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Smoothelin

About: Smoothelin is a research topic. Over the lifetime, 264 publications have been published within this topic receiving 14069 citations.


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Journal ArticleDOI
TL;DR: A novel transgenic mouse model of scleroderma offers insight into the altered biomechanical properties previously reported for large elastic arteries in human SSc and suggests a role for perturbed TGF-β and endothelin activity in this process.
Abstract: Introduction: Vasculopathy, including altered vasoreactivity and abnormal large vessel biomechanics, is a hallmark of systemic sclerosis (SSc). However, the pathogenic link with other aspects of the disease is less clear. To assess the potential role of transforming growth factor beta (TGF-β) overactivity in driving these cardiovascular abnormalities, we studied a novel transgenic mouse model characterized by ligand-dependent activation of TGF-β signaling in fibroblasts. Methods: The transgenic mouse strain Tβ RIIΔk-fib is characterized by balanced ligand-dependent upregulation of TGF-β signaling. Aortic and cardiac tissues were examined with histologic, biochemical, and isolated organ bath studies. Vascular and perivascular architecture was examined by hematoxylin and eosin (H&E) and special stains including immunostaining for TGF-β1 and phospho-Smad2/3 (pSmad2/3). Confirmatory aortic smooth muscle cell proliferation, phenotype, and functional assays, including signaling responses to exogenous TGF-β and endothelin-1, were performed. Aortic ring contractile responses to direct and receptor-mediated stimulation were assessed. Results: Aortic ring contractility and relaxation were diminished compared with wild-type controls, and this was associated with aortic adventitial fibrosis confirmed histologically and with Sircol assay. TGF-β1 and pSmad 2/3 expression was increased in the adventitia and smooth muscle layer of the aorta. Aortic smooth muscle cells from transgenic animals showed significant upregulation of TGF-β- responsive genes important for cytoskeletal function, such as transgelin and smoothelin, which were then resistant to further stimulation with exogenous TGF-β1. These cells promoted significantly more contraction of free floating type I collagen lattices when compared with the wildtype, but were again resistant to exogenous TGF-β1 stimulation. Aortic ring responses to receptor-mediated contraction were reduced in the transgenic animals. Specifically, bosentan reduced endothelin-mediated contraction in wild-type animals, but had no effect in transgenic animals, and endothelin axis gene expression was altered in transgenic animals. Transgenic mice developed cardiac fibrosis. Conclusions: The histologic, biochemical, and functional phenotype of this transgenic mouse model of scleroderma offers insight into the altered biomechanical properties previously reported for large elastic arteries in human SSc and suggests a role for perturbed TGF-β and endothelin activity in this process.

32 citations

Journal ArticleDOI
TL;DR: The results strongly suggest that the smoothelin gene contains a vascular and a visceral smooth muscle promoter that plays a role in the modulation of the contractile properties of different smooth muscle cell types.
Abstract: Smoothelin is a cytoskeletal protein specifically expressed in differentiated smooth muscle cells and has been shown to colocalize with smooth muscle alpha actin In addition to the small smoothelin isoform of 59 kD, we recently identified a large smoothelin isoform of 117 kD The aim of this study was to identify and characterize novel smoothelin isoforms The genomic structure and sequence of the smoothelin gene were determined by genomic PCR, RT-PCR and DNA sequencing Comparison of the cDNA and genomic sequences shows that the small smoothelin isoform is generated by transcription initiation 10 kb downstream of the start site of the large isoform In addition to the known smoothelin cDNA (c1 isoform) we identified two novel cDNA variants (c2 and c3 isoform) that are generated by alternative splicing within a region, which shows similarity to the spectrin family of F-actin cross-linking proteins Visceral organs express the c1 form, while the c2 form prevails in well-vascularized tissue as analyzed by RT-PCR We then generated specific antibodies against the major smoothelin isoforms and could show by Western blotting and immunohistochemistry that the large isoform is specifically expressed in vascular smooth muscle cells, while the small isoform is abundant in visceral smooth muscle These results strongly suggest that the smoothelin gene contains a vascular and a visceral smooth muscle promoter The cell-type-specific expression of smoothelin isoforms that are associated with actin filaments may play a role in the modulation of the contractile properties of different smooth muscle cell types

