Regulation Of High Glucose/ox-LDL On Sortilin Expression In Liver Sinusoidal Endothelial Cells

Object:Both type 2 diabetes mellitus(T2DM)and non-alcoholic fatty liver disease(NAFLD)are closely associated with the elevated levels of low-density lipoprotein cholesterol(LDL-C)and its oxidized form(ox-LDL).Sortilin,participate in liver lipid metabolism by lysosomal degradation of sortilin protein.Recent studies demonstrated that high glucose/ox-LDL is critically involved in the LSECs sortilin expression and inflammatory cytokine secretion.This study aims to investigate the role of sortilin induced by high glucose and ox-LDL in LSECs in vitro and effect of inflammatory cytokine secretion.Methods:Extracted and isolated mouse primary LSECs.Anti-CD31 was used to identify LSECs.Control group(glucose concentration 5.5mmol/L),high glucose group(glucose concentration 25mmol/L),ox-LDL group(ox-LDL concentration100?g/ml),high glucose+ox-LDL group(glucose concentration 25mmol/L+ox-LDL concentration 100?g/ml)and LV-Sort1 group(glucose concentration 25mmol/L+ox-LDL concentration 100?g/ml+LV-S o r t 1 1×1 0 ~8 T U/m l)were established.Oil red O(ORO)staining observed lipid droplets accumulation in LSECs,the apoptosis rate of LSECs was determined by AnnexinV-FITC/PI double staining,immunofluorescence technique was used to detect the expression levels of sortilin,extraction of mRNA and protein from each group after culture 24h,real-time quantitative PCR(RT-PCR)and Western blotting were used to detect the levels of genes and proteins in each group of LSECs.SPSS 21.0 software was used for statistical analysis.Results:We first found that sortilin was expressed in LSECs by anti-CD31staining,ORO and AnnexinV-FITC/PI staining showed that high glucose and ox-LDL increased lipid droplets of accumulation and the rate of apoptosis of cells.We then provided evidence demonstrating that LSECs sortilin protein was significantly decreased in cultured with high glucose and ox-LDL when assessed by immunofluorescence confocal microscope,western blotting and RT-PCR.In addition,overexpression of sortilin in LSECs via a lentiviral-mediated gene infection increased sortilin expression,ameliorated cell apoptosis rate and decreased IL-6 and TNF-?release.Conclusions:sortilin is expressed in LSECs,a synergistic effect on the accumulation of lipid droplets and induction of apoptosis and decrease sortilin expression and inflammatory cytokine release when combine high glucose with ox-LDL in LSECs.We also show that overexpression of sortilin attenuate apoptosis of LSECs and decrease inflammatory cytokine release.These findings provide new insights into the molecular mechanism of sortilin and its potential therapeutic in the treatment of T2DM and NAFLD.