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目的 Hedgehog信号通路参与了肿瘤的发生发展,本研究探讨该通路关键信号分子Smoothened(SMO)在肾透明细胞癌中的表达及对肾癌细胞增殖和凋亡的影响。方法选取2012-01-01-2013-06-30青岛大学附属医院泌尿外科,行手术治疗的80例肾透明细胞癌患者的临床及病理资料,采用免疫组织化学方法检测SMO在肾透明细胞癌组织中的表达并分析其与临床病理特征间的关系。采用小干扰RNA下调SMO在人肾癌细胞786-O中的表达,分别应用CCK-8法、流式细胞术及蛋白质印迹法检测下调SMO表达对细胞增殖、凋亡及Gli1和Gli2表达的影响。结果 SMO在73例(91.25%)肾透明细胞癌组织中有表达,其在高级别肾癌中表达(80.49%)较低级别(56.41%)显著升高,χ2=5.39,P=0.02。RT-PCR检测结果显示,SMO在人肾癌细胞系786-O中表达量为0.704±0.059;蛋白质印迹结果显示,SMO在786-O中表达量为0.651±0.074。在786-O细胞中应用小干扰RNA沉默SMO表达后,实验组细胞相较于对照组其活力百分比在48、72和96h分别为92.7%、80.9%和79.9%,3个时间点差异有统计学意义(P=0.003),细胞凋亡显著增加,t=-29.2,P<0.001;与空白对照组和阴性对照组相比,其下游分子Gli1和Gli2蛋白表达明显减少(Gli1:3.2 vs 2.9 vs 1;Gli2:2.5 vs 2.1vs 1)。结论 SMO可能通过调控细胞增殖和凋亡,以及调节Gli蛋白表达参与了肾癌的发生发展。
Objective Hedgehog signaling pathway is involved in tumorigenesis. This study was aimed to investigate the expression of Smoothened (SMO), a key signaling pathway in renal clear cell carcinoma, and its effect on proliferation and apoptosis of renal cell carcinoma. Methods 2012-01-01-2013-06-30 Department of Urology, Affiliated Hospital of Qingdao University, surgical treatment of 80 cases of clear cell renal cell carcinoma of the clinical and pathological data using immunohistochemical detection of SMO in renal clear cell carcinoma In the expression and analysis of the relationship between clinicopathological features. Small interfering RNA (siRNA) down-regulated the expression of SMO in 786-O cells. CCK-8 assay, flow cytometry and Western blotting were used to detect the effect of SMO expression on cell proliferation, apoptosis and Gli1 and Gli2 expression . Results The expression of SMO was detected in 73 cases of clear cell renal cell carcinoma (91.25%). The expression of SMO in high grade renal cell carcinoma (80.49%) was significantly higher than that of lower grade (56.41%) (χ2 = 5.39, P = 0.02). The results of RT-PCR showed that the expression of SMO in human renal cell carcinoma cell line 786-O was 0.704 ± 0.059; Western blotting showed that the expression of SMO in 786-O was 0.651 ± 0.074. After SMO siRNA was silenced in 786-O cells, the percentage of viability of SMO cells in experimental group was 92.7%, 80.9% and 79.9% at 48, 72 and 96 h, respectively, with statistical difference at 3 time points (P = 0.003), the apoptosis increased significantly (t = -29.2, P <0.001). Compared with the blank control group and the negative control group, the expression of Gli1 and Gli2 protein was significantly decreased (Gli1: 3.2 vs 2.9 vs 1; Gli2: 2.5 vs 2.1 vs 1). Conclusion SMO may participate in the development of renal cell carcinoma by regulating cell proliferation and apoptosis and regulating the expression of Gli protein.