Showing papers on "Smoothelin published in 2008"
TL;DR: To determine whether skeletal muscle‐derived stem cells convert into smooth muscle cells (SMCs) both in vitro and in vivo, and in so doing ameliorate the erectile dysfunction of aged rats, and whether endogenous stem cells are present in the rat corpora cavernosa, MDSCs are studied.
Abstract: Objective To determine whether skeletal muscle-derived stem cells (MDSCs) convert into smooth muscle cells (SMCs) both in vitro and in vivo, and in so doing ameliorate the erectile dysfunction (ED) of aged rats, and whether endogenous stem cells are present in the rat corpora cavernosa. Materials and methods MDSCs were obtained from mouse muscle, and shown by immunocytochemistry for alpha-smooth muscle actin (alpha SMA) to originate in vitro in myofibroblasts and SMCs, discriminating SMCs by calponin 1 expression. In vivo these MDSCs, labelled with 4',6-diamidino-2-phenylindole, were implanted into the corpora cavernosa of young adult (5-month old) and aged (20-month old) rats for 2 and 4 weeks. Histological changes were assessed by immunohistochemistry and quantitative Western blot. Functional changes were determined by electrical field stimulation (EFS) of the cavernosal nerve. Results The exogenous cells replicated and converted into SMCs, as shown in corporal tissue sections by confocal immunofluorescence microscopy for proliferating cell nuclear antigen (PCNA), alpha SMA, and smoothelin, and also by Western blot for alpha SMA and PCNA. MDSC differentiation was confirmed by the activation of the alpha SMA promoter-linked beta-galactosidase in transfected cells, both in vitro and after implantation in the corpora. Putative endogenous stem cells were shown in corporal tissue sections and Western blots by detecting CD34 and a possible Sca1 variant. EFS showed that implanted MDSCs raised in aged rats the maximal intracavernosal pressure/mean arterial pressure levels above (2 weeks) or up to (4 weeks) those of young adult rats. Conclusions MDSCs implanted into the corpora cavernosa of aged rats converted into SMCs and corrected ED, and endogenous cells expressing stem cell markers were also found in untreated tissue. This suggests that exogenous stem cell implantation and/or endogenous stem cell modulation might be viable therapeutic approaches for ageing-related ED.
109 citations
TL;DR: It is concluded that the mature podocyte in vitro possesses functional apparatus of contractile smooth muscle cells, with potential implications for its in vivo ability to regulate glomerular dynamic and permeability characteristics.
Abstract: The glomerular podocyte is a highly specialized cell, with the ability to ultrafilter blood and support glomerular capillary pressures. However, little is known about either the genetic programs leading to this functionality or the final phenotype. We approached this question utilizing a human conditionally immortalized cell line, which differentiates from a proliferating epithelial phenotype to a differentiated form. We profiled gene expression during several time points during differentiation and grouped the regulated genes into major functional categories. A novel category of genes that was upregulated during differentiation was of smooth muscle-related molecules. We further examined the smooth muscle phenotype and showed that podocytes consistently express the differentiated smooth muscle markers smoothelin and calponin and the specific transcription factor myocardin, both in vitro and in vivo. The contractile contribution of the podocyte to the glomerular capillary is controversial. We demonstrated using two novel techniques that podocytes contract vigorously in vitro when differentiated and in real time were able to demonstrate that angiotensin II treatment decreases monolayer resistance, morphologically correlating with enhanced contractility. We conclude that the mature podocyte in vitro possesses functional apparatus of contractile smooth muscle cells, with potential implications for its in vivo ability to regulate glomerular dynamic and permeability characteristics.
92 citations
TL;DR: The results suggest that hair follicles may be an easily accessible, autologous, and rich source of functional SMPC for cardiovascular tissue engineering and regenerative medicine.
Abstract: Aims We hypothesized that hair-follicle stem cells can differentiate toward smooth contractile muscle cells, providing an autologous cell source for cardiovascular tissue regeneration.
