression and cell aggregation. Methods and Results: MiRNA profiles in AVICs from normal and diseased human aortic valves were analyzed by miRNA array and real-time qPCR. Diseased AVICs displayed higher levels of miR-486. Immunoblotting and immunofluorescence staining revealed that diseased AVICs had higher levels of α-SMA and α-SMA fibers. Inhibition of miR-486 by lentiviral-delivered miR-486 antagomir in diseased AVICs suppressed α-SMA expression and cell aggregation, resulting in reduced calcification nodule formation. Conversely, lentiviral-delivered miR-486 mimic in normal AVICs induced α-SMA expression and cell aggregation, leading to exacerbated calcification nodule formation. Stimulation of normal AVICs with pro-osteogenic mediators TGF-β1 and BMP-2 up-regulated miR-486 levels. MiR-486 antagomir reduced α-SMA expression, cell aggregation and calcification nodule formation in cells exposed to TGF-β1 or BMP-2. Further, miR-486 mimic induced AKT phosphorylation. Inhibition of AKT decreased α-SMA expression and cell aggregation induced by miR-486 mimic in normal AVICs. Knockdown of α-SMA suppressed cell aggregation and calcification nodule formation. Conclusions: The pro-osteogenic phenotype of AVICs of diseased aortic valves is associated with up-regulated levels of miR-486 and α-SMA. MiR-486 modulates the AKT pathway to up-regulate α-SMA expression and cell aggregation that are required for calcification nodule formation. These novel findings indicate that miR-486 contributes to the mechanism underlying aortic valve calcification and appears to be a therapeutic target for suppression of valvular osteogenic activity.

scholarly journals Host-Derived Pericytes and Sca-1+ Cells Predominate in the MART-1− Stroma Fraction of Experimentally Induced Melanoma

2011 ◽  
Vol 59 (12) ◽  
pp. 1060-1075 ◽  
Author(s):  
J. Humberto Treviño-Villarreal ◽  
Douglas A. Cotanche ◽  
Rosalinda Sepúlveda ◽  
Magda E. Bortoni ◽  
Otto Manneberg ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Endothelial Cells ◽  
Smooth Muscle ◽  
Blood Vessel ◽  
Smooth Muscle Actin ◽  
Cell Types ◽  
Muscle Actin ◽  
Cellular Composition ◽  
Tumor Capsule ◽  
Form Blood Vessel

Identification of cell types in tumor-associated stroma that are involved in the development of melanoma is hampered by their heterogeneity. The authors used flow cytometry and immunohistochemistry to demonstrate that anti–MART-1 antibodies can discriminate between melanoma and stroma cells. They investigated the cellular composition of the MART-1−, non-hematopoietic melanoma-associated stroma, finding it consisted mainly of Sca-1+ and CD146+ cells. These cell types were also observed in the skin and muscle adjacent to developing melanomas. The Sca-1+ cell population was observed distributed in the epidermis, hair follicle bulges, and tumor capsule. The CD146+ population was found distributed within the tumor, mainly associated with blood vessels in a perivascular location. In addition to a perivascular distribution, CD146+ cells expressed α-smooth muscle actin, lacked expression of endothelial markers CD31 and CD34, and were therefore identified as pericytes. Pericytes were found to be associated with CD31+ endothelial cells; however, some pericytes were also observed associated with CD31−, MART-1+ B16 melanoma cells that appeared to form blood vessel structures. Furthermore, the authors observed extensive nuclear expression of HIF-1α in melanoma and stroma cells, suggesting hypoxia is an important factor associated with the melanoma microenvironment and vascularization. The results suggest that pericytes and Sca-1+ stroma cells are important contributors to melanoma development.

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