沉默生物钟基因PER1对口腔鳞状细胞癌细胞中生物钟基因网络的影响

Effect of clock gene PER1 knockdown on clock gene networks in human oral squamous cell carcinoma

目的探讨口腔鳞状细胞癌细胞中生物钟基因PER1对生物钟基因网络中其他生物钟基因表达的影响。方法采用RNA干扰技术沉默人口腔鳞状细胞癌SCC15细胞中PER1基因,应用流式细胞仪检测沉默后细胞的增殖和凋亡水平,实时荧光定量聚合酶链式反应检测生物钟基因CLOCK、BMAL1、PER1、PER2、PER3、DEC1、DEC2、CRY1、CRY2、TIM、CKIE、RORA、NPAS2、REV-ERBA m RNA的表达情况。结果 PER1基因沉默后,SCC15细胞增殖指数上升,凋亡指数下降(P<0.05);PER1、PER2、DEC1、DEC2、CRY1、CRY2和NPAS2 m RNA的表达水平降低(P<0.05),PER3、TIM、RORA和REV-ERBA m RNA的表达水平升高(P<0.05),CLOCK、BMAL1和CKIE m RNA的表达水平无统计学改变(P>0.05)。结论生物钟基因PER1能够调控生物钟基因网络中众多其他生物钟基因PER2、DEC1、DEC2、CRY1、CRY2、NPAS2、PER3、TIM、RORA和REV-ERBA的表达,PER1在生物钟基因网络中具有重要调控作用。

Objective This study investigated the effect of clock gene PER1 on the expression levels of other clock genes in clock gene networks in oral squamous cell carcinoma cells. Methods We used RNA interference mediated by short hairpin RNAs （shRNAs） to effectively knock down PER1 in SCC15 human oral squamous cell carcinoma cells. Flow cytometry was used to detect the degree of proliferation and apoptosis of the cells after PER1 knockdown, and quantitative real-time PCR was used to detect the mRNA expression levels of the clock genes CLOCK, BMAL1, PER1, PER2, PER3, DEC1, DEC2, CRY1, CRY2, TIM, CKIE, RORA, NPAS2, and REV-ERBA. Results The proliferation index of SCC15 cells increased significantly while the apoptotic index decreased significantly after PER1 knockdown （P〈0.05）. The mRNA expression levels of PER1, PER2, DEC 1, DEC2, CRY1, CRY2, and NPAS2 markedly decreased （P〈0.05） while those of PER3, TIM, RORA, and REV-ERBA markedly increased （P〈0.05）. By contrast, no obvious changes were observed in the mRNA expression levels of CLOCK, BMAL 1, and CKIE （P〉0.05）. Conclusion The clock gene PER1 can regulate the expression levels of other clock genes in the clock gene networks; these genes include PER2, DEC1, DEC2, CRY1, CRY2, NPAS2, PER3, TIM, RORA, and REV-ERBA. PER1 gene thus plays an important role in the regulation of clock gene networks.