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目的明确Sigma-1受体在非酒精性脂肪性肝病中的表达变化及意义。方法应用软脂酸诱导HepG2细胞脂肪变建立非酒精性脂肪性肝病(non-alcoholic fatty liver disease,NAFLD)体外模型,并按0、2、4、8、12 h时相点收获细胞,利用尼罗红(nile red)染色检测细胞脂肪变程度,通过Real-time PCR及Western blot检测Sigma-1受体(Sigma-1 recep-tor,Sigma-1R)及内质网应激标志蛋白葡糖调节蛋白78(glucose regulating protein,GRP78)的表达,二甲基噻唑二苯基四氮唑溴盐[3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide,MTT]检测细胞脂肪变过程中细胞活性的变化。结果 HepG2细胞经脂肪酸诱导后发生了脂肪变,脂肪变程度随时间点的延长而加重,细胞的平均脂变面积在12 h组(435.14±39.87)较0 h组明显升高(P<0.01);8、12 h组GRP78 mRNA的相对表达量[(2.35±0.51),(8.71±1.20)]较0 h组明显升高(P<0.05),而Sigma-1R mRNA相对表达量在2 h组(0.19±0.02)即开始出现明显下降(P<0.05);在蛋白水平上,GRP78的表达在8、12 h组[(25.59±0.51),(28.43±1.20)]较0 h组明显上调,Sigma-1R在8、12 h组[(15.92±0.42),(10.73±0.63)]较0 h组明显下调(P<0.05);MTT检测提示细胞在脂肪变过程中细胞活性逐渐下降,12 h组细胞活性[(59.86±4.47)%]相对0 h组明显下降(P<0.05)。结论 Sigma-1R的低表达可能诱发了NAFLD过程中的内质网应激,进而导致肝细胞的损伤。
Objective To clarify the changes of sigma-1 receptor in non-alcoholic fatty liver disease and its significance. Methods The model of non-alcoholic fatty liver disease (NAFLD) was induced by palmitate in the HepG2 cell steatosis. The cells were harvested at 0, 2, 4, 8 and 12 h, The degree of fatty degeneration was detected by nile red staining. Sigma-1 recep-tor (Sigma-1R) and endoplasmic reticulum stress marker glucose regulation The expression of glucose regulating protein (GRP78), the detection of cells by MTT (dimethylthiazolium-2-yl) -2,5-diphenyltetrazolium bromide Changes in cell viability during steatosis. Results Steatosis occurred in HepG2 cells after fatty acid induction, and the degree of steatosis increased with the prolongation of time point. The mean area of adipocytosis in HepG2 cells was significantly higher at 12 h (435.14 ± 39.87) than that at 0 h (P <0.01) ; The relative expression of GRP78 mRNA in 8,12 h group was significantly higher than that in 0 h group [(2.35 ± 0.51), (8.71 ± 1.20)], while the relative expression level of Sigma-1R mRNA in 2 h group (0.19 ± 0.02) (P <0.05). At the protein level, GRP78 expression was significantly up-regulated in 8,12 h group [(25.59 ± 0.51), (28.43 ± 1.20) The levels of Sigma-1R in 8,12 h group [(15.92 ± 0.42), (10.73 ± 0.63)] were significantly lower than those in 0 h group (P <0.05). The MTT assay showed that the cell viability decreased gradually during 12 h The cell viability of [(59.86 ± 4.47)%] was significantly lower than that of 0 h group (P <0.05). Conclusion Low expression of Sigma-1R may induce endoplasmic reticulum stress during NAFLD, leading to hepatocellular injury.