32 citations

Journal ArticleDOI
TL;DR: This paper outlined a step-by-step procedure for isolating aortic VSMCs from adult C57BL6J male mice by enzymatic digestion of the aorta using collagenase to characterize VSMC phenotype by gene expression and immunofluorescence.
Abstract: Vascular smooth muscle cells (VSMCs) play important roles in cardiovascular disorders and biology. Outlined in this paper is a step-by-step procedure for isolating aortic VSMCs from adult C57BL6J male mice by enzymatic digestion of the aorta using collagenase. The plating, culturing, and subculturing of the isolated cells are discussed in detail along with techniques to characterize VSMC phenotype by gene expression and immunofluorescence. Traction force microscopy was used to characterize contractility of single subcultured VSMCs at baseline.

31 citations

Journal ArticleDOI
TL;DR: The results confirmed that rat prostatic SMCs are affected by androgen deprivation, and support the idea that SMCs may modulate their phenotypes (contractile vs synthetic) without changing their differentiation states.
Abstract: Smooth muscle (SM) is an important component of the prostatic stroma. We previously showed that, despite extensive morphologic changes, smooth muscle cells (SMCs) of the rat ventral prostate preserve some differentiation markers 21 days after castration. In the present study, we investigated whether the expression of SMC markers is preserved in the rat ventral prostate after long-term castration. Adult Wistar rats were castrated and sacrificed 100 days after surgery. The ventral prostates were processed for histology, stereology, immunocytochemistry (SM alpha-actin and SM-myosin heavy chain [MHC]), transmission electron microscopy (TEM), and reverse transcription polymerase chain reaction (smoothelin, sm22, and calponin). The prostates of castrated rats showed significant weight reduction, corresponding to only 5.6% of the control. Stereology showed that SMCs occupied the same proportion of the prostate volume but suffered a significant reduction in absolute volume (5.5% of control). The SMCs were retracted and showed spinous outlines. TEM revealed the presence of an abundant myofibrillar component, dense plaques, and an external lamina in these cells. SMCs were reactive to antibodies against SM alpha-actin and SM-MHC and expressed mRNA for smoothelin, sm22, and calponin. The results confirmed that rat prostatic SMCs are affected by androgen deprivation. Although showing marked phenotypic changes, these cells expressed SMC markers at the protein (SM alpha-actin and SM-MHC) and mRNA (smoothelin, sm22, and calponin) levels. These observations support the idea that SMCs may modulate their phenotypes (contractile vs synthetic) without changing their differentiation states.

31 citations

Journal ArticleDOI
TL;DR: It is shown here that hAFSCs under selective culture conditions are able to give rise to functional SMCs, for the first time that efficient SMCs can be obtained from human amniotic fluid stem cells.
Abstract: Functional smooth muscle engineering requires isolation and expansion of smooth muscle cells (SMCs), and this process is particularly challenging for visceral smooth muscle tissue where progenitor cells have not been clearly identified. Herein we showed for the first time that efficient SMCs can be obtained from human amniotic fluid stem cells (hAFSCs). Clonal lines were generated from c-kit(+) hAFSCs. Differentiation toward SM lineage (SMhAFSCs) was obtained using a medium conditioned by PDGF-BB and TGF-β1. Molecular assays revealed higher level of α smooth muscle actin (α-SMA), desmin, calponin, and smoothelin in SMhAFSCs when compared to hAFSCs. Ultrastructural analysis demonstrated that SMhAFSCs also presented in the cytoplasm increased intermediate filaments, dense bodies, and glycogen deposits like SMCs. SMhAFSC metabolism evaluated via mass spectrometry showed higher glucose oxidation and an enhanced response to mitogenic stimuli in comparison to hAFSCs. Patch clamp of transduced hAFSCs with lentiviral vectors encoding ZsGreen under the control of the α-SMA promoter was performed demonstrating that SMhAFSCs retained a smooth muscle cell-like electrophysiological fingerprint. Eventually SMhAFSCs contractility was evident both at single cell level and on a collagen gel. In conclusion, we showed here that hAFSCs under selective culture conditions are able to give rise to functional SMCs.

31 citations


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Performance
Metrics
No. of papers in the topic in previous years
YearPapers
202113
202012
20196
20188
201713
20165