Endoglin, a homodimeric transmembrane glycoprotein, is a part of the transforming growth factor-beta (TGF-beta) receptor cascade. It has been demonstrated that endoglin can affect TGF-beta signaling and eNOS expression by affecting SMAD proteins in vitro. We planned to go one step forward and evaluate whether endoglin is co-expressed with SMAD2, phosphorylated SMAD2/3 protein and eNOS in endothelium of normocholesterolemic C57BL/6J mice, and in advanced atherosclerotic lesions in hypercholesterolemic apoE/LDLr-deficient mice by means of fluorescence immunohistochemistry. Female C57BL/6J mice were fed with a chow diet (standard laboratory diet) for 12 weeks after weaning (at the age of 4 weeks). Two-month-old female apoE/LDLr-deficient mice were fed the western type diet (atherogenic diet) containing 21% fat (11% saturated fat) and 0.15% cholesterol for 2 months. Immunohistochemical analysis of endoglin, SMAD2, phosphorylated SMAD2/3 and eNOS expression was performed in mice aortic sinus. Immunohistochemical analysis showed the expression of endoglin in intact endothelium in both C57BL/6J and apoE/LDLr-deficient mice and in endothelium covering the atherosclerotic lesion in apoE/LDLr-deficient mice. Fluorescence immunohistochemistry revealed co-expression of endoglin with SMAD2, phosphorylated SMAD2/3 and eNOS in intact aortic endothelium in C57BL/6J mice. Moreover, strong co-localization of endoglin, SMAD2, phosphorylated SMAD2/3 and eNOS was also detected in endothelium covering atherosclerotic lesions in apoE/LDLr-deficient mice. In conclusion, we suggest that endoglin, SMAD2, phosphorylated SMAD2/3 and eNOS may be important in vessel endothelium homeostasis underlying their role in atherogenesis.

• MeSH
• aorta cytologie   MeSH
• apolipoproteiny E genetika   nedostatek   MeSH
• cévní endotel metabolismus   MeSH
• dieta aterogenní MeSH
• fluorescenční protilátková technika přímá MeSH
• fosforylace MeSH
• hypercholesterolemie metabolismus   MeSH
• imunohistochemie MeSH
• intracelulární signální peptidy a proteiny genetika   metabolismus   MeSH
• myši inbrední C57BL MeSH
• myši MeSH
• protein Smad2 genetika   metabolismus   MeSH
• protein Smad3 genetika   metabolismus   MeSH
• synthasa oxidu dusnatého, typ III metabolismus   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• práce podpořená grantem MeSH

AIM: Endoglin is a homodimeric transmembrane glycoprotein that has been demonstrated to affect transforming growth factor beta (TGF-beta) signaling and endothelial nitric oxide synthase (eNOS) expression by affecting SMAD proteins in vitro. Thus, in this study we stepped forward to elucidate whether endoglin is co-expressed with SMAD2, phosphorylated SMAD2/3 proteins and eNOS in vivo in atherosclerotic lesions in ApoE/LDLR double knockout mice. In addition, we sought whether endoglin expression as well as the expression of SMAD2, phosphorylated SMAD2/3 and eNOS is affected by atorvastatin treatment. METHODS: Two-month-old female ApoE/LDLR double knockout mice were divided into two groups. The control group was fed with the western type diet whereas in the atorvastatin group, atorvastatin at dose 100 mg/kg per day was added to the same diet. Immunohistochemical and western blot analysis of endoglin, SMAD2, phosphorylated SMAD2/3 and eNOS expressions in aorta were performed. RESULTS: The biochemical analysis showed that administration of atorvastatin significantly decreased level of total cholesterol, VLDL, LDL, TAG, and significantly increased level of HDL cholesterol. Fluorescence immunohistochemistry showed endoglin co-expression with SMAD2, phosphorylated SMAD2/3 and eNOS in aortic endothelium covering atherosclerotic lesions in both control and atorvastatin treated mice. Western blot analysis demonstrated that atorvastatin significantly increased expression of endoglin, SMAD2, phosphorylated SMAD2/3, and eNOS in mice aorta. CONCLUSION: These findings suggest, that endoglin might be interesting marker of endothelial dysfunction and/or atherogenesis which is upregulated by statins implicating potential beneficial role of endoglin and its pathway in atherosclerosis.

