过氧化体增殖物激活型受体γ和δ在高密度脂蛋白介导细胞胆固醇流出中的作用

The Role of Peroxisome Proliferator-Activated Receptors-gamma and-delta in the Cellular Cholesterol Efflux Mediated by High Density Lipoprotein

为探讨过氧化体增殖物激活型受体γ和δ在高密度脂蛋白介导的细胞胆固醇流出中的作用。取新鲜抗凝血浆 ,用超速离心机进行密度梯度离心 ,收集密度为 1 .0 6 3～ 1 .2 1 0组分。U937细胞用 5 0～ 4 0 0nmol L反义过氧化体增殖物激活型受体γ和δ预处理细胞 1 2h。培养U937细胞与 0 .2 8mCi L [3H] 胆固醇乙醇液共孵育 2 4h ,15 0 0r min离心 1 0min ,收集细胞 ,PBS漂洗 2次 ,RPMI 1 6 4 0重悬细胞 ,加HDL继续培养 0～ 4 8h。液体闪烁计数仪测细胞相对放射活性。Westernblot检测过氧化体增殖物激活型受体γ和δ蛋白表达水平。不同浓度HDL处理后细胞胆固醇流出具有明显差异 ,呈剂量 -效应关系。用 1 0 0nmol L反义过氧化体增殖物激活型受体γ处理 2 4h后 ,HDL介导的细胞胆固醇流出减少 (4 5 78± 2 0 6 ) ,与对照组 (4 0 2 4± 385 )比较差别有显著性。过氧化体增殖物激活型受体γ激动剂ciglitizone预处理后 ,HDL介导的细胞胆固醇流出增加 ,用 0、2 5、5 0、1 0 0、2 0 0 μmol Lciglitizone处理的放射活性分别为 4 371± 2 4 3、386 9± 2 1 2、346 9± 2 0 9、31 5 6± 31 5和 30 2 0± 2 96。反义过氧化体增殖物激活型受体δ处理后 ,HDL介导的细胞胆固醇流出略有增加 ,处理浓度达 4 0 0nmol

Aim This study examines the relationship between PPAR gamma and delta and cholesterol efflux mediated by high density lipoprotein. Methods Pooled plasma were collected from health human. Component (density 1.063 1.21 g/cm 3) was prepared by density ultracentrifugation. U937 cells were treated by cholesterol for 24 h, cell pellets were washed with PBS, then added RPMI 1640 supplemented with 1% FBS. The cells were treated by the antisense PPAR gamma and delta oligo DNA, PPAR gamma activator ciglitazone, and PPAR responsive element decoy, respectively. The effects of the oligo DNA on efflux of cholesterol was evaluated by relative radioactivity of the cell pellets. PPAR gamma and delta protein expression levels were assayed by Western blot. Results The results demonstrated that HDL promoted cellular cholesterol efflux in dose and time dependent manner reflected by radioactivity of cell pellets. It was also observed an increase in HDL mediated cholesterol efflux associated with a decrease of antisense oligo DNA of PPAR gamma, and an decrease in HDL mediated cholesterol efflux associated with decrease of antisense oligo DNA of PPAR delta, both in a dose dependent manner. PPAR activator ciglitazone stimulated the HDL mediated cholesterol efflux. Treatment with PPAR responsive element decoy inhibited the cholesterol efflux mediated by HDL. Western blot assay demonstrated that PPARgamma protein level was increased, while PPARdelta level decreased by HDL. Conclusion This study suggests that PPAR gamma may improve the cholesterol efflux mediated by HDL, PPAR delta may prevent the HDL mediated cholesterol efflux.