下调lncRNA LINC00176对肺癌A549/DDP细胞顺铂耐药和自噬的影响

目的：研究下调lncRNA LINC00176对肺癌A549/DDP细胞顺铂耐药和自噬的影响及机制。方法：qRT-PCR方法测定正常支气管上皮16HBE细胞和肺癌A549、A549/DDP、NCI-H1299、SK-MES-1细胞中LINC00176的表达。将A549/DDP细胞分成对照组、si-NC组、si-LINC00176组、si-LINC00176+Anti-miR-NC组、si-LINC00176+Anti-miR-138-5p组。MTT实验检测顺铂对A549/DDP的半数抑制浓度（IC50）。流式细胞术测定细胞凋亡。Western blot测定LC3-I、LC3-II、Beclin 1、C-Caspase-3蛋白表达。利用荧光素酶报告系统鉴定LINC00176和miR-138-5p的靶向关系。结果：与16HBE细胞比较，A549、A549/DDP、NCI-H1299、SK-MES-1细胞中LINC00176表达水平显著升高（P<0.01）。与A549细胞比较，A549/DDP细胞中LINC00176表达水平显著升高（P<0.01）。与对照组、si-NC组比较，si-LINC00176组A549/DDP细胞IC50显著降低（P<0.01），LC3-II/LC3-I、Beclin 1蛋白表达显著降低（P<0.01），凋亡率、C-Caspase-3蛋白表达、miR-138-5p表达水平显著升高（P<0.01）。LINC00176与miR-138-5p直接结合。与si-LINC00176+Anti-miR-NC组比较，si-LINC00176+Anti-miR-138-5p组A549/DDP细胞IC50显著升高（P<0.01），LC3-II/LC3-I、Beclin 1蛋白表达显著升高（P<0.01），凋亡率、C-Caspase-3蛋白表达显著降低（P<0.01）。 结论：下调lncRNA LINC00176通过靶向上调miR-138-5p能够抑制肺癌A549/DDP细胞顺铂耐药，抑制细胞自噬，诱导细胞凋亡。

Effects of down-regulating lncRNA LINC00176 on cisplatin resistance and autophagy in lung cancer A549/DDP cells

AIM: To study the effects and mechanism of down-regulating lncRNA LINC00176 on cisplatin resistance and autophagy in lung cancer A549/DDP cells. METHODS: The qRT-PCR method was used to determine the expression changes of LINC00176 in normal bronchial epithelial 16HBE cells and lung cancer A549, A549/DDP, NCI-H1299 and SK-MES-1 cells. A549/DDP cells were divided into Control group, si-NC group, and si-LINC00176 group, si-LINC00176+ Anti-miR-NC group, and si-LINC00176+ Anti-miR-138-5p group. MTT experiment detected the half inhibitory concentration (IC50) of cisplatin on A549/DDP. Flow cytometry was used to measure cell apoptosis. Western blot was used to determine the protein expression of LC3-I, LC3-II, Beclin 1, C-Caspase-3. The luciferase reporter system was used to identify the targeting relationship between LINC00176 and miR-138-5p. RESULTS: The expression level of LINC00176 in A549, A549/DDP, NCI-H1299 and SK-MES-1 cells was higher than that in 16HBE cells (P<0.01). The expression level of LINC00176 in A549/DDP cells was significantly higher than that in A549 cells (P<0.01). Compared with the Control and si-NC groups, the IC50 of A549/DDP cells in the si-LINC00176 group was significantly reduced (P<0.01), the expression of LC3-II/LC3-I, Beclin 1 protein was significantly reduced (P<0.01), the apoptosis rate, and the expression of C-Caspase-3 protein and miR-138-5p were significantly increased (P<0.01). LINC00176 directly bound to miR-138-5p. Compared with the si-LINC00176+Anti-miR-NC group, the IC50 of A549/DDP cells in the si-LINC00176+Anti-miR-138-5p group was significantly increased (P<0.01), and the expression of LC3-II/LC3-I and Beclin 1 protein was significantly increased (P<0.01), the apoptosis rate and C-Caspase-3 protein expression were significantly reduced (P<0.01).CONCLUSION: Down-regulating lncRNA LINC00176 could inhibit cisplatin resistance, inhibit autophagy and induce apoptosis in lung cancer A549/DDP cells by targeting and up-regulating miR-138-5p.