Methods and results Smooth muscle progenitor cells (SMPCs) were obtained from ovine hair follicles using a tissue-specific promoter and fluorescence-activated cell sorting. Hair-follicle smooth muscle progenitor cells (HF-SMPCs) expressed several markers of vascular smooth muscle including α-actin, calponin, myosin heavy chain (MHC), caldesmon, smoothelin, and SM22. HF-SMPCs were highly proliferative and showed high clonogenic potential without any signs of chromosomal abnormalities as evidenced by karyotype analysis. HF-SMPCs compacted fibrin hydrogels to a similar extent as vascular smooth muscle cells from ovine umbilical veins (V-SMCs), indicating the development of the force-generating machinery. In addition, cylindrical tissue equivalents prepared with HF-SMPCs displayed significant contractility in response to vasoactive agonists including KCl and the thromboxane A2 mimetic U46619, suggesting that these cells had developed receptor and non-receptor-mediated pathways of contractility. Finally, transforming growth factor-β1 promoted differentiation of HF-SMPCs toward a mature SMC phenotype as suggested by increased expression of MHC and enhanced matrix compaction.
Conclusion Our results suggest that hair follicles may be an easily accessible, autologous, and rich source of functional SMPC for cardiovascular tissue engineering and regenerative medicine.
83 citations
TL;DR: Different aspects of the phenotypic changes associated with hypertension occurred at different rates: cell proliferation and collagen production occurred early and peaked by 2 weeks, whereas changes in contractile protein expression continued to decrease over the entire 8-week study period.
Abstract: Arteries undergo marked structural and functional changes in human and experimental hypertension that generally involve smooth muscle cell (SMC) hypertrophy/hyperplasia as well as abnormal extracellular matrix turnover. In this study we examined time courses of changes in SMC activity and matrix protein content in a novel mini-pig aortic coarctation model. Cell proliferation was evaluated by immunostaining of Ki-67, apoptosis was assessed by TUNEL, and phenotypic changes were monitored by immunostaining three SMC contractile markers (caldesmon, calponin, and smoothelin). Changes in medial collagen and elastin were examined by picrosirius red and Verhoeff–van Gieson staining, respectively. LabVIEW-based image analysis routines were developed to objectively and efficiently quantify the (immuno)histochemical results. We found that significant cell proliferation and matrix production occurred in the early stages of this coarctation model and then declined gradually; the SMCs also tended to exhibit a less cont...
65 citations
TL;DR: It is demonstrated that fibrocytes contribute to formation of the fibrous cap, and it is suggested that monocytes may play a more crucial role in the clinical outcome of atherosclerosis than previously realized as they not only contribute directly to plaque instability (through foam cell formation), but also promote plaque stability by transformation into a Fibrocyte.
64 citations
Journal Article•
TL;DR: In this article, the authors examined human carotid endarterectomy specimens for the presence of fibrocytes by immunohistochemistry staining for CD34/procollagen I and leukocyte specific protein-1/pro-collagen-I. They found that the cells possessed a smooth muscle-like spindle shape, produced collagen, and consistent with being SM-like cells they co-localized with transformation growth factor beta, but not serum amyloid P factors, known to promote and inhibit their formation.
Abstract: Aim The stability of an atherosclerotic plaque is a key-determining factor in the clinical outcome of cardiovascular disease. In this respect, smooth muscle (SM) alfa actin positive cells play an important role in maintaining plaque stability through formation of a fibrous cap. Recent evidence suggests that circulating progenitors may be a source of these cells. We hypothesized that they may be fibrocytes bone-marrow derived cells that acquire SM-like characteristics, including the expression of SM alfa actin. Methods We examined human carotid endarterectomy specimens for the presence of fibrocytes by immunohistochemistry staining for CD34/procollagen I and leukocyte specific protein-1/procollagen I) and examined fibrocyte differentiation in vitro. Results Fibrocytes were found in regions of plaque growth/healing. They possessed a SM-like spindle shape, produced collagen, and consistent with being fibrocytes they co-localized with transformation growth factor beta, but not serum amyloid P factors, known to promote and inhibit their formation, respectively. While fibrocytes were detected in regions of new growth in 35/40 specimens, only 1/3 of the specimens expressed the SM cell marker calponin, and smoothelin was absent, in these regions. Conclusion Our results demonstrate that fibrocytes contribute to formation of the fibrous cap. With fibrocytes being a monocyte derived cell, we suggest that monocytes may play a more crucial role in the clinical outcome of atherosclerosis than previously realized as they not only contribute directly to plaque instability (through foam cell formation), but also promote plaque stability by transformation into a fibrocyte.
