生物钟基因PER1对口腔鳞癌细胞中生物钟基因网络的调控作用

发布时间：2018-05-05 04:22

本文选题：生物钟 + 癌　；参考：《重庆医科大学》2017年硕士论文

【摘要】：目的:探讨口腔鳞癌细胞内生物钟基因PER1对生物钟基因网络中其它生物钟基因表达的影响。方法:通过RNA干扰技术沉默PER1并构建3组PER1短发夹(short hairpin)RNA慢病毒载体质粒,然后分别感染SCC15口腔鳞癌细胞株,经Western blot和实时荧光定量PCR(Quantitative real-time PCR,qRT-PCR)筛选并确定PER1沉默效果最佳组定为实验组,以转染scramble质粒(不含任何干扰基因片段)的SCC15细胞为阴性对照组,以相同培养条件下不曾做处理的SCC15细胞为空白对照组。分别在体外和体内应用qRT-PCR检测PER1沉默前后生物钟基因CLOCK、BMAL1、PER2、PER3、DEC1、DEC2、CRY1、CRY2、TIM、CKIε、RORα、NPAS2和REV-ERBαmRNA的表达情况,应用流式细胞仪检测口腔鳞癌细胞的增殖和凋亡水平的改变情况,并应用裸鼠体内成瘤实验观察SCC15癌细胞体内成瘤能力改变情况。结果:PER1基因沉默后,体内和体外癌细胞中生物钟基因PER2、DEC1、DEC2、CRY1、CRY2和NPAS2 mRNA的表达显著降低(P0.05),PER3、TIM、RORɑ和REV-ERBɑmRNA的表达显著升高(P0.05),而CLOCK、BMAL1和CKIεmRNA的表达无显著改变(P0.05)。PER1沉默之后,SCC15癌细胞的凋亡降低,增殖和体内成瘤加强。结论:生物钟基因PER1可以调控钟基因网络中众多其他钟基因如PER2、DEC1、DEC2、CRY1、CRY2、NPAS2、PER3、TIM、RORɑ和REV-ERBɑ的表达,PER1在机体的钟基因网络中具有十分重要的调控功能。PER1对癌症形成的作用并非单一通过对其下游基因的调控,而是同时通过对生物钟基因网络中诸多生物钟基因的异常调控所共同导致。
[Abstract]:Aim: to investigate the effect of clock gene PER1 on the expression of other clock genes in oral squamous cell carcinoma (OSCC) cells. Methods: PER1 was silenced by RNA interference technique and three groups of PER1 short hairpin)RNA vector plasmids were constructed and then infected with SCC15 oral squamous cell carcinoma cell lines. Western blot and real-time quantitative PCR(Quantitative real-time PCR qRT-PCR were used to screen and determine the best effect of PER1 silencing. SCC15 cells transfected with scramble plasmid (without any interfering gene fragments) were used as negative control group. The SCC15 cells which were not treated under the same culture conditions were used as the blank control group. In vitro and in vivo, qRT-PCR was used to detect the expression of circadian clock gene CLOCKC BMAL1, PER2PER3, DEC1, CRY2, CRY2, CRY2, CRY2, and REV-ERB 伪 mRNA, and flow cytometry was used to detect the changes of proliferation and apoptosis in oral squamous carcinoma cells, respectively, before and after PER1 silencing, and in vitro and in vivo, respectively, were used to detect the expression of REV-ERB 伪 -NPAS2 and REV-ERB 伪 mRNA in oral squamous carcinoma cells, respectively, before and after PER1 silencing. The tumorigenic ability of SCC15 cancer cells in vivo was observed by tumorigenesis test in nude mice. Results the expression of clock gene PER2 / DEC1 / CRY1CRY2 and NPAS2 mRNA in cancer cells decreased significantly after the gene was silenced. The expression of P0.05, PER3TIMROR and REV-ERB mRNA increased significantly, while the expression of CLO CKBMAL1 and CKI 蔚 mRNA did not change the apoptosis of SCC15 cells. Proliferation and tumorigenesis are enhanced. Conclusion: biological clock gene PER1 can regulate the expression of many other clock genes in the clock gene network, such as PER2DEC1, DEC1, CRY1, CRY2, NPAS2, PER3, TIMROR and REV-ERB. PER1 plays a very important regulatory role in the clock gene network. PER1 plays an important role in cancer formation. The regulation of its downstream genes, At the same time, it is caused by the abnormal regulation of many biological clock genes in the biological clock gene network.
【学位授予单位】：重庆医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R739.8

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