• MeSH
• anticholesteremika farmakologie   MeSH
• aorta chemie   MeSH
• apolipoproteiny E genetika   MeSH
• ateroskleróza metabolismus   MeSH
• cévní endotel chemie   MeSH
• fosforylace MeSH
• imunohistochemie MeSH
• intracelulární signální peptidy a proteiny analýza   MeSH
• kyseliny heptylové farmakologie   MeSH
• myši knockoutované MeSH
• myši MeSH
• protein Smad2 analýza   MeSH
• protein Smad3 analýza   MeSH
• pyrroly farmakologie   MeSH
• receptory LDL genetika   MeSH
• synthasa oxidu dusnatého, typ III analýza   MeSH
• western blotting MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Cardiac fibrosis is a final common pathology in inherited and acquired heart diseases that causes cardiac electrical and pump failure. Here, we use systems genetics to identify a pro-fibrotic gene network in the diseased heart and show that this network is regulated by the E3 ubiquitin ligase WWP2, specifically by the WWP2-N terminal isoform. Importantly, the WWP2-regulated pro-fibrotic gene network is conserved across different cardiac diseases characterized by fibrosis: human and murine dilated cardiomyopathy and repaired tetralogy of Fallot. Transgenic mice lacking the N-terminal region of the WWP2 protein show improved cardiac function and reduced myocardial fibrosis in response to pressure overload or myocardial infarction. In primary cardiac fibroblasts, WWP2 positively regulates the expression of pro-fibrotic markers and extracellular matrix genes. TGFβ1 stimulation promotes nuclear translocation of the WWP2 isoforms containing the N-terminal region and their interaction with SMAD2. WWP2 mediates the TGFβ1-induced nucleocytoplasmic shuttling and transcriptional activity of SMAD2.

• MeSH
• dospělí MeSH
• extracelulární matrix - proteiny metabolismus   MeSH
• fibróza genetika   metabolismus   MeSH
• genetická predispozice k nemoci * genetika   MeSH
• genové regulační sítě * MeSH
• kardiomyopatie genetika   metabolismus   MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• myši transgenní MeSH
• myši MeSH
• nemoci srdce genetika   metabolismus   MeSH
• protein - isoformy MeSH
• protein Smad2 genetika   metabolismus   MeSH
• regulace genové exprese MeSH
• senioři MeSH
• transformující růstový faktor beta metabolismus   MeSH
• ubikvitinligasy genetika   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• myši MeSH
• senioři MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Lower respiratory tract infection due to Pseudomonas aeruginosa has become increasingly challenging, resulting in a worse morbidity and mortality. Airway remodeling is a common phenomenon in this process, to which epithelial-mesenchymal transition (EMT) may contribute as an important promoter. Previous studies showed that epithelium-specific integrin αvβ6-mediated EMT was involved in pulmonary fibrosis via transforming growth factor-β1 (TGF-β1) signaling, but whether integrin αvβ6 plays a role in the P. aeruginosa-associated airway remodeling remains unknown. BEAS-2B cells were incubated with lipopolysaccharide (LPS) from P. aeruginosa in the presence or the absence of integrin αvβ6-blocking antibodies. Morphologic changes were observed by an inverted microscopy. The EMT markers were detected using Western blotting and immunofluorescence. The activation of TGF-β1-Smad2/3 signaling pathway was assessed. Furthermore, matrix metalloproteinase (MMP)-2 and -9 in the medium were measured using ELISA. P. aeruginosa's LPS decreased the expression of the epithelial marker E-cadherin and promoted the mesenchymal markers, vimentin and α-smooth muscle actin in BEAS-2B cells. The expression of integrin αvβ6 was significantly increased during EMT process. Blocking integrin αvβ6 could attenuate P. aeruginosa's LPS-induced EMT markers' expression via TGF-β1-Smad2/3 signaling pathway. Furthermore, blocking integrin αvβ6 could prevent morphologic changes and oversecretion of MMP-2 and -9. Integrin αvβ6 mediates epithelial-mesenchymal transition in human bronchial epithelial cells induced by lipopolysaccharides of P. aeruginosa via TGF-β1-Smad2/3 signaling pathway and might be a promising therapeutic target for P. aeruginosa-associated airway remodeling.

• MeSH
• antigeny nádorové genetika   metabolismus   MeSH
• epitelo-mezenchymální tranzice * MeSH
• epitelové buňky cytologie   účinky léků   metabolismus   MeSH
• integriny genetika   metabolismus   MeSH
• lidé MeSH
• lipopolysacharidy metabolismus   MeSH
• matrixové metaloproteinasy genetika   metabolismus   MeSH
• protein Smad2 genetika   metabolismus   MeSH
• protein Smad3 genetika   metabolismus   MeSH
• pseudomonádové infekce genetika   metabolismus   mikrobiologie   patofyziologie   MeSH
• Pseudomonas aeruginosa metabolismus   MeSH
• signální transdukce MeSH
• transformující růstový faktor beta1 genetika   metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