63 citations
TL;DR: Stromal cells failed to adhere to the scaffold in serum-free conditions, but laminin pre-coating of the scaffolds prevented serum adsorption and promoted cell attachment and differentiation.
59 citations
TL;DR: These results identify smoothelins as key determinants of arterial smooth muscle contractility and cardiovascular performance and studies on mutations in the Smtn gene or alterations in smoothelin levels in connection to hypertension in humans are warranted.
Abstract: Background— Smoothelins are actin-binding proteins that are abundantly expressed in healthy visceral (smoothelin-A) and vascular (smoothelin-B) smooth muscle. Their expression is strongly associated with the contractile phenotype of smooth muscle cells. Analysis of mice lacking both smoothelins (Smtn-A/B−/− mice) previously revealed a critical role for smoothelin-A in intestinal smooth muscle contraction. Here, we report on the generation and cardiovascular phenotype of mice lacking only smoothelin-B (Smtn-B−/−). Methods and Results— Myograph studies revealed that the contractile capacity of the saphenous and femoral arteries was strongly reduced in Smtn-B−/− mice, regardless of the contractile agonist used to trigger contraction. Arteries from Smtn-A/B−/− compound mutant mice exhibited a similar contractile deficit. Smtn-B−/− arteries had a normal architecture and expressed normal levels of other smooth muscle cell–specific genes, including smooth muscle myosin heavy chain, α-smooth muscle actin, and smo...
41 citations
TL;DR: Gene Identification by Nonsense-mediated mRNA decay Inhibition (GINI) analysis has identified the ICR191-induced frameshift mutations in the TP53, smoothelin, Ras association (RalGDS/AF-6) domain family 6 (RASSF6) and other genes in the transformed MCF-10A cells.
Abstract: Widely accepted somatic mutation theory of carcinogenesis states that mutations in oncogenes and tumor suppressor genes in genomes of somatic cells is the cause of neoplastic transformation. Identifying frequent mutations in cancer cells suggests the involvement of mutant genes in carcinogenesis. To develop an in vitro model for the analysis of genetic alterations associated with breast carcinogenesis, we used random mutagenesis and selection of human non-tumorigenic immortalized breast epithelial cells MCF-10A in tissue-culture conditions that mimic tumor environment. Random mutations were generated in MCF-10A cells by cultivating them in a tissue-culture medium containing the frameshift-inducing agent ICR191. The first selective condition we used to transform MCF1-10A cells was cultivation in a medium containing mutagen at a concentration that allowed cell replication despite p53 protein accumulation induced by mutagen treatment. The second step of selection was either cell cultivation in a medium with reduced growth-factor supply or in a medium that mimics a hypoxia condition or growing in soft agar. Using mutagenesis and selection, we have generated several independently derived cultures with various degrees of transformation. Gene Identification by Nonsense-mediated mRNA decay Inhibition (GINI) analysis has identified the ICR191-induced frameshift mutations in the TP53, smoothelin, Ras association (RalGDS/AF-6) domain family 6 (RASSF6) and other genes in the transformed MCF-10A cells. The TP53 gene mutations resulting in the loss of protein expression had been found in all independently transformed MCF-10A cultures, which form large progressively growing tumors with sustained angiogenesis in nude mice. Identifying genes containing bi-allelic ICR191-induced frameshift mutations in the transformed MCF-10A cells generated by random mutagenesis and selection indicates putative breast-tumor suppressors. This can provide a model for studying the role of mutant genes in breast carcinogenesis.
32 citations
TL;DR: This work has determined the solution structure of the CH-domain of mouse SMTNL1 (SMTNL 1-CH; residues 346-459) and characterized the binding of apo-CaM to SMtNL1-CH through its IQ-motif by isothermal titration calorimetry and NMR chemical shift perturbation studies.
28 citations
TL;DR: To characterize a paracrine effect of prostatic epithelial cells in the presence or absence of oestradiol on the differentiation and proliferation of prostatics stromal cells, the objective is to establish a baseline for this study and establish an upper bound on this effect.
Abstract: OBJECTIVE
To characterize a paracrine effect of prostatic epithelial cells in the presence or absence of oestradiol on the differentiation and proliferation of prostatic stromal cells.
