油酰乙醇胺对大鼠原代血管平滑肌细胞泡沫化病变的治疗作用

Therapeutic effect of oleoyl ethanolamine on foaming lesions of rat primary vascular smooth muscle cells

目的 探讨动脉粥样硬化的脂质代谢过程中,油酰乙醇胺(oleoyl ethanolamine,OEA)作为重要参与者的作用及其可能的分子机制。方法 通过氧化型低密度脂蛋白(oxidized low-density lipoprotein,ox-LDL)诱导大鼠原代血管平滑肌细胞泡沫化病变。将大鼠原代血管平滑肌细胞分为正常组(正常培养)、模型组(100μg/ml的ox-LDL刺激48 h)和OEA组(100μg/ml的ox-LDL与50μmol/L OEA刺激48 h),使用实时荧光定量PCR检测细胞泡沫化密切相关的受体与酶的mRNA表达水平,包括过氧化物酶体增殖物激活受体-α(peroxisome proliferator-activated receptor-α,PPAR-α)、B类清道夫受体白细胞分化抗原36(cluster of differentiation 36,CD36)、B族Ⅰ型清道夫受体(scavenger receptor class B typeⅠ,SR-BⅠ)、胆固醇酰基转移酶A(acyl-coenzyme A:cholesterol acyltransferase-1,ACAT-1)、胆固醇调节元件结合蛋白-1c(sterol regulatory element binding protein-1c,SREBP-1c)和肉毒碱棕榈酰转移酶-1(carnitine palmitoyl transferase-1,CPT-1)。使用化学酶促法检测各组实验细胞中总胆固醇(TC)、甘油三酯(TG)的含量,并计算细胞内胆固醇酯(cholesteryl ester,CE)。采用油红O染色法在显微镜下直观观察各组实验细胞内脂质堆积的情况。结果 与正常组相比,模型组ACAT-1、CD36和SREBP-1c表达升高(P<0.001),SR-BⅠ、CPT-1表达降低(P<0.001),脂代谢基因PPAR-α表达降低(P<0.05);细胞内TC、TG含量和CE/TC显著升高(P<0.001),细胞内脂滴明显增加。与模型组相比,OEA组ACAT-1、CD36和SREBP-1c表达降低(P<0.05),SR-BⅠ、CPT-1表达升高(P<0.001),脂代谢基因PPAR-α表达升高(P<0.01);细胞内TC、TG含量和CE/TC降低(P<0.001),并且可从显微镜直观观察到细胞内脂质堆积程度降低。结论 OEA上调脂代谢基因PPAR-α表达,可以改善细胞内脂质堆积情况,这一效果可能是通过PPAR-α通路调控CD36、ACAT-1、SREBP-1c、SR-BⅠ、CPT-1等重要脂质代谢基因实现的。

Objective To investigate the role of oleoyl ethanolamine(OEA)as an important participant in the lipid metabolism process of atherosclerosis and its possible molecular mechanisms.Methods The foam of rat primary vascular smooth muscle cells(VSMCs)was induced by oxidized low-density lipoprotein(ox-LDL).The cells were divided into blank group(normal culture),model group(stimulated with 100μg/ml ox-LDL for 48 h),and OEA group(stimulated with 100μg/ml ox-LDL and 50μmol/L OEA for 48 h).Real-time PCR was used to detect the mRNA expression levels of receptors and enzymes closely related to foam cell formation,including peroxisome proliferator-activated receptor-α(PPAR-α),cluster of differentiation 36(CD36),scavenger receptor class B typeⅠ(SR-BⅠ),acyl-coenzyme A:cholesterol acyltransferase-1(ACAT-1),sterol regulatory element binding protein-1c(SREBP-1c),and carnitine palmitoyl transferase-1(CPT-1).Total cholesterol(TC)and triglyceride(TG)contents were measured using a chemical enzyme kit,and cholesteryl ester(CE)content was calculated.Oil red O staining was used to observe the lipid accumulation under a microscope.Results Compared to normal group,the expression levels of ACAT-1,CD36,and SREBP-1c were increased in model group(P<0.001),and the expression levels of SR-BⅠand CPT-1 were decreased(P<0.001),and the expression of the lipid metabolism gene PPAR-αwas decreased(P<0.05).Compared to normal group,the intracellular contents of TC,TG,and CE/TC were significantly increased in model group(P<0.001),accompanied by a noticeable increase in intracellular lipid droplets.Compared with model group,the expression levels of ACAT-1,CD36,and SREBP-1c were reduced(P<0.05 or P<0.001),the expression levels of SR-BⅠand CPT-1 were increased(P<0.001),and the expression of lipid metabolism gene PPAR-αwas increased in OEA group(P<0.01);and the intracellular TC,TG contents,and CE/TC were also reduced in OEA group(P<0.001),and the degree of lipid accumulation in the cells was visually observed under the microscope.Conclusion OEA can upregulate the expression of the lipid metabolism gene PPAR-αand improve the intracellular lipid accumulation.This effect may be achieved by regulating the important lipid metabolism genes CD36,ACAT-1,SREBP-1c,SR-BⅠ,and CPT-1 through the PPAR-αpathway.