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目的:检测不同前列腺癌(PCa)细胞系及组织中S期激酶相关蛋白(Skp2)的表达水平,探讨其表达改变对雄激素受体(AR)信号通路及去势抵抗型前列腺癌(CRPC)发生可能的影响。方法:通过Western印迹法检测不同PCa细胞系中Skp2、AR表达;利用RNA干扰技术,转染shRNA敲低CRPC细胞系C4-2或22RV1中Skp2表达,通过Western印迹法检测雄激素处理后细胞中AR、P27表达,通过双荧光素酶报告基因法检测雄激素处理后细胞中雄激素反应元件(ARE)报告基因ARR3-Luc活性;利用免疫组化染色法检测未行内分泌治疗的患者PCa和CRPC组织标本中Skp2、AR表达,分析两者表达差异及相关性。结果:CRPC细胞系C4-2及22RV1较雄激素依赖性细胞系LNCa P中Skp2表达水平显著升高;C4-2细胞中雄激素处理可诱导Skp2表达,而shRNA敲除Skp2不仅可显著上调其下游经典靶分子P27蛋白的表达,还显著降低AR蛋白表达水平。相一致地,C4-2及22RV1细胞系中雄激素处理可增强ARR3-Luc活性,而敲除Skp2可显著抑制雄激素处理前或后的ARR3-Luc活性(P<0.05)。此外,CRPC组织较未行内分泌治疗患者PCa组织中Skp2、AR染色显著增强(P<0.05),两者表达具有正相关性(r=0.658 1,P<0.05)。结论:CRPC中Skp2可增强AR蛋白表达及转录活性,有望成为重要的分子治疗靶点。
OBJECTIVE: To detect the expression of S-phase kinase-related protein (Skp2) in different prostate cancer (PCa) cell lines and tissues, and to explore the effect of its expression on the expression of androgen receptor (AR) Possible impact. Methods: The expression of Skp2 and AR in different PCa cell lines was detected by Western blotting. The expression of Skp2 in CRPC cell line C4-2 or 22RV1 was knocked down by RNA interference and the expression of Skp2 was detected by Western blotting in the cells treated with androgen AR and P27 were detected by enzyme-linked immunosorbent assay (ELISA). ARR3-Luc activity of androgen receptor (ARE) reporter gene in androgen-treated cells was detected by dual luciferase reporter assay. Immunohistochemical staining was used to detect PCa and CRPC The expression of Skp2 and AR in the tissue samples was analyzed, and the difference and correlation between them were analyzed. Results: The expression of Skp2 in CRPC cell lines C4-2 and 22RV1 was significantly higher than that in androgen-dependent cell line LNCa P, while the expression of Skp2 was induced by androgen treatment in C4-2 cells. Downstream classical target molecule P27 protein expression, but also significantly reduce the AR protein expression level. Consistently, androgen treatment in C4-2 and 22RV1 cell lines enhanced ARR3-Luc activity, whereas knockdown of Skp2 significantly inhibited ARR3-Luc activity (P <0.05) before and after androgen treatment. In addition, the expression of Skp2 and AR in PCa tissues of patients with CRPC was significantly higher than that of patients without endocrine therapy (P <0.05), and there was a positive correlation between them (r = 0.658 1, P <0.05). Conclusion: Skp2 in CRPC can enhance AR protein expression and transcription activity, which is expected to be an important target of molecular therapy.