Tob1通过诱导自噬抑制肾上腺皮质癌细胞SW-13增殖促进细胞凋亡的机制研究

Tob1 Activates Autophagy to Inhibits Cell Proliferation and Promotes Apoptosis of Adrenocortical Carcinoma Cells

目的:探讨Tob1对肾上腺皮质癌细胞自噬的作用及其影响。方法:将体外培养的SW-13细胞分为对照组、空载质粒转染组以及pcDNA3.1-Tob1质粒转染组。实时荧光定量PCR检测各组细胞中Tob1的mRNA表达。Western blot检测Tob1以及自噬相关蛋白LC3-Ⅰ、LC3-Ⅱ、p62和Beclin 1的表达。LC3免疫荧光检测细胞自噬水平。随后在转染pcDNA3.1-Tob1质粒的SW-13细胞中加入自噬抑制剂3-MA处理,CCK-8检测细胞增殖,Tunel检测细胞凋亡。结果:pcDNA3.1-Tob1质粒转染显著上调SW-13细胞中Tob1的mRNA和蛋白表达,提升LC3-Ⅱ/LC3-Ⅰ和Beclin 1水平,降低p62蛋白表达,并显著提升细胞自噬水平(P<0.05)。与对照组相比,pcDNA3.1-Tob1组SW-13细胞增殖降低,细胞凋亡增加;与pcDNA3.1-Tob1组相比,pcDNA3.1-Tob1+3-MA组细胞增殖提升,细胞凋亡降低(P<0.05)。结论:Tob1能够通过激活细胞自噬降低人皮质癌细胞SW-13增殖并促进其凋亡。

Objective:To study the effect of Tob1 on autophagy of adrenocortical carcinoma cells.Methods:The cultured SW-13 cells were divided into control group,empty plasmid transfection group and pcDNA3.1-Tob1 plasmid transfection group.Real-time quantitative PCR was used to detect Tob1 mRNA expression in each group.Protein expression of Tob1,LC3-Ⅰ,LC3-Ⅱ,p62 and Beclin 1 were measured by Western blot.LC3 immunofluorescence was used to detect cell autophagy level.Subsequently,SW-13 cells transfected with pcDNA3.1-Tob1 were treated with autophagy inhibitor 3-MA.CCK-8 was used to detect cell proliferation and Tunel assay was used to detect cell apoptosis.Results:pcDNA3.1-Tob1 transfection significantly up-regulated the mRNA and protein expression of Tob1,elevated LC3-Ⅱ/LC3-Ⅰand Beclin 1 protein level,decreased p62 expression,and significantly promoted the autophagy level in SW-13 cells(P<0.05).Compared with the control group,SW-13 cell proliferation was decreased and apoptosis was increased in pcDNA3.1-Tob1 group.Compared with pcDNA3.1-Tob1 group,cell proliferation was increased and cell apoptosis was decreased in pcDNA3.1-Tob1+3-MA group(P<0.05).Conclusion:Tob1 can inhibit the proliferation of human cortical adenocarcinoma cell SW-13 and promote its apoptosis by activating autophagy.