PRC1通过MAPK途径促进肺癌对顺铂耐药的机制研究

Mechanism of PRC1 promoting cisplatin resistance of lung cancer through MAPK pathway

探讨PRC1与肺癌顺铂耐药的关系及可能机制。方法采用Western blotting检测不同浓度顺铂处理不同时间下A549细胞中PRC1蛋白水平。采用慢病毒转染和PCMV-PRC1过表达质粒转染构建稳定干扰和过表达PRC1的A549细胞(shPRC1组和PCMV-PRC1组),采用CCK-8法和克隆形成实验检测不同PRC1表达对顺铂杀伤作用的影响。Western blotting检测shPRC1组和PCMV-PRC1组p-ERK1/2、cleaved PARP、cleaved caspase-3蛋白水平。结果PRC1蛋白水平随着顺铂浓度升高而升高,随作用时间的延长而升高。A549/DDP细胞中PRC1的蛋白水平显著高于A549细胞(4.04±0.50 vs.0.99±0.13,P<0.01)。5μg/ml顺铂处理下,与阴性对照组比较,shPRC1组细胞存活数量下降,PCMV-PRC1组细胞存活数量升高;PCMV-PRC1组细胞克隆形成能力升高。与阴性对照组比较,PCMV-PRC1组p-ERK1/2蛋白水平升高,cleaved PARP、cleaved caspase-3蛋白水平明显降低。PCMV-PRC1组同时加入特异性的ERK抑制剂U0126后,p-ERK1/2蛋白水平降低,cleaved PARP、cleaved caspase-3蛋白水平升高。结论PRC1通过激活MAPK信号途径促进A549细胞对顺铂耐药。

Objective To explore the relationship between PRC1 and cisplatin resistance in lung cancer and its possible mechanism.Methods Western blotting was used to detect the expression of PRC1 protein in A549 cells treated with different cisplatin concentrations at different times.The A549 cell lines with stable interference and overexpression of PRC1 were constructed by lentivirus transfection(shPRC1 group)and PCMV-PRC1 overexpression plasmid transfection(PCMV-PRC1 group).CCK-8 method and clone formation experiment were used to detect the effect of different PRC1 expression on the killing effect of cisplatin.Western blotting was used to detect the protein levels of p-ERK1/2,cleaved PARP and cleaved caspase-3 in shPRC1 group and PCMV-PRC1 group.Results The expression of PRC1 protein increased with the increase of cisplatin concentration,and increased with the extension of action time.The expression of PRC1 in A549/DDP cells was significantly higher than that in A549 cells(4.04±0.50 vs.0.99±0.13,P<0.01).Compared with the negative control group,the number of cells in shPRC1 group decreased,while that in PCMV-PRC1 group increased;the ability of cell clone formation in PCMV-PRC1 group increased.Compared with the negative control group,the protein level of p-ERK1/2 in PCMV-PRC1 group increased,while the protein level of cleaved PARP and cleaved caspase-3 decreased significantly.In PCMV-PRC1 group,the protein level of p-ERK1/2 decreased and the protein levels of cleaved PARP and cleaved caspase-3 increased after the addition of specific ERK inhibitor U0126.Conclusion PRC1 promotes cisplatin resistance of A549 cells by activating MAPK signaling pathway.