Notch1信号途径在皮肤鳞癌SCL-1细胞中的激活及其对细胞周期的影响

Activation of Notch 1 signaling pathway in cutaneous squamous cell carcinoma SCL-1 cell line and its effect on the cell cycle

目的:研究Notch1及其下游靶基因Hes1在皮肤鳞状细胞癌(简称鳞癌)细胞株SCL-1细胞中的表达及上调Notch1表达对细胞周期的影响。方法:将Notch1真核表达载体通过脂质体2000转染SCL-1细胞,通过反转录(RT)-PCR和Westernblotting检测Notch1和Hes1基因在皮肤鳞癌SCL-1细胞中的表达,并利用CCK-8试剂分析上调Notch1表达对皮肤鳞癌SCL-1细胞增殖的影响,通过流式细胞仪观察Notch1过表达对细胞周期的影响。结果:SCL-1细胞中存在Notch1和Hes1基因表达,提示该信号途径在皮肤鳞癌细胞中处于激活状态,转染Notch1真核表达载体后,与未处理组和空载体组相比,转染组中Notch1和Hes1的mRNA和蛋白表达都明显增加(P<0.05)。此外,细胞增殖实验结果表明,过表达Notch1能明显抑制皮肤鳞癌SCL-1细胞增殖。流式细胞仪检测结果显示,Notch1过表达使细胞周期静止在G_0/G_1期。结论:Notch1过表达能引起皮肤鳞癌SCL-1细胞的细胞周期静止在G_0/G_1期,Notch1信号途径可能在皮肤鳞癌的进展中起作用。

Objective： To investigate the expression of Notch 1 and its down-stream target gene Hes 1 in cutaneous squamous cell carcinoma （CSCC） SCL-1 cell line and the effect of over-expression of Notch 1 on cell cycle of SCL-1. Methods： Notch 1 eukaryotic expression vector was transfected into SCL-1 cells by Lipofectamine 2000. The expression levels of Notch 1 and Hes 1 genes were detected by RT-PCR and Western blotting in SCL-1 cells. In addition,the assay of proliferation of SCL_cell line was performed using CCK-8 Kit. Further, the cell cycle distribution was measured by flow cytometry（FCM）. Results：The expression levels of Notch 1 and Hes 1 genes were found in SCL-1 cells, implying an activated status of Notch 1 signaling pathway in SCL-1. Meanwhile, the expres- sion levels of Notch 1 and Hes 1 mRNAs and proteins were obviously increased in SCL-1 cells transfected with Notch 1 eukaryotic expression vector compared to untreated and empty vector treated groups （P〈0.05）. In addition, the result of the cell proliferation ex- periment showed that over-expression of Notch 1 could markedly inhibit the cell proliferation in SCL-1 cells. Further, the result of flow cytometry revealed that over-expression of Notch 1 could arrest cell cycle at G0/G1 phase in SCL-1 cells. Conclusions：Over- expression of Notch 1 could arrest cell cycle at Go/G1 phase in SCL-1 cells, which suggests that Notch 1 gene may play an important role in inhibiting the proliferation of CSCC cell line.