MATERIALS AND METHODS
Conditioned media (CM) collected from a prostatic epithelial cell line (BPH-1), which was pre-treated with different concentration of oestradiol, were added to cultures of primary prostatic stromal cells. The proliferation rates of stromal cells were determined using a tetrazolium assay. The mRNA level was analysed by real-time reverse transcription-polymerase chain reaction (RT-PCR), and the protein level of smooth muscle myosin heavy chain (SM-MHC), fibronectin and collagen IV were determined with Western blotting, enzyme- linked immunosorbent assay and radioimmunoassay, respectively. The expression of transforming growth factor β1 (TGFβ1) in the BPH-1 cell line was analysed.
RESULTS
The rate of proliferation of stromal cells increased when they were cultured with CM harvested from oestradiol-treated BPH-1 cells, but there was no remarkable change when they were cultured with CM from untreated cells. The level of smoothelin mRNA and SM-MHC protein increased after treatment with CM from BPH-1. The CM from BPH-1 with oestradiol stimulation was more effective in stimulating smoothelin mRNA and SM-MHC protein level. The protein level of collagen type IV, but not fibronectin, was up-regulated in the supernatants and cell extracts of CM-treated stromal cells. Oestradiol enhanced the expression and secretion of TGFβ1 in BPH-1 cells. TGFβ1-neutralizing antibody abrogated the effect of BPH-1 CM on the synthesis of collagen IV and SM-MHC in stromal cells.
CONCLUSION
These results suggest that oestradiol-stimulated proliferation and differentiation of prostatic stromal cells could be regulated by factors secreted from prostatic epithelial cells.
TL;DR: It is concluded that PTX treatment induces less proliferation within the vessel wall early after angioplasty and increases late lumen size after angyplasty by a positive effect on vascular remodeling.
Abstract: Pentoxifylline (PTX) inhibits the effects of several cytokines and reduces injury-related collagen accumulation. The aim of the present study was to investigate the effect of PTX on the vascular response to injury. We treated rabbits with PTX (100 mg/kg/day) or placebo (saline) subcutaneously from 2 days before angioplasty of an iliac artery until euthanasia 7 or 28 days later. At 7 days after injury, PTX treatment was associated with a more differentiated (less proliferation, more smoothelin-positive) intimal smooth muscle cell phenotype. Furthermore, PTX reduced myofibroblast accumulation in adventitia. At 28 days after injury, PTX-treated rabbits had a 48.5% larger lumen area (P = 0.03) and a 28.1% larger area within the external elastic lamina (P = 0.04). There were no significant differences between PTX-treated rabbits and the placebo group with regard to neointima and media area. Angioplasty induced marked neoadventitial hyperplasia, which was reduced by 20.5% (P = 0.01) in the PTX-treated group. Finally, PTX reduced collagen density in all three arterial layers. We conclude that PTX treatment induces less proliferation within the vessel wall early after angioplasty and increases late lumen size after angioplasty by a positive effect on vascular remodeling.
Journal Article•
TL;DR: High cell density,obvious atypia, high karyokinesis,cystification,hemorrage or necrosis in few cases were easy to be found (the character of malignant gastrointestinal stromal tumors).
