scholarly journals Rapamycin activates TGF receptor independently of its ligand: implications for endothelial dysfunction

2018 ◽  
Vol 132 (4) ◽  
pp. 437-447 ◽  
Author(s):  
Ayumi A. Miyakawa ◽  
Thais Girão-Silva ◽  
Jose E. Krieger ◽  
Elazer R. Edelman
Keyword(s):  
Cell Proliferation ◽  
Kinase Inhibitor ◽  
Umbilical Vein ◽  
Type I ◽  
Mesenchymal Transition ◽  
Smad Phosphorylation ◽  
Endothelial To Mesenchymal Transition ◽  
Umbilical Vein Endothelial Cells ◽  
Pai 1

Rapamycin, the macrolide immunosuppressant and active pharmaceutic in drug-eluting stents (DES), has a well-recognized antiproliferative action that involves inhibition of the mTOR pathway after binding to the cytosolic protein FKBP12. TGF receptor-type I (TGFRI) spontaneous activation is inhibited by the association with FKBP12. We hypothesized that rapamycin, in addition to inhibition of mTOR signaling, activates TGFRI independent of TGFβ. Human umbilical vein endothelial cells (HUVECs) were treated with rapamycin (10 nmol/l) and/or TGFβ RI kinase inhibitor (TGFRIi, 100 nmol/l) for 24 h. Rapamycin induced SMAD phosphorylation (SMAD1, SMAD2, and SMAD5) and PAI-1 up-regulation, which was specifically abrogated by SMAD2 knockdown. TGFRIi efficiently blocked phosphorylation of SMAD2, but not SMAD1/5. Interestingly, the inhibitor did not alter cell proliferation arrest induced by rapamycin. Active TGFβ secretion was not affected by the treatment. Neutralizing TGFβ experiments did not influence SMAD2 phosphorylation or PAI-1 expression indicating that activation of this pathway is independent of the ligand. In addition, rapamycin induction of endothelial-to-mesenchymal transition (EndMT) was potentiated by IL-1β and efficiently blocked by TGFRIi. In vivo, the prothrombogenic effects of rapamycin and up-regulation of PAI-1 in murine carotid arteries were reduced by TGFRIi treatment. In conclusion, we provide evidence that rapamycin activates TGF receptor independent of its ligand TGFβ, in concert with promotion of PAI-1 expression and changes in endothelial phenotype. These undesirable effects, the prothrombogenic state, and activation of EndMT are SMAD2-dependent and independent of the therapeutic rapamycin-induced cell proliferation arrest.

scholarly journals Dasatinib increases endothelial permeability leading to pleural effusion

2018 ◽  
Vol 51 (1) ◽  
pp. 1701096 ◽  
Author(s):  
Carole Phan ◽  
Etienne-Marie Jutant ◽  
Ly Tu ◽  
Raphaël Thuillet ◽  
Andrei Seferian ◽  
...  
Keyword(s):  
Pleural Effusion ◽  
Kinase Inhibitor ◽  
Umbilical Vein ◽  
Endothelial Permeability ◽  
Cell Junctions ◽  
Dependent Manner ◽  
Vascular Endothelial ◽  
Umbilical Vein Endothelial Cells

Pleural effusion is a frequent side-effect of dasatinib, a second-generation tyrosine kinase inhibitor used in the treatment of chronic myelogenous leukaemia. However, the underlying mechanisms remain unknown. We hypothesised that dasatinib alters endothelial integrity, resulting in increased pulmonary vascular endothelial permeability and pleural effusion.To test this, we established the first animal model of dasatinib-related pleural effusion, by treating rats with a daily regimen of high doses of dasatinib (10 mg·kg−1·day−1 for 8 weeks).Pleural ultrasonography revealed that rats chronically treated with dasatinib developed pleural effusion after 5 weeks. Consistent with these in vivo observations, dasatinib led to a rapid and reversible increase in paracellular permeability of human pulmonary endothelial cell monolayers as reflected by increased macromolecule passage, loss of vascular endothelial cadherin and zonula occludens-1 from cell–cell junctions, and the development of actin stress fibres. These results were replicated using human umbilical vein endothelial cells and confirmed by decreased endothelial resistance. Interestingly, we demonstrated that this increased endothelial permeability is a reactive oxygen species (ROS)-dependent mechanism in vitro and in vivo using a cotreatment with an antioxidant agent, N-acetylcysteine.This study shows that dasatinib alters pulmonary endothelial permeability in a ROS-dependent manner in vitro and in vivo leading to pleural effusion.

