对氧磷酶-1参与调控非酒精性脂肪肝脂质代谢与胰岛素抵抗作用及机制

目的探究对氧磷酶-1(Paraoxonase 1,PON1)与非酒精性脂肪肝(Nonalcoholic fatty liver disease,NAFLD)脂质代谢与胰岛素抵抗关系及机制。方法选取129例NAFLD患者和130非NAFLD患者为研究对象,收集患者年龄、身高、体重、腰围,血压等信息,检测血脂、空腹血糖(Fast Glucose)、胰高血糖素(Glucagon)、胰岛素(Fast Insulin)、评估胰岛素抵抗的稳态指数(homeostatic index of insulin resistance,HOMA-IR)、肝肾功能、脂肪因子等,LO2细胞接种12孔板培养24 h后分别加入0~1.2 mmol OA溶液处理24 h或10 ng/mL TNF-α处理6 h,以PEI转染空载质粒和PON1质粒1μg 24 h。使用尼罗红染料测定细胞内脂质含量。ELISA法检测PON1活性,蛋白质印迹检测PON1、p-IRS1、T-IRS1、p-AKT T-AKT、FAS、HMGCR、SREBP2和SREBP1C蛋白表达。结果NAFLD患者腰围、BMI、外周血ALT、AST、FFA、TG、TC、LDL-C、VLDL-C、空腹血糖、胰岛素、HOMA-IR、抵抗素、瘦素、IL-6和TNF-α显著高于非NAFLD,PON1活性、含量、HDL-C、胰高血糖素和脂联素显著降低;血PON1活性和含量与FFAs、TG、TC、LDL-C、VLDL-C、空腹血糖、胰岛素、HOMA-IR、抵抗素、瘦素、IL-6和TNF-α显著负相关,与HDL-C、胰高血糖素和脂联素显著正相关;OA组LO2细胞PON1的活性逐渐降低,且0.2~1.2 mM OA组LO2细胞PON1的活性显著低于Control组,过表达PON1不影响PON1活性,0.2~1.2 mM OA组LO2细胞内脂质含量显著高于Control组,显著低于OA组;TNF-α组LO2细胞PON1、p-IRS1、p-AKT、FAS、HMGCR、SREBP2和SREBP1C蛋白表达量相比于Control组显著降低,转染PON1质粒显著增加PON1,同时p-IRS1、p-AKT、FAS、HMGCR、SREBP2和SREBP1C蛋白表达量显著增加。结论PON1活性与含量和NAFLD血脂代谢及胰岛素抵抗显著相关,且可能通过调控p-IRS1/AKT通路影响胰岛素抵抗,并调控脂质代谢。

The role and mechanism of paraoxonase 1 in the regulation of lipid metabolism and insulin resistance in nonalcoholic fatty liver disease

Objective To investigate the role and mechanism of paraoxonase 1(PON1)and lipid metabolism,insulin resistance in nonalcoholic fatty liver disease(NAFLD).Methods One hundred and twenty-nine NAFLD patients and 130 non-NAFLD individuals were enrolled in the study.The age,height,weight,waist circumference,blood pressure and other information were collected.Plasma blood lipids,fast glucose,and glucagon were detected by routine blood tests,homeostatic index of insulin resistance(HOMA-IR)was calculated,liver and kidney function were evaluated in biochemical assay,LO2 cells were inoculated in 12-well plates for 24 h and then added 0~1.2 mM OA solution was treated for 24 h or 10 ng/mL TNF-αfor 6 h,and the 1μg control and PON1 plasmid were transfected with PEI for 24 h.The intracellular lipid content was determined using a Nile Red dye.PON1 activity was detected by ELISA,and PON1,p-IRS1,T-IRS1,p-AKT T-AKT,FAS,HMGCR,SREBP2 and SREBP1C protein were detected by Western Blot.Results Waist circumference,BMI,peripheral blood ALT,AST,FFAs,TG,TC,LDL-C,VLDL-C,fasting blood glucose,insulin,HOMA-IR,resistin,leptin,IL-6 and TNF-αwere significantly higher in patients with NAFLD.In non-NAFLD,PON1 activity,content,HDL-C,glucagon and adiponectin were significantly reduced;PON1 activity and content were significantly negatively correlated with FFAs,TG,TC,LDL-C,VLDL-C,fasting blood glucose,insulin,HOMA-IR,resistin,leptin,IL-6 and TNF-α,and positively correlated with HDL-C,glucagon and adiponectin;PON1 activity in LO2 cells of OA group gradually decreased,and the activity of PON1 in LO2 cells of 0.2~1.2 mM OA group was significantly lower than that of control group.Overexpression of PON1 did not affect PON1 activity,the lipid content of LO2 cells in 0.2~1.2 mM OA group was significantly higher.In the control group,it was significantly lower than the OA group;the expression levels of PON1,p-IRS1,p-AKT,FAS,HMGCR,SREBP2 and SREBP1C in the TNF-αgroup were significantly lower than those in the control group,and the transfected PON1 plasmid was significantly increased.PON1,while the expression levels of p-IRS1,p-AKT,FAS,HMGCR,SREBP2 and SREBP1C protein were significantly increased.Conclusion PON1 activity and content are significantly correlated with NAFLD lipid metabolism and insulin resistance,and may affect insulin resistance and regulate lipid metabolism by regulating p-IRS1/AKT pathway.