| 摘要: |
| 目的 研究甲硫氨酸饥饿诱导胃癌细胞凋亡的分子机制。方法 利用癌症基因组图谱（TCGA）数据库分析胃癌组织中程序性死亡配体1（PD-L1）的表达及其与临床病理特征的关系。用普通培养液和甲硫氨酸饥饿培养液分别培养胃癌AGS细胞及抑制PD-L1表达的siRNA（siPD-L1）胃癌细胞，根据处理方法分为对照组、甲硫氨酸饥饿处理组、siPD-L1处理组、甲硫氨酸饥饿联合siPD-L1处理组，采用CCK-8检测胃癌细胞活力，吖啶橙/溴化乙锭双染检测胃癌细胞凋亡数目，流式细胞术检测胃癌细胞凋亡率，蛋白质印迹法检测胃癌细胞PD-L1、Bcl-2、B淋巴细胞瘤2相关X蛋白（Bax）、Caspase 3蛋白表达。starBase数据库分析PD-L1与Bcl-2抗凋亡蛋白家族（BCL2A1、MCL1、BCL2、BCL2L1）之间的联系。结果 PD-L1在胃癌组织中高表达（P<0.01），且PD-L1的表达与胃癌G分级有关（P<0.01）。甲硫氨酸饥饿和siPD-L1处理均能够降低胃癌细胞存活率（P<0.01），促进凋亡（P<0.01），抑制PD-L1和Bcl-2表达（P<0.05），上调促凋亡蛋白Bax、Caspase 3表达（P<0.01）；甲硫氨酸饥饿联合siPD-L1处理上述作用更强（P<0.05）。PD-L1与Bcl-2抗凋亡蛋白家族表达水平之间呈正相关（P<0.01），说明PD-L1是一个关键抗凋亡基因。结论 甲硫氨酸饥饿通过抑制PD-L1的表达下调抗凋亡蛋白Bcl-2并上调促凋亡蛋白Bax和Caspase 3表达，从而诱导胃癌细胞凋亡。 |
| 关键词: 甲硫氨酸饥饿 胃肿瘤 程序性死亡配体1 细胞凋亡 |
| DOI：10.16781/j.0258-879x.2020.12.1329 |
| 投稿时间：2019-08-07修订日期：2020-02-21 |
| 基金项目:国家自然科学基金（81760549，81872480）. |
|
| Methionine starvation induces apoptosis of gastric cancer cells by inhibiting programmed death ligand-1 |
| ZHOU Li-qiang,LI Shi-hao,WU You,ZHOU Qi,YUAN Yi-wu,XIN Lin* |
(Department of Gastrointestinal Surgery, the Second Affiliated Hospital of Nanchang University, Nanchang 330006, Jiangxi, China
*Corresponding author) |
| Abstract: |
| Objective To explore the molecular mechanism of methionine starvation induced apoptosis in gastric cancer cells. Methods The expression of programmed death ligand-1 (PD-L1) in gastric cancer tissues was analyzed by the cancer genome atlas (TCGA) database. The gastric cancer AGS cells and the gastric cancer cells treated with siRNA that inhibit the expression of PD-L1 (siPD-L1) were cultured with ordinary medium and methionine-starved medium. According to the treatment method, they were divided into control group, methionine starvation treatment group, siPD-L1 treatment group, and methionine starvation combined siPD-L1 treatment group. The cell viability of gastric cancer was detected by cell counting kit 8, the apoptosis of gastric cancer cells was detected by acridine orange/ethidium bromide double staining, and the apoptosis rate of gastric cancer cells was detected by flow cytometry. The expression levels of PD-L1, B-cell lymphoma-2 (Bcl-2), B-cell lymphoma-2-related X protein (Bax) and Caspase 3 were detected by Western blotting analysis. Finally, the relationship between PD-L1 and Bcl-2 anti-apoptotic protein family (Bcl-2 related protein A1[BCL2A1], myeloid cell leakemia 1[MCL1], BCL2, and Bcl-2 like 1[BCL2L1]) was analyzed by starBase database. Results PD-L1 was highly expressed in gastric cancer tissues (P<0.01), and the expression of PD-L1 was correlated with G grade of gastric cancer (P<0.01). Methionine starvation and siPD-L1 significantly decreased the survival rate of gastric cancer cells (P<0.01), promoted apoptosis (P<0.01), inhibited the expression of PD-L1 and Bcl-2 (P<0.05), and up-regulated the expression of proapoptotic proteins Bax and Caspase 3 (P<0.01). The effect of methionine starvation combined with siPD-L1 was even stronger (P<0.05). There was a positive correlation between PD-L1 and Bcl-2 anti-apoptotic protein family (P<0.01), indicating that PD-L1 is a key anti-apoptotic gene. Conclusion Methionine starvation can induce gastric cancer cell apoptosis by inhibiting the expression of PD-L1, downregulating the expression of anti-apoptotic protein Bcl-2 and up-regulating the expression of Bax and Caspase 3. |
| Key words: methionine starvation stomach neoplasms programmed death ligand-1 apoptosis |
