共抑制RAD18和REV1基因显著增加顺铂对肺癌细胞的毒性

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Abstract Objective: To explore the effects of RAD18 and REV1 gene knockdowns on cisplatin cytotoxicity in cisplatinresistant A549/DDP cell line. Methods: The expression of RAD18 and REV1 protein, and the FANCD2 and PCNA monoubiquitination level were detected by Western blotting. The siRNAs against RAD18 and REV1 genes(RAD18siRNA and REV1siRNA) were transfected into A549/DDP cells using Lipofectamine. The CCK8 assay was used to detect the changes of cell proliferation rate following treatment with cisplatin before and after transfection of RAD18siRNA and REV1siRNA in A549/DDP cells. Cisplatininduced cell apoptosis and cell cycle were detected by flow cytometry in A549/DDP cells pre and posttransfection of RAD18siRNA and REV1siRNA. Results: The expressions of RAD18 and REV1 proteins were significantly decreased after transfection of RAD18siRNA or REV1siRNA, indicating that the transfections were successful. Cisplatininduced monoubiquitinations of FANCD2 and PCNA were significantly decreased after transfection with RAD18siRNA, suggesting that the activation of the FA pathway and the TLS pathway was suppressed. Individual knockdown of RAD18 or REV1 by siRNAs transfection enhanced the cytotoxicity of cisplatin to A549/DDP cells, promoted cell apoptosis induced by cisplatin, and resulted in the arrest of S/G2 phase. Coknockdown of RAD18 and REV1 genes further enhanced the cytotoxicity of cisplatin to A549/DDP cells compared to RAD18 or REV1 knockdown alone. Conclusion: Knockdowns of RAD18 gene of the TLS pathway and the Fanconi anemia（FA） pathway upstream and REV1 gene of the TLS pathway could increase the sensitivity of cisplatinresistant A549/DDP cells to cisplatin by inhibiting the DNA damage repair capacity,and the sensitization can be further potentiated by coknockdown of RAD18 and REV1.

Received: 27 December 2017

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http://zzs.ujs.edu.cn/xbyxb/EN/ OR http://zzs.ujs.edu.cn/xbyxb/EN/Y2018/V28/I02/109

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