基于SCAP/SREBP-2通路探讨电针对高脂血症大鼠胆固醇代谢的影响

Electroacupuncture mitigates hyperlipidemia via improving cholesterol metabolism mediated by SCAP/SREBP-2 signaling in liver tissue in rats

目的:观察电针对高脂血症(HLP)大鼠固醇调节元件结合蛋白裂解活化蛋白(SCAP)、固醇调节元件结合蛋白-2(SREBP-2)及其下游胆固醇代谢靶基因3羟基3甲基戊二酰辅酶A还原酶(HMGCR)、前蛋白转化酶枯草溶菌素9(PCSK9)、低密度脂蛋白受体(LDLR)表达的影响,探讨电针调节胆固醇代谢从而治疗HLP的机制。方法:SD大鼠随机挑选10只作为空白组饲喂基础饲料,其余大鼠饲喂高脂饲料28 d建立HLP模型。造模成功大鼠随机分为模型组、电针组,每组10只。电针组电针双侧“丰隆”“阴陵泉”,留针30 min,每天治疗1次,治疗28 d。全自动生化分析仪测定血清中总胆固醇(TC)、甘油三酯(TG)、高密度脂蛋白胆固醇(HDL-C)、低密度脂蛋白胆固醇(LDL-C)含量及谷草转氨酶(AST)、谷丙转氨酶(ALT)活性;高效液相色谱法检测肝组织的TC含量;Western blot法检测肝组织SCAP、SREBP-2及HMGCR、PCSK9、LDLR的蛋白表达水平;荧光定量PCR法检测肝组织SCAP、SREBP-2及HMGCR、PCSK9、LDLR mRNA表达水平;免疫荧光法检测肝组织SCAP蛋白表达水平。结果:与空白组相比,模型组大鼠肝组织内TC含量,血清AST、ALT活性及TC、LDL-C含量显著升高(P<0.01);肝组织SCAP、SREBP-2、HMGCR、PCSK9蛋白及mRNA表达均显著升高(P<0.01),LDLR mRNA及蛋白表达水平显著降低(P<0.01)。与模型组相比,电针组大鼠肝组织TC含量,血清AST、ALT活性及TC、LDL-C含量显著降低(P<0.01);肝组织SCAP、SREBP-2、HMGCR、PCSK9蛋白及mRNA表达均显著降低(P<0.01),LDLR蛋白表达水平明显升高(P<0.05)。结论:电针能够抑制肝组织胆固醇合成,改善HLP,其机制可能与抑制肝组织SCAP/SREBP-2表达,下调下游胆固醇合成代谢靶基因HMGCR、PCSK9表达,上调LDLR表达有关。

Objective To explore the effect of electroacupuncture(EA) on sterol regulatory element-binding protein(SREBP) cleavage-activating protein(SCAP)/SREBP-2 signaling and the expressions of its downstream cholesterol metabolism related molecules 3-hydroxy-3-methylglutaryl-CoA reductase(HMGCR),proprotein convertase subtilisin/kexin type 9(PCSK9),and low-density lipoprotein receptor(LDLR) in the liver tissue in rats with hyperlipidemia(HLP),so as to reveal its mechanisms underlying improvement of HLP.Methods Male SD rats were randomly divided into normal control,HLP model and EA groups(n=10/group).The HLP model was established by feeding the rats with high-fat diet for 28 d.Rats in the EA group received EA stimulation(2 Hz/100 Hz,2 mA) at “Fenglong”(ST40) and “Yinlingquan”(SP9) for 30 min,once daily for 28 d.The contents of total cholesterol(TC),triglyceride(TG),high density lipoprotein-cholesterol(HDL-C),low-density lipoprotein cholesterol(LDL-C) in the serum,the activity of glutamic oxaloacetic transaminase(AST) and glutamic pyruvic transaminase(ALT) were detected by automatic biochemical analysis.The content of TC in the liver tissue was detected using high performance liquid chromatography.The mRNA and protein expression levels of SCAP,SREBP-2,HMGCR,PCSK9 and LDLR in the liver tissue were measured by using quantitative real-time PCR and Western blot,respectively.The immunofluorescence density of liver SCAP was determined by using immunofluorescence histochemistry.Results Compared with the normal control group,the contents of liver TC,serum TC,LDL-C,the activities of AST and ALT,and the mRNA and protein expression levels of SCAP,SREBP-2,HMGCR,PCSK9 as well as SCAP immunoactivity were significantly increased(P<0.01),while the LDLR mRNA and protein levels were markedly decreased(P<0.01) in the model group.In comparison with the model group,the contents of liver TC,serum TC,LDL-C,the activities of AST and ALT and the expression of SCAP,SREBP-2,HMGCR,PCSK9 mRNAs and proteins and SCAP immunoactivity were considerably decreased in the EA group(P<0.01),while the LDLR protein level was evidently increased in the EA group(P<0.05).Conclusion EA intervention can inhibit the synthesis of cholesterol in the liver and thus improve hyperlipidemia in HLP rats,which may be realized by down-regulating the protein and mRNA expressions of hepatic SCAP/SREBP-2,HMGCR and PCSK9,and up-regulating LDLR protein.