OBJECTIVE: GDF11 is a member of the TGF-β superfamily that was recently implicated as potential "rejuvenating" factor, which can ameliorate metabolic disorders. The main objective of the presented study was to closely characterize the role of GDF11 signaling in the glucose homeostasis and in the differentiation of white adipose tissue. METHODS: We performed microscopy imaging, biochemical and transcriptomic analyses of adipose tissues of 9 weeks old ob/ob mice and murine and human pre-adipocyte cell lines. RESULTS: Our in vivo experiments employing GDF11 treatment in ob/ob mice showed improved glucose/insulin homeostasis, decreased weight gain and white adipocyte size. Furthermore, GDF11 treatment inhibited adipogenesis in pre-adipocytes by ALK5-SMAD2/3 activation in cooperation with the WNT/β-catenin pathway, whose inhibition resulted in adipogenic differentiation. Lastly, we observed significantly elevated levels of the adipokine hormone adiponectin and increased glucose uptake by mature adipocytes upon GDF11 exposure. CONCLUSION: We show evidence that link GDF11 to adipogenic differentiation, glucose, and insulin homeostasis, which are pointing towards potential beneficial effects of GDF11-based "anti-obesity" therapy.

• MeSH
• adipogeneze * MeSH
• adiponektin metabolismus   MeSH
• beta-katenin * metabolismus   MeSH
• buněčná diferenciace fyziologie   MeSH
• glukosa metabolismus   MeSH
• inzulin metabolismus   MeSH
• kostní morfogenetické proteiny metabolismus   MeSH
• lidé MeSH
• myši MeSH
• protein Smad2 MeSH
• protein Smad3 MeSH
• receptory regulované proteiny Smad MeSH
• růstové diferenciační faktory metabolismus   MeSH
• signální dráha Wnt MeSH
• TGF-beta receptor I. typu MeSH
• transformující růstový faktor beta metabolismus   MeSH
• tukové buňky metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Gluteal muscle contracture (GMC) is a chronic fibrotic disease of gluteal muscles due to multiple etiologies. Emilin 1 plays a determinant role in fibers formation, but its role in the progression of GMC remains unclear. The present study was aimed to search for the predictive role and regulatory mechanism of Emilin 1 on GMC. Here, Protein and mRNA expression of Emilin 1 were decreased in GMC tissues compared to normal muscle tissues. Using the anslysis of target prediction, Emilin 1 was observed to be a potential downstream sponge of miR-491-5p. In comparison to Emilin 1, miR-491-5p showed a aberrant elevation in GMC tissues, which was further proven to have a negative correlation with Emilin 1. The direct binding of miR-491-5p to Emilin 1 mRNA was confirmed by luciferase reporter gene assay, and miR-491-5p mimics inhibited, while miR-491-5p inhibitor promoted the protein expression and secretion of Emilin 1 in contraction bands (CB) fibroblasts. Additionally, miR-491-5p mimics promoted the expression of cyclin-dependent kinase 2 and cyclin D1 and the proliferation of CB fibroblasts, which could be reversed by Emilin 1 overexpression. Mechanistically, miR-491-5p mimics possibly activated transforming growth factor beta1 (TGF-beta1)/Smad3 signal cascade via binding to 3'-untranslated region of Emilin 1 mRNA, thereby promoting the progression of fibrosis of CB fibroblasts. Collectively, miR-491-5p inhibited Emilin 1 expression, and subsequently promoted CB fibroblasts proliferation and fibrosis via activating TGF-beta1/Smad3 signal axis. MiR-491-5p might be a potentially effective biomarker for predicting GMC, providing a novel therapeutic strategy for GMC.

• MeSH
• fibroblasty metabolismus   MeSH
• fibróza MeSH
• kontraktura * patologie   MeSH
• kosterní svaly metabolismus   MeSH
• lidé MeSH
• membránové glykoproteiny MeSH
• messenger RNA metabolismus   MeSH
• mikro RNA * genetika   metabolismus   MeSH
• proliferace buněk MeSH
• protein Smad2 metabolismus   MeSH
• transformující růstový faktor beta1 genetika   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Several lines of evidence suggest that in mice the activation of SMAD2/3 signaling by oocyte secreted factors, together with epidermal growth factor receptor (EGFR) activation, is essential to induce cumulus expansion. Here we show that inhibition of EGFR kinase in follicle stimulating hormone (FSH)-stimulated porcine oocyte-cumulus cell complex (OCCs) strongly decreases hyaluronan (HA) synthesis and its retention in the matrix, as well as progesterone synthesis. Although porcine cumulus cells undergo expansion independently of oocytes, we use biochemical and gene expression analyses to show that they do require activation of SMAD2/3 for optimal stimulation of HA synthesis and proteins involved in the organization of this polymer in the expanded matrix. Furthermore, FSH-induced progesterone synthesis by porcine cumulus cells was increased by blocking SMAD2/3 activation. In conclusion, these results support the hypothesis that an FSH-EGF autocrine loop is active in porcine OCCs, and provide the first evidence that the SMAD2/3 signaling pathway is induced by paracrine/autocrine factors in porcine cumulus cells and is involved in the control of both cumulus expansion and steroidogenesis.