The role of miR-328 in high glucose-induced endothelial-to-mesenchymal transition in human umbilical vein endothelial cells

Life Sciences ◽  
2018 ◽  
Vol 207 ◽  
pp. 110-116 ◽  
Author(s):  
Yunxiao Chen ◽  
Qin Yang ◽  
Yuliang Zhan ◽  
Junsong Ke ◽  
Ping Lv ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
High Glucose ◽  
Umbilical Vein ◽  
Mesenchymal Transition ◽  
Human Umbilical Vein ◽  
Endothelial To Mesenchymal Transition ◽  
Umbilical Vein Endothelial Cells

scholarly journals Calcitriol Attenuates Doxorubicin-Induced Cardiac Dysfunction and Inhibits Endothelial-to-Mesenchymal Transition in Mice

Cells ◽  
2019 ◽  
Vol 8 (8) ◽  
pp. 865 ◽  
Author(s):  
Tsai ◽  
Lin ◽  
Hang ◽  
Chen
Keyword(s):  
Myocardial Fibrosis ◽  
Cardiac Fibrosis ◽  
Transforming Growth Factor ◽  
Cardiac Fibroblasts ◽  
Umbilical Vein ◽  
Active Form ◽  
Mesenchymal Transition ◽  
Endothelial To Mesenchymal Transition ◽  
Umbilical Vein Endothelial Cells

Doxorubicin (Dox) is an effective anti-neoplasm drug, but its cardiac toxicity limits its clinical use. Endothelial-to-mesenchymal transition (EndMT) has been found to be involved in the process of heart failure. It is unclear whether EndMT contributes to Dox-induced cardiomyopathy (DoIC). Calcitriol, an active form Vitamin D3, blocks the growth of cancer cells by inhibiting the Smad pathway. To investigate the effect of calcitriol via inhibiting EndMT in DoIC, C57BL/6 mice and endothelial-specific labeled mice were intraperitoneally administered Dox twice weekly for 4 weeks (32 mg/kg cumulative dose) and were subsequently treated with or without calcitriol for 12 weeks. Echocardiography revealed diastolic dysfunction at 13 weeks following the first Dox treatment, accompanied by increased myocardial fibrosis and up-regulated pro-fibrotic proteins. Calcitriol attenuated Dox-induced myocardial fibrosis, down-regulated pro-fibrotic proteins and improved diastolic function. Endothelial fate tracing revealed that EndMT-derived cells contributed to Dox-induced cardiac fibrosis. In vitro, human umbilical vein endothelial cells and mouse cardiac fibroblasts were treated with Transforming growth factor (TGF)-β with or without calcitriol. Morphological, immunofluorescence staining, and Western blot analyses revealed that TGF-β-induced EndMT and fibroblast-to-myofibroblast transition (FMT) were attenuated by calcitriol by the inhibition of the Smad2 pathway. Collectively, calcitriol attenuated DoIC through the inhibition of the EndMT and FMT processes.

Autophagy mediates 2-methoxyestradiol-inhibited scleroderma collagen synthesis and endothelial-to-mesenchymal transition induced by hypoxia

Rheumatology ◽  
2019 ◽  
Vol 58 (11) ◽  
pp. 1966-1975 ◽  
Author(s):  
Chaofan Liu ◽  
Xing Zhou ◽  
Jinghao Lu ◽  
Lubing Zhu ◽  
Ming Li
Keyword(s):  
Collagen Synthesis ◽  
Umbilical Vein ◽  
Smooth Muscle Actin ◽  
Tissue Growth ◽  
Collagen I ◽  
Bafilomycin A1 ◽  
Mesenchymal Transition ◽  
Transmission Electron ◽  
Endothelial To Mesenchymal Transition ◽  
Umbilical Vein Endothelial Cells

