Crosstalk between endothelial cell-specific calpain inhibition and the endothelial-mesenchymal transition via the HSP90/Akt signaling pathway - 11/02/20
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Graphical abstract |
Highlights |
• | Canonical TGF-β1 triggers Endothelial-Mesenchymal Transition by activating calpain. |
• | Endothelial cell calpain deletion ameliorates cardiac fibrosis and hypertrophy. |
• | Overexpression of calpastatin reverses Endothelial-Mesenchymal Transition in vitro. |
• | Calpain inhibition contributes to alleviating EndMT via blocking HSP90/Akt signaling. |
Abstract |
Hypothesis |
The role of non-cardiomyocytes in cardiac remodeling and fibrosis has not been totally understood until now. This study investigated if endothelial cell (EC)-specific calpain participates in myocardial endothelial injury via the endothelial- mesenchymal transition (EndMT) and in cardiac fibroblasts during cell proliferation, thereby contributing to cardiac fibrosis eventually.
Methods |
in vitro cultured mouse cardiac ECs were induced with transforming growth factor (TGF)-β1 (10 ng/ml) and calpain inhibitor III (20 μM) or Akt inhibitor (LY294002, 20 μM). Isolated cardiac fibroblasts were induced by TGF-β1 and an HSP90 inhibitor (17AAG, 20 μM), and EndMT were analysed. Capn4-knockout (KO) specific to ECs of mice was generated. We induced the pathological process mimicking cardiac hypertrophy and fibrosis in both Capn4-KO mice and their wild-type littermates. The histological analysis was used to measure cardiomyocyte size and collagen contained in the heart. The immunofluorescence analysis was performed to demonstrate that the ECs went through the EndMT, transforming mesenchymal cells into fibroblasts and myofibroblasts.
Results |
Capn4 deletion specific to ECs abrogated activity of both calpain 1 and calpain 2 in ECs, lowered the volume of cardiac collagen and cardiomyocytes size, and ameliorated myocardial dysfunction in the isoproterenol-treated cardiac fibrosis model. An ex vivo analysis of cardiomyocytes by Evans Blue staining revealed that isoproterenol increased cell death compared with the control, and Capn4-KO alleviated this result. Inhibiting calpain in cultured cardiac microvascular endothelial cells (MCECs) reversed the EndMT process, which was induced by TGF-β1. Overexpression of calpastatin decreased the pathological EndMT process, showing that the cultured MCECs have more mesenchymal markers, such as α-smooth muscle actin (SMA), and fewer endothelial markers, such as VE-cadherin. Activating calpain elevated phosphorylated Akt in mice cultured ECs, and inhibiting calpain decreased phosphorylated Akt. Upregulation of phosphorylated Akt by calpain promoted the EndMT, whereas inhibiting calpain switched on the protective mechanism during the EndMT via the heat shock protein (HSP)90/Akt signaling way in cultured ECs.
Conclusions |
This study demonstrated a vital role of calpain in ECs for inducing myocardiocyte hypertrophy, cell death and the EndMT via the HSP90/Akt signaling pathway, thereby promoting cardiac fibrosis. The results indicate that inhibiting ECs calpain is a novel therapeutic target to retard cardiac fibrosis and has positive effects on heart failure.
Le texte complet de cet article est disponible en PDF.Keywords : EC-calpain, Cardiac fibrosis, HSP90, p-Akt, VE-cadherin, α-SMA, Endothelial-mesenchymal transition
Plan
Vol 124
Article 109822- avril 2020 Retour au numéroBienvenue sur EM-consulte, la référence des professionnels de santé.
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