Expression and mechanism of miR-15a in the endothelial cell model of septic shock

LAN Qing; CHEN Hao; WANG Bin; ZHENG Xiangde; LIU Cheng; LI Chongpei

doi:10.13431/j.cnki.immunol.j.20180016

Abstract：To investigate the role of miR-15a in septic shock and the underlying mechanism,qRT-PCR was employed to quantify the expression level of miR-15a in healthy controls (n=25) and septic patients with (n=26) or without shock (n=34).MicroRNA-15a mimic was applied in the HUVECs culture to examine its effect on endogenous miR-15a and LPS-induced cell permeability.Direct targeting of miR-15a on myosin light chain kinase (MYLK) was assessed by luciferase assay,qRT-PCR and Western blotting were employed to quantify the expression level of MYLK in HUVECs after transfection with miR-15a.Data showed that the level of miR-15a was significantly up-regulated in septic patients,more importantly,miR-15a was significantly down-regulated in septic shock patients compared with septic patients without shock (P＜0.05).MiR-15a mimic upregulated endogenous miR-15a gene level,but inhibited LPS-induced permeability;MYLK was directly targeted by miR-15a.MiR-15a mimic significantly decreased the mRNA (P＜0.05) and protein levels of MYLK in HUVECs cells,while MiR-15a inhibitor significantly increased the mRNA (P＜0.05) and protein levels of MYLK in HUVECs cells.Taken together,Upregulating miR-15a protects HUVECs against permeability change and maintains endothelial barrier integrity,probably through down-regulating MYLK.The plasma levels of miRNA are altered in patients with severe sepsis complicated by shock,which may offer new insights into the mechanisms of endothelial dysfunction in sepsis.