Abstract Objectives To investigate whether autophagy mediates 2-methoxyestradiol (2-ME)-inhibited hypoxia-induced fibrosis and endothelial-to-mesenchymal transition (endoMT) in SSc. Methods Autophagy in the skin of SSc patients was assessed by transmission electron microscopy. SSc skin fibroblasts and human umbilical vein endothelial cells (HUVECs) were cultured under hypoxic (1% O2) conditions with 2-ME or autophagy inhibitor. Collagen I and connective tissue growth factor (CTGF) in fibroblasts and vascular endothelial (VE)-cadherin, CD31, vimentin and α-smooth muscle actin (α-SMA) in HUVECs were examined by western blotting. Autophagic markers were evaluated by confocal microscopy and immunofluorescence. Results SSc skins presented increased autolysosomes, LC3-II, collagen I and CTGF. Hypoxia-challenged fibroblasts and HUVECs formed more autophagosomes and autolysosomes, with increased LC3 and decreased P62. Meanwhile, hypoxia increased collagen I and CTGF in fibroblasts and increased vimentin and α-SMA but decreased VE-cadherin and CD31 in HUVECs. Bafilomycin A1 increased LC3-II and P62 in fibroblasts and HUVECs and decreased collagen I and CTGF in fibroblasts and vimentin and α-SMA in HUVECs, while upregulating VE-cadherin and CD31. 3-methyladenine decreased autophagy and fibrosis in fibroblasts and endothelial-to-mesenchymal transition in HUVECs. 2-ME-treated HUVECs showed more autophagosomes and fewer autolysosomes while 2-ME-treated fibroblasts showed fewer of both. Moreover, 2-ME decreased LC3-II and increased P62 in fibroblasts and increased both in HUVECs. Inhibition of autophagy by 2-ME showed the same effect with bafilomycin A1 on fibroblast collagen synthesis as well as endothelial and mesenchymal markers in HUVECs. Conclusion Autophagy mediated hypoxia-induced fibroblast collagen synthesis and endoMT in SSc, which could be reversed by 2-ME.

scholarly journals CXCR7 Inhibits Fibrosis via Wnt/β-Catenin Pathways during the Process of Angiogenesis in Human Umbilical Vein Endothelial Cells

2020 ◽  
Vol 2020 ◽  
pp. 1-10
Author(s):  
MinQian Shen ◽  
YiFan Feng ◽  
Jing Wang ◽  
YuanZhi Yuan ◽  
Fei Yuan
Keyword(s):  
Endothelial Cells ◽  
Umbilical Vein ◽  
Tube Formation ◽  
Erk Signaling ◽  
Cell Type ◽  
Mesenchymal Transition ◽  
Human Umbilical Vein ◽  
Endothelial To Mesenchymal Transition ◽  
Umbilical Vein Endothelial Cells ◽  
Tissue Origin

Although SDF-1/CXCR7 plays an important role in angiogenesis, the function and the pathway of the SDF-1/CXCR7 axis might depend on the cell type or tissue origin and not fully understood. In this study, we investigated the effect of CXCR7 in SDF-1-induced proliferation, migration, apoptosis, tube formation, and endothelial-to-mesenchymal transition (EndMT) of human umbilical vein endothelial cells (HUVECs), and the potential pathway of SDF-1/CXCR7. We confirmed that the silencing of CXCR7 inhibited the proliferation of HUVECs and contributed the apoptosis, while overexpressed CXCR7 increased SDF-1-induced HUVECs migration and tube formation. However, upregulated CXCR7 inhibited the expression of α-SMA, suggesting that CXCR7 might attenuate EndMT. In addition, overexpressed CXCR7 activated AKT and ERK signaling pathways but suppressed Wnt/β-catenin pathways in HUVECs. The inhibition of Wnt/β-catenin pathways decreased the expression of α-SMA. Altogether, these results suggest that CXCR7 might inhibit fibrosis via Wnt/β-catenin pathways during the process of angiogenesis.

scholarly journals MiR-200c-3p promotes ox-LDL-induced endothelial to mesenchymal transition in human umbilical vein endothelial cells through SMAD7/YAP pathway

2021 ◽  
Vol 71 (1) ◽  
Author(s):  
Yongzhong Mao ◽  
Ling Jiang
Keyword(s):  
Endothelial Cells ◽  
Umbilical Vein ◽  
Critical Role ◽  
Luciferase Reporter ◽  
Rhodamine Phalloidin ◽  
Mesenchymal Transition ◽  
Human Umbilical Vein ◽  
Endothelial To Mesenchymal Transition ◽  
Umbilical Vein Endothelial Cells

