阿米替林对非酒精性脂肪性肝病细胞模型脂质沉积及生化代谢的影响

目的探讨阿米替林通过调节酸性鞘磷脂酶(ASM)/神经酰胺(CE)通路对非酒精性脂肪性肝病(NAFLD)细胞模型脂质及生化代谢的影响。方法体外培养HepG2和L02细胞构建NAFLD细胞模型,MTT比色法测定细胞增殖率,油红O染色观察细胞内脂滴变化。实验分组:正常对照组、模型组、Ami组、TNFα组、Ami+TNFα组。全自动生化分析仪检测细胞内TG、TC及细胞上清液ALT、AST水平,ELISA法检测细胞内总的CE、ASM水平,Western Blot检测细胞内ASM蛋白的表达;实时荧光定量PCR检测细胞内ASM mRNA水平的表达。计量资料多组间比较采用单因素方差分析,进一步两两比较采用Turkey检验。结果与正常对照组相比,NAFLD模型组ASM蛋白和mRNA表达量以及CE、TG、TC、ALT、AST水平明显升高(P值均<0.05);与模型组相比,Ami组ASM蛋白和mRNA表达量以及CE、TG、TC、ALT、AST水平明显降低(P值均<0.05),TNFα组ASM蛋白和mRNA表达量以及CE、TG、ALT、AST水平明显升高(P值均<0.05);与TNFα组比较,Ami+TNFα组ASM蛋白和mRNA表达量以及CE、TG、TC、ALT、AST水平明显降低(P值均<0.05)。结论ASM/CE通路促进脂质积聚、导致脂肪变,阿米替林可通过抑制该通路改善NAFLD肝细胞的脂质沉积。

Effect of amitriptyline on lipid deposition and biochemical metabolism in a cell model of nonalcoholic fatty liver disease

Objective To investigate the effect of amitriptyline on lipid deposition and biochemical metabolism in a cell model of nonalcoholic fatty liver disease(NAFLD)by regulating the acid sphingomyelinase(ASM)/ceramide(CE)pathway.Methods HepG2 and L02 cells were cultured in vitro to establish a cell model of NAFLD.MTT colorimetry was used to measure cell proliferation rate,and oil red O staining was used to observe the change of lipid droplets in cells.In the experiment,the cells were divided into normal control group,model group,Ami group,TNFαgroup,and Ami+TNFαgroup.An automatic biochemical analyzer was used to measure the levels of triglyceride(TG)and total cholesterol(TC)in cells and the levels of alanine aminotransferase(ALT)and aspartate aminotransferase(AST)in supernatant;ELISA was used to measure the levels of CE and ASM in cells;Western blot was used to measure the protein expression ASM in cells,and RT-PCR was used to measure the mRNA expression of ASM in cells.A one-way analysis of variance was used for comparison of continuous data between multiple groups,and the Turkey test was used for further comparison between two groups.Results Compared with the normal control group,the NAFLD model group had significant increases in the protein and mRNA expression of ASM and the levels of CE,TG,TC,ALT,and AST(all P<0.05).Compared with the model group,the Ami group had significant reductions in the protein and mRNA expression of ASM and the levels of CE,TG,TC,ALT,and AST(all P<0.05),and the TNFαgroup had significant increases in the protein and mRNA expression of ASM and the levels of CE,TG,ALT,and AST(all P<0.05).Compared with the TNFαgroup,the Ami+TNFαgroup had significant reductions in the protein and mRNA expression of ASM and the levels of CE,TG,TC,ALT,and AST(all P<0.05).Conclusion The ASM/CE pathway promotes lipid accumulation and may lead to hepatocyte steatosis,and amitriptyline can alleviate lipid deposition in NAFLD hepatocytes by inhibiting the ASM/CE pathway.