复方谷氨酰胺胶囊通过靶向自噬相关基因5介导自噬诱导胃癌细胞凋亡的研究

Compound glutamine capsules induce gastric cancer cell apoptosis by targeting autophagy-related gene 5 and mediating autophagy

目的 探讨复方谷氨酰胺胶囊(CGEC)对人胃癌细胞AGS凋亡的影响及其作用机制。方法 将AGS细胞分为4组:对照组(不作处理)、CGEC组(100μg·mL^(-1)的CGEC处理)、CGEC+si-NC组(100μg·mL^(-1)的CGEC处理+转染si-NC)和CGEC+si-ATG5组(100μg·mL^(-1)的CGEC处理+转染si-ATG5)。用实时荧光定量聚合酶链反应(RT-qPCR)法检测各组细胞ATG5 mRNA相对表达水平;用蛋白质印迹法检测各组细胞自噬相关基因5(ATG5)以及增殖和凋亡相关蛋白表达情况;用克隆形成实验检测各组细胞集落形成;用流式细胞术检测各组细胞凋亡率。结果 对照组,CGEC组,CGEC+si-NC组,CGEC+si-ATG5组的ATG5蛋白表达水平分别为0.35±0.06,1.08±0.10,1.06±0.10和0.52±0.07;细胞克隆数分别为(312.06±27.34),(175.12±15.56),(188.36±18.56)和(291.10±29.23)个;细胞凋亡率分别为(3.74±0.48)%,(21.01±2.79)%,(21.22±2.20)%和(11.62±1.58)%;抗增殖细胞核抗原(PCNA)蛋白表达水平分别为1.08±0.13,0.50±0.04,0.56±0.08和0.79±0.11;B淋巴细胞瘤-2相关X蛋白(Bax)表达水平分别为0.31±0.03,0.88±0.08,0.91±0.09和0.50±0.04。对照组的上述指标与CGEC组比较,CGEC+si-NC组的上述指标与CGEC+si-ATG5组比较,差异均有统计学意义(均P<0.05)。结论 CGEC可通过上调ATG5的表达诱导自噬进而抑制AGS细胞增殖,并促进凋亡。

Objective To investigate the effect of compound glutamin entresoluble capsules(CGEC) on the apoptosis of human gastric cancer cells AGS and its mechanism. Methods AGS cells were divided into four groups: control group(no treatment), CGEC group(100 μg·mL^(-1)CGEC treatment), CGEC +si-NC group(100 μg·mL^(-1)CGEC treatment + transfect si-NC) and CGEC + si-ATG5 group(100 μg·mL^(-1)CGEC treatment + transfect si-ATG5). The relative expression level of autophagy-related gene 5(ATG5) mRNA was detected by real-time polymerase chain reaction. The expressions of ATG5 and proliferation and apoptosis related proteins were detected by Western blot. Colony formation was detected by clonal formation assay. The apoptosis rate of each group was detected by flow cytometry. Results The expression levels of ATG5 protein in control group, CGEC group CGEC + si-NC group and CGEC + si-ATG5 group were 0. 35 ± 0. 06, 1. 08 ± 0. 10, 1. 06 ± 0. 10 and 0. 52 ± 0. 07,respectively. The number of cell clones were 312. 06 ± 27. 34, 175. 12 ± 15. 56, 188. 36 ± 18. 56 and291. 10 ± 29. 23, respectively. The apoptosis rates were(3. 74 ± 0. 48) %,(21. 01 ± 2. 79) %(21. 22 ± 2. 20) %,(11. 62 ± 1. 58) %. The expression levels of proliferating cell nuclear antigen(PCNA) protein were 1. 08 ± 0. 13,0. 50 ± 0. 04, 0. 56 ± 0. 08, 0. 79 ± 0. 11, respectively. The expression levels of B-cell lymphoma-2 associated Xprotein(Bax) were 0. 31 ± 0. 03, 0. 88 ± 0. 08, 0. 91 ± 0. 09, 0. 50 ± 0. 04, respectively. The above indexes weresignificantly different between the control group and the CGEC group, between the CGEC + si-NC group and theCGEC + si-ATG5 group.Conclusion CGEC can inhibit AGS cells proliferation and promote apoptosis by up-regulating ATG5 expression and inducing autophagy.