SIRT1-mediated deacetylation of NF-kB inhibits the MLCK/MLC2 pathway and the expression of ET-1, thus alleviating the development of coronary artery spasm

摘要

Coronary artery spasm (CAS) is an intense vasoconstriction of coronary arteries that causes total or subtotal vessel occlusion. The cardioprotective effect of sirtuin-1 (SIRT1) has been extensively highlighted in coronary artery diseases. The aims within this study include the investigation of the molecular mechanism by which SIRT1 alleviates CAS. SIRT1 expression was first determined by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and Western blot analysis in an endothelin-1 (ET-1)-induced rat CAS model. Interaction among SIRT1, nuclear factor-kappaB (NF-kB), myosin light chain kinase/myosin light chain-2 (MLCK/MLC2), and ET-1 was analyzed using luciferase reporter assay, RT-qPCR, and Western blot analysis. After ectopic expression and depletion experiments in vascular smooth muscle cells (VSMCs), contraction and proliferation of VSMCs and expression of contraction-related proteins (a-SMA, calponin, and SM22a) were measured by collagen gel contraction, 5-ethynyl-2'-deoxyuri-dine (EdU) assay, RT-qPCR, and Western blot analysis. The obtained results showed that SIRT1 expression was reduced in rat CAS models. However, overexpression of SIRT1 inhibited the contraction and proliferation of VSMCs in vitro. Mechanistic investigation indicated that SIRT1 inhibited NF-kB expression through deacetylation. Moreover, NF-kB could activate the MLCK/MLC2 pathway and upregulate ET-1 expression by binding to their promoter regions, thus inducing VSMC contraction and proliferation in vitro. In vivo experimental results also revealed that SIRT1 alleviated CAS through regulation of the NF-kB/MLCK/MLC2/ET-1 signaling axis. Collectively, our data suggested that SIRT1 could mediate the deacetylation of NF-kB, disrupt the MLCK/MLC2 pathway, and inhibit the expression of ET-1 to relieve CAS, providing a theoretical basis for the prospect of CAS treatment and prevention.