• MeSH
• benzamidy farmakologie   MeSH
• C-reaktivní protein metabolismus   MeSH
• chinazoliny farmakologie   MeSH
• dioxoly farmakologie   MeSH
• epidermální růstový faktor metabolismus   MeSH
• erbB receptory antagonisté a inhibitory   metabolismus   MeSH
• folikuly stimulující hormon metabolismus   MeSH
• glukuronosyltransferasa antagonisté a inhibitory   metabolismus   MeSH
• isochinoliny farmakologie   MeSH
• kumulární buňky metabolismus   MeSH
• kyselina hyaluronová biosyntéza   MeSH
• meióza účinky léků   MeSH
• molekuly buněčné adheze antagonisté a inhibitory   metabolismus   MeSH
• myši MeSH
• oocyty enzymologie   fyziologie   MeSH
• prasata MeSH
• progesteron biosyntéza   MeSH
• protein Smad2 antagonisté a inhibitory   metabolismus   MeSH
• protein Smad3 antagonisté a inhibitory   metabolismus   MeSH
• pyridiny farmakologie   MeSH
• pyrroly farmakologie   MeSH
• sérový amyloidový protein metabolismus   MeSH
• signální transdukce účinky léků   MeSH
• tyrphostiny farmakologie   MeSH
• vývojová regulace genové exprese účinky léků   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The incidence of cutaneous squamous cell carcinoma (cSCC) is rising. Whilst the majority are cured surgically, aggressive metastatic cSCC carry a poor prognosis. Inactivating mutations in transforming growth factor beta (TGF-β) receptors have been identified amongst genetic drivers of sporadic tumours and murine models of cSCC, suggesting a tumour suppressor function for TGF-β in normal skin. However, paradoxically, TGF-β acts as a tumour promoter in some murine model systems. Few studies have analysed the role of TGF-β/activin signalling in human normal skin, hyper-proliferative skin disorders and cSCC. Antibodies recognising phospho-SMAD proteins which are activated during canonical TGF-β/activin signalling were validated for use in immunohistochemistry. A tissue microarray comprising FFPE lesional and perilesional tissue from human primary invasive cSCC (n=238), cSCCin-situ(n=2) and keratocanthoma (n=9) were analysed in comparison with tissues from normal human scalp (n=10). Phosphorylated SMAD2 and SMAD3 were detected in normal interfollicular epidermal keratinocytes and were also highly localised to inner root sheath, matrix cells and Keratin 15 positive cells. Lesional cSCC tissue had significantly reduced activated SMAD2/3 compared to perilesional tissue, consistent with a tumour suppressor role for SMAD2/3 activators in cSCC. Increased cSCC tumour thickness inversely correlated with the presence of phospho-SMADs in tumour tissue suggesting that a reduction in canonical TGF-β/activin signalling may be associated with disease progression.

• Publikační typ
• časopisecké články MeSH

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

• Publikační typ
• tisková chyba MeSH

The hearts of spontaneously hypertensive rats (SHR) are prone to malignant arrhythmias, mainly due to disorders of electrical coupling protein Cx43 and the extracellular matrix. Cold acclimation may induce cardio-protection, but the underlying mechanisms remain to be elucidated. We aimed to explore whether the adaptation of 9-month-old hairless SHRM to cold impacts the fundamental cardiac pro-arrhythmia factors, as well as the response to the thyroid status. There were no significant differences in the registered biometric, redox and blood lipids parameters between hairless (SHRM) and wild type SHR. Prominent findings revealed that myocardial Cx43 and its variant phosphorylated at serine 368 were increased, while an abnormal cardiomyocyte Cx43 distribution was attenuated in hairless SHRM vs. wild type SHR males and females. Moreover, the level of β-catenin, ensuring mechanoelectrical coupling, was increased as well, while extracellular matrix collagen-1 and hydroxyproline were lower and the TGF-β1 and SMAD2/3 pathway was suppressed in hairless SHRM males compared to the wild type strain. Of interest, the extracellular matrix remodeling was less pronounced in females of both hypertensive strains. There were no apparent differences in response to the hypothyroid or hyperthyroid status between SHR strains concerning the examined markers. Our findings imply that hairless SHRM benefit from cold acclimation due to the attenuation of the hypertension-induced adverse downregulation of Cx43 and upregulation of extracellular matrix proteins.

• Publikační typ
• časopisecké články MeSH