Abstract Background Endothelial to mesenchymal transition (EndMT) participates in the progression of atherosclerosis (AS). MiR-200c-3p has been implicated in EndMT. However, the functional role of miR-200c-3p in AS remains largely unknown. Here, we demonstrated the critical role of miR-200c-3p in regulating EndMT in AS. Methods ApoE−/− mice were fed with high-fat diet to establish AS mouse model, and human umbilical vein endothelial cells (HUVECs) were treated with oxidized low-density lipoprotein (ox-LDL) to mimic AS cell model. The expression of miR-200c-3p, SMAD7 and YAP in ApoE−/− mice and HUVECs was detected by quantitative real-time PCR. Rhodamine phalloidin staining and Western blot were performed to observe cell morphology and EndMT marker expression of HUVECs. Luciferase reporter assay and Co-Immunoprecipitation were performed to verify the relationship among miR-200c-3p, SMAD7, and YAP. Results MiR-200c-3p was highly expressed, and SMAD7 and YAP were down-regulated in the aortic tissues of ApoE−/− mice and ox-LDL-treated HUVECs. MiR-200c-3p overexpression promoted the transformation of ox-LDL-treated HUVECs from cobblestone-like epithelial phenotype to a spindle-like mesenchymal phenotype. Meanwhile, miR-200c-3p up-regulation repressed the expression of endothelial markers CD31 and vWF and promoted the expression of mesenchymal markers α-SMA and vimentin in the ox-LDL-treated HUVECs. MiR-200c-3p inhibited SMAD7 and YAP expression by interacting with 3′ untranslated region of SMAD7. Moreover, miR-200c-3p promoted EndMT in ox-LDL-treated HUVECs by inhibiting SMAD7/YAP pathway. Conclusion This work demonstrated that MiR-200c-3p promoted ox-LDL-induced EndMT in HUVECs through SMAD7/YAP pathway, which may be important for the onset of atherosclerosis.

Mechanistic studies on the role of TGF-β1 in angiogenesis through EndMT

Vascular ◽  
2020 ◽  
pp. 170853812095366
Author(s):  
Sheng-Jun Cao ◽  
Lei Hong ◽  
Xiao-Qiang Li
Keyword(s):  
Growth Factor ◽  
Transforming Growth Factor ◽  
Umbilical Vein ◽  
Transforming Growth Factor Β1 ◽  
Tube Formation ◽  
Mesenchymal Transition ◽  
Phosphorylated Akt ◽  
Endothelial To Mesenchymal Transition ◽  
Umbilical Vein Endothelial Cells ◽  
Tgf Β1

Objective This study aims to investigate the mechanism of transforming growth factor-β1 (TGF-β1) in promoting angiogenesis through endothelial-to-mesenchymal transition (EndMT). Methods The mesenchymal transition of human umbilical vein endothelial cells (HUVECs) was induced by TGF-β1. The angiogenesis, migration, and proliferation of HUVECs undergoing EndMT were examined by tube formation assay, scratch assay, Transwell assay, and CCK-8 assay. Results The outcomes revealed that EndMT promoted angiogenesis, migration, and proliferation of HUVECs and the secretion of the vascular endothelial growth factor (VEGF) of HUVECs. Phosphorylated AKT (p-AKT) increased in EndMT by inhibiting the mitigation of angiogenesis. Conclusion EndMT induces angiogenesis by promoting the secretion of VEGF, and p-AKT participates in this regulation.

scholarly journals Irradiated Human Umbilical Vein Endothelial Cells Undergo Endothelial-Mesenchymal Transition via the Snail/miR-199a-5p Axis to Promote the Differentiation of Fibroblasts into Myofibroblasts

2018 ◽  
Vol 2018 ◽  
pp. 1-10 ◽  
Author(s):  
Minxiao Yi ◽  
Bo Liu ◽  
Yang Tang ◽  
Fang Li ◽  
Wan Qin ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Umbilical Vein ◽  
Smooth Muscle Actin ◽  
Lung Fibroblasts ◽  
Type I ◽  
Mesenchymal Transition ◽  
Human Umbilical Vein ◽  
Radiation Induced ◽  
Umbilical Vein Endothelial Cells ◽  
Endothelial Mesenchymal Transition

Radiation induced pulmonary fibrosis (RIPF) is one of the major side effects of radiotherapy for lung cancer. Previous studies have shown that endothelial cells and activated myofibroblasts play a key role in RIPF. However, the interaction between irradiated endothelial cells and activation of myofibroblasts has not been reported. The aim of the present study was to examine whether irradiated endothelial cells would affect the differentiation of fibroblasts into myofibroblasts in the process of RIPF. In the current study, we used a coculture system that allowed direct contact between human fetal lung fibroblasts (MRC-5) and irradiated human umbilical vein endothelial cells (HUVECs). After 24 or 48 h, cells were sorted by flow cytometry. Radiation induced endothelial-mesenchymal transition (EndMT) by significantly increasing the expression of Snail and vimentin and reducing the expression of CD31 in HUVECs. In addition, irradiation of HUVECs induced the expression of collagen type I and α-smooth muscle actin (α-SMA) in MRC-5 cells. Further investigation indicated that irradiation of HUVECs induced the differentiation of fibroblasts into myofibroblasts through the Snail/miR-199a-5p axis. We conclude that irradiated endothelial cells undergo EndMT to promote differentiation of fibroblasts into myofibroblasts via the Snail/miR-199a-5p axis.

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