SENP3对ox-LDL诱导的巨噬细胞泡沫化的影响

Effect of SENP3 on formation of foam cells from macrophages induced by ox-LDL

目的:探讨小泛素样修饰物(SUMO)/sentrin特异性蛋白酶3(SENP3)对氧化型低密度脂蛋白(ox-LDL)诱导的巨噬细胞泡沫化的影响及其机制。方法:利用慢病毒感染并筛选构建4组稳转细胞,分为SENP3过表达组和对照组人源THP-1单核-巨噬细胞,以及SENP3敲减表达组和对照组小鼠RAW264.7巨噬细胞,利用ox-LDL诱导巨噬细胞构建泡沫细胞模型。采用油红O染色检测细胞内脂滴沉积;用试剂盒酶法测定细胞内胆固醇含量;Westernblot检测SENP3的表达量,以及pro-caspase-1的表达量和剪切情况;ELISA法检测细胞中促炎因子白细胞介素1β(IL-1β)的分泌。结果:与对照组相比,SENP3过表达显著降低THP-1细胞内脂滴含量,SENP3敲减表达显著提升RAW264.7细胞内脂滴含量;与对照组相比,SENP3过表达细胞内总胆固醇、胆固醇酯和胆固醇酯化率均显著降低(P<0.05或P<0.01);SENP3的表达量随ox-LDL刺激时间增加而显著下降(P<0.05或P<0.01);与对照组相比,SENP3过表达组pro-caspase-1的表达量没有显著差异,而剪切体caspase-1(p20)的含量显著下降(P<0.01);与对照组相比,SENP3过表达组细胞分泌的IL-1β蛋白量显著下降(P<0.05)。结论:SENP3蛋白可显著抑制巨噬细胞泡沫化,该作用可能是通过抑制炎症小体的激活实现的。

AIM:To investigate the effect of small ubiquitin-like modifier(SUMO)/sentrin-specific protease 3(SENP3)on the formation of foam cells from macrophages induced by oxidized low-density lipoprotein(ox-LDL)and its mechanism.METHODS:Cells were established by infection with lentivirus.After selection,they were divided into SENP3-overexpressing and control human THP-1 monocyte-macrophages,and SENP3-knockdown and control mouse RAW264.7 macrophages.All macrophages were induced by ox-LDL to establish foam cell model.Lipid droplet accumula-tion was determined by oil red O staining.Cholesterol levels were measured by enzyme testing kits.The expression of SENP3 and pro-caspase-1,and the cleavage of pro-caspase-1 were quantified by Western blot.Secretion of pro-inflamma-tory cytokine interleukin-1β(IL-1β)was detected by ELISA.RESULTS:Overexpression of SENP3 significantly de-creased THP-1 cell lipid droplets,while knockdown of SENP3 increased RAW264.7 cell lipid droplets.Overexpression of SENP3 significantly decreased the levels of total cholesterol and cholesterol ester,and cholesterol esterification rate(P<0.05 or P<0.01).Treatment with ox-LDL inhibited SENP3 expression in a time-dependent manner(P<0.05 or P<0.01).Compared with control group,overexpression of SENP3 decreased the amount of caspase-1(p20)without change of pro-caspase-1 expression.Compared with control group,over-expression of SENP3 significantly inhibited the secretion of IL-1β(P<0.05).CONCLUSION:The SENP3 significantly inhibits foam cell formation from macrophages through in-hibiting the activation of inflammasome.