Abstract: Objective To investigate the clinicopathologic and electron microscopic features of intra-abdominal inflammatory myofibroblastic tumor (IAIMT),Non-Abdominal inflammatory myofibroblastic tumor(NAIMT),Abdominal fibromatosis(AF),Gastrointestinal stromal tumor(GIST)and to discuss their differential diagnosis.Methods IAIMT,NAIMT,AF and GIST were observed by light microscopy,immunohistochemistry and electron microscopy.Results Objective To investigate the clinicopathologic and electron microscopic features of intra-abdominal inflammatory myofibroblastic tumor (IAIMT),Non-Abdominal inflammatory myofibroblastic tumor(NAIMT),Abdominal fibromatosis(AF),Gastrointestinal stromal tumor(GIST)and to discuss their differential diagnosis.Methods IAIMT,NAIMT,AF and GIST were observed by light microscopy,immunohistochemistry and electron microscopy.Results The clinical manifestations of IAIMT,NAIMT,AF and GIST were similar: Except the patients with NAIMT,the presenting symptoms in majority cases were abdominal mass,vague abdominal pain.The masses were either solitary or multiple and adherent to the surroundingtissues.Incomplete resection of the lesions can result in recurrence of the tumor and even death of the patients.There were similar characters in pathologic morphology between IAIMT and NAIMT,which composed of myofibroblasts,fibroblasts and a large number of inflammatory cells.The tumors had no capsules and clear boundary.There were three basic histological pantterns of IAIMT and NAIMT:①myxoid-vascular pattern,② compact spindle cell pattern,③hypocellular-fibroscar pattern.Calcification,ossification and necrosis were present in few cases.The masses of AF were composed of spindle and corpulent shaped cells,separated by a variable amount of collagenous tissue.GIST were composed of spindle cells and epithelioid cells.The vacuoles appeared in the cytoplasm of the neoplastic cells.The spindle cells were arranged in plexiform or bunchiness,Among them there are some cell nests which were the character of epithelial cells.High cell density,obvious atypia,high karyokinesis(≥5/50HPF),cystification,hemorrage or necrosis in few cases were easy to be found (the character of malignant gastrointestinal stromal tumors).In immunohistochemical analyses,all of IAIMT and NAIMT showed diffused strong positive for vimentin(15/15),SMA(15/15) and MSA(15/15),focal positive for ALK(7/15),Calponin(6/15),Desmin (5/15),CK(5/15),and negative for S-100,CD34,CD117,Caldesmon,β-catenin,Smoothelin,Pgp9.5 and MBP;AF showed diffuse strong positive for Vimentin(5/5),SMA(5/5),MSA(5/5),β-catenin(5/5),focal positive for S-100(1/5),Caponin(2/5),Desmin(1/5)and CK(1/5);GIST showed diffuse strong positive for Vimentin(5/5),CD117(5/5),focal positive for CD34(4/5)SMA(2/5)、MSA(2/5)、S-100(1/5),Calponin(3/5),Caldesmon(3/5),Desmin(1/5),Smoothelin(3/5),Pgp9.5(1/5)and MBP(1/5).The characteristic expressions of ALK,β-catenin,CD117 and CD34 were detected in IMT(IAIMTNAIMT),AF and GIST respectively.Moreover,Caldesmon and Smoothelin were both focal positive in GIST with smooth muscle-differentiation,but negative in tumors with myofibroblastic differentiation.Electron microscopically,IAIMT,NAMIT and AF all showed the characteristic ultrastructure of myofibroblasts and fibroblasts,but a large number of inflammatory cells and a amount of extracellular collagen appear in IMT and AF.Electronic microscopy of GIST show the presence of interdigitating cell processes,in some areas,synapse-like structure and numerous desmosome-like junctions as well as a few gap junctions and small round neurosecretory granules.There were also abundant intermediate filaments and thin filaments of actin-type with longitudina condensations (dense bodies) and a spot of undifferentiated mesenchymal cells.Conclusion A certain specific clinical characteristics,pathologic morphology,immunohistochemistry and ultrastructural change appear in IAIMT,NAIMT,AF and GIST.Among them,pathologic morphology and immunohistochemical phenotype are two important means to diagnose and differentiate IMT(IAIMTNAIMT) from AF and GIST.
Journal Article•
TL;DR: LBX reduces the mRNA expressions of TGF-beta1 and Smoothelin in human prostatic stromal cells and can be used in the treatment of BPH.
Abstract: OBJECTIVE To investigate the effects of the Chinese herbal medicine of Longbixiao (LBX) Capsule on the expressions of TGF-beta1 and Smoothelin in human prostatic stromal cells cultured in vitro. METHODS Blood serum medicated with LBX was incubated with the stromal cells isolated from men with benign prostatic hyperplasia (BPH) and cultured in vitro. The mRNA expression levels of TGF-beta1 and Smoothelin were detected by real-time RT-PCR and other relevant techniques. RESULTS In the high and low concentration groups, the gene relative expressions of TGF-beta1 were (0.158 +/- 0.020) and (0.169 +/- 0.020) , while those of Smoothelin were (0.035 +/- 0.007) and (0.036 +/- 0.007) respectively, both significantly decreased in comparison with the control group(P < 0.01). CONCLUSION LBX reduces the mRNA expressions of TGF-beta1 and Smoothelin in human prostatic stromal cells and can be used in the treatment of BPH.