肝再生磷酸酶3对肾透明细胞癌细胞A-498的影响

目的]研究肝再生磷酸酶3 (phosphatase of regenerating liver-3,PRL-3)对肾透明细胞癌(clear cell renal cell cancinoma.CCRCC)细胞A-498的影响.方法]本实验以肾透明细胞癌细胞A-498为实验对象,采用siRNA干涉的方法下调A-498中PRL-3表达水平,以不做任何处理为空白对照组,脂质体加随机序列的siRNA为阴性对照组,脂质体加siRNA处理组为实验组,之后通过RT-PCR及Western blotting方法检测siRNA干涉效率,绘制细胞生长曲线、流式细胞仪检测细胞周期及凋亡以观察下调PRL-3对A-498细胞的影响.结果]同阴性对照组相比,本实验所设计的siRNA能够下调PRL-3在A498细胞中的表达,其中siRNA2干涉效率更高,细胞生长曲线显示自siRNA转染60h开始,下调PRL-3能够抑制细胞的生长(P＜0.01),流式细胞仪检测发现转染72h后,下调PRL-3的表达可以促进A-498细胞G1期阻滞,S期减少,凋亡增加(P＜0.05).结论]下调PRL-3能够明显抑制A498细胞的增殖,PRL-3很可能成为肾透明细胞癌基因治疗的新靶点.

Analysis on effect of phosphatase of regenerating liver-3 in clear cell renal cell carcinoma cell line-A-498

Objective] To investigate the function of phosphatase of regenerating liver-3 （PRL-3） in clear cell renal cell carcinoma （CCRCC） cell line-A-498. Methods] In this research, clear renal cell carcinoma cell line-A-498 was used as the experimental subject. PRL-3 siRNA was designed and transfected into A-498 cell（the transfection was performed by using LipotectamineTM 2000 ）. lntreated Cells served as a blank control, cells treated with lipofec, tamine and random sequence siRNA served as negative control, and cells tl~ated with lipotectamine and siRNA served as expremental group. The siRNA interfering effects were detected on mRNA and protein levels by RT-PCR and Western blotting respectively. The affection of the siRNA on cell proliferation was el）served through cell growth curve. The variation of cell cycle and apoptosis were detected by Flow cytometry. Result] The results of RT-PCR and Western blotting indicated that the siRNA could down-regulate the expression of PRL--3 in A498 cell properly, and tile siRNA2 was more effectively. Cell growth curves indicated that down-regulation of PRL-3 conld suppress the proliferation of the A-498 cells 60 h after transfection as compared with the negative control （P 〈 0.01 ）. siRNA-transfected cells showed an increase in G1 phase and a decrease in S phase after transfection （ P 〈 0.05 ）as compared with the negative control. PRL-3 siRNA induced apoptosis increased in comparison with the control. Conclusion ] Down-regulation of PRL-3 can suppress the proliferation of the A498 cells significantly. These results indicate that PRL-3 may become a new target gene for CCRCC therapy.