不同浓度桦木酸对人胃癌MGC-803细胞自噬的影响

Effects of betulinic acid at different concentrations on autophagy of human gastric cancer MGC-803 cells

目的:本研究利用不同浓度桦木酸处理人胃癌MGC-803细胞,以探究其对细胞自噬的影响。方法:将人胃癌MGC-803细胞分为4组,每组3个复孔,对照组不加桦木酸处理,其余三组分别加入终浓度为10、20、30 mg/L桦木酸。桦木酸处理细胞48 h后,qRT-PCR检测桦木酸对人胃癌MGC-803细胞自噬相关基因mRNA表达的影响。Western blot检测药物处理细胞自噬相关基因的蛋白表达。利用免疫荧光检测药物处理后MGC-803细胞内LC3蛋白的胞内定位及表达。结果:与对照组相比,在10~30 mg/L浓度范围内,桦木酸处理的人胃癌MGC-803细胞LC3、Beclin-1 mRNA的表达明显升高(P<0.05),Beclin-1、LC3-Ⅱ蛋白的表达显著升高(P<0.05),LC3-Ⅰ蛋白的表达明显降低(P<0.05),其中30 mg/L处理组表现最佳。此外,桦木酸还可诱导MGC-803细胞LC3蛋白在细胞质内形成点状聚集。结论:在10~30 mg/L浓度范围内,桦木酸能诱导人胃癌MGC-803细胞发生自噬。

Objective:In this study,human gastric cancer MGC-803 cells were treated with betulinic acid at different concentrations to investigate its effect on cell autophagy.Methods:The human gastric cancer MGC-803 cells were divided into 4 groups,each group was set with 3 replicate.The control group was not treated with betulinic acid,the other three groups were added with final concentration of 10,20,30 mg/L betulinic acid,respectively.Cells were treated with betulinic acid for 48 h,qRT-PCR was applied to detect the effect of betulinic acid on mRNA expressions of autophagy-related genes in human gastric cancer MGC-803 cells.Western blot was performed to determine the protein expressions of cell autophagy-related genes after drug treatment.Immunofluorescence was used to detect the localization and expression of LC3 protein in MGC-803 cells after drug treatment.Results:Compared with the control group,in the concentration range of 10~30 mg/L,the mRNA expression of LC3 and Beclin-1 in human gastric cancer MGC-803 cells treated with betulinic acid were increased significantly,the expressions of Beclin-1 and LC3-Ⅱprotein were also increased significantly,while the expression of LC3-Ⅰprotein was decreased significantly.Among them,betulinic acid at the concentration of 30 mg/L showed the best effects.In addition,betulinic acid induced the LC3 protein in MGC-803 cells to form spot aggregates in the cytoplasm.Conclusion:At the concentrations of 10~30 mg/L,betulinic acid can induce autophagy in human gastric cancer MGC-803 cells.