沙苑子苷A对L02细胞脂肪变性模型的降脂作用和机制探讨  

Lipid-lowering effect and mechanism study of Complanatoside A on steatosis model of L02 cells

作　　者：孟靓 魏益谦 高晶 李丽[1] 邵紫萱 孟智睿 高学敏[1] 王景霞[1] MENG Liang;WEI Yiqian;GAO Jing;LI Li;SHAO Zixuan;MENG Zhirui;GAO Xuemin;WANG Jingxia(BeijingUniversity of Chinese Medicine,Beijing 100029,China)

机构地区：[1]北京中医药大学,100029 [2]北京中医药大学第三附属医院内分泌科

出　　处：《环球中医药》2021年第7期1204-1211,共8页Global Traditional Chinese Medicine

基　　金：国家自然科学基金(81673617)。

摘　　要：目的建立人正常肝细胞株(L02)脂肪变体外模型,探讨沙苑子苷A对脂肪变性肝细胞的降脂作用及相关机制。方法采用1 mmol/L的游离脂肪酸FFA(油酸∶棕榈酸为2∶1)诱导24小时建立L02细胞脂肪变模型,然后分为空白组对照组、模型组、沙苑子苷A低、中、高剂量组(25、50、100μM),按照试剂盒检测甘油三酯(triglyceride, TG)、胆固醇(cholesterol, TC)、谷丙转氨酶(glutamic pyruvic transaminase,ALT)、谷草转氨酶(glutamic oxaloacetic transaminase,AST)、乳酸脱氢酶(lactate dehydrogenase, LDH)、丙二醛(malondialdehyde, MDA)、超氧化物歧化酶(superoxide dismutase, SOD)的水平,Elisa法检测白介素6(interleukin-6,IL-6)、肿瘤坏死因子(tumor necrosis factor-α,TNF-α)的水平,qPCR检测固醇调节元件结合蛋白-1c(sterol regulatory element-binding protein 1C,SREBP-1c)、脂肪酸合成酶(fatty acid synthase, FAS)、过氧化物酶体增殖物激活受体α(peroxisome proliferator activated receptorα,PPARα)、肉毒碱棕榈酰基转移酶1A(carnitine Palmitoyltransferase 1A,CPT-1A) mRNA的表达,Western blot检测SREBP-1c、FAS、PPARα、CPT-1A蛋白的表达。结果与模型组比较,沙苑子苷A预处理能显著降低细胞TG、TC、ALT、AST、LDH、MDA的含量(P<0.05,P<0.01),显著升高SOD活性(P<0.01),显著降低IL-6、TNF-α的水平(P<0.01);同时,与模型组比较,沙苑子苷A预处理能显著降低肝细胞SREBP-1c、FAS mRNA和蛋白的表达量(P<0.01),显著增加肝细胞PPARα、CPT-1A mRNA和蛋白的表达量(P<0.01)。结论 FFA成功诱导了L02细胞的脂肪变性,沙苑子苷A对脂肪变的肝细胞具有良好的降脂和保肝作用,其降脂机制可能是通过抑制肝细胞SREBP-1c的表达,降低TG合成途径中限速酶FAS水平,降低TG合成速度;同时上调PPARα蛋白表达,提高脂肪酸β氧化途径中CPT-1A水平,加速脂肪酸的β氧化,减少TG合成,从两方面共同作用从而达到降脂和保肝效果。Objective To construct an in vitro steatosis model with normal human hepatocytes line(L02),and to explore the lipid-lowering effect of Complanatoside A on Steatosis hepatocytes.and its mechanism.Methods The model of Steatosis L02 cells was induced with 1 mmol/L FFA(oleic acid∶palmitic acid=2∶1)for 24 h.And then cells were divided into blank group,control group,model group,and complanatoside A low-dose,medium-dose and high-dose groups(25,50,and 100μM).The levels of TG,TC,ALT,AST,LDH,MDA,and SOD were detected according to the kit.Elisa was used to determine the levels of IL-6 and TNF-α.QPCR was used to assess the mRNA expressions of SREBP-1c,FAS,PPARα,and CPT-1a.Western blot was used to examine the protein expressions of SREBP-1c,FAS,PPARα,and CPT-1A.Results Complanatoside A with pretreatment significantly decreased the contents of TG,TC,ALT,AST,LDH and MDA(compared with the model group,P<0.05,P<0.01),significantly increased the activity of SOD(compared with the model group,P<0.01),and significantly decreased the levels of IL-6 and TNF-α(compared with the model group,P<0.01).At the same time,Complanatoside A with pretreatment significantly reduced the mRNA and protein expressions of SREBP-1c and FAS in hepatocytes(compared with the model group,P<0.01),and significantly increased the mRNA and protein expressions of PPARαand CPT-1A in hepatocytes(compared with the model group,P<0.01).Conclusion FFA can successfully induce L02 cells steatosis.And complanatoside A has good lipid-lowering and hepatoprotective effects on steatosis liver cells.The mechanism of the lipid-lowering effect may be the interaction of the following two aspects.On the one hand,by inhibiting the expression of SREBP-1c in hepatocytes,the level of FAS in TG synthesis process decreases,and the rate of TG synthesis reduces;on the other hand,by up-regulating the protein expression of PPARα,the level of CPT-1A in the fatty acidsβ-oxidation process increases,theβ-oxidation of fatty acids is accelerated,and TG synthesis was reduced.These two

关 键 词：沙苑子苷A L02细胞 脂质代谢紊乱 qPCR检测固醇调节元件结合蛋白-1c 过氧化物酶体增殖物激活受体Α 

分 类 号：R285.5[医药卫生—中药学]