RNAi抑制△ANp63表达对头颈鳞癌细胞增殖的影响

目的 研究 △△Np63促进头颈鳞癌细胞增殖及存活的作用及相关分子机制.方法 筛选高表达△Np63 的头颈鳞癌细胞系,采用RNAi方法分别抑制△Np63及P63表达.RT-PCR、Western印迹、MTT法、流式细胞术等检测细胞增殖和细胞周期等生物学行为改变;并检测相关周期调控蛋白的表达.结果 △Np63在FaDu细胞中高表达,定位于细胞核.抑制△Np63及P63表达后,FaDu细胞增殖能力明显下降(t=3.66,P＜0.01),发生G1期细胞周期阻滞;CDK抑制物P21及P27蛋白表达上调,P53表达上调.结论 △Np63对FaDu细胞增殖及存活发挥主导作用,△Np63通过抑制p21、p27的表达,促进肿瘤细胞的生长和增殖.

Abstract：

Objective To investigate the functional role of △Np63, the main isoform of P63, in promoting cell proliferation and survival in head and neck squamous cell carcinoma (HNSCC). Methods △Np63 overexpressing cell line was selected from several HNSCC cell lines. Specific △Np63 siRNAs and p63 siRNAs were individually transfected into FaDu cells for 48h and 72h which overexpressing △Np63. The expression levels of △Np63 mRNA and protein were analyzed by RT-PCR,Western blot and immunohistochemistry respectively.The cell proliferation was determined by MTT assay.The distribution of cell cycle was assessed by flow cytometry.The protein expression level of P21, P27 and P53 were detected by Western blot. Results △Np63 is highly expressed in FaDu cell line, especially located in the nucleus.The growth rate of FaDu cells transfected with △Np63-siRNA and p63-siRNA was significantly decreased(t=3.66,P＜0.01),and cell cycle mainly arrested in G1 phase. Further studies show that the down-requlation of △Np63 results in the overexpression of P53 and upregulation of CKIs , including P21 and P27 at protein levels. Conclusion △Np63 inhibit the expression of p21 and p27 to promote the proliferation of HNSCC cells , playing a role in maintenance of cell proliferation and survival.