FasL的特异性miRNA阻断舌鳞癌免疫逃逸的研究

发布时间：2018-04-01 12:26

本文选题：FasL　切入点：miR-590　出处：《安徽医科大学》2017年硕士论文

【摘要】：目的通过生物信息学预测并找出确实针对Fas L的特异性Micro RNA(miRNA),研究其对舌鳞癌细胞SCC-3 Fas配体(Fas Lignad,Fas L)的表达的影响,并最终验证其对舌鳞癌免疫逃逸的影响,从而了解肿瘤的发生发展和调控机制。方法将合成的miR-590 mimics或miR-590的抑制剂(inhibior)转染至SCC-3细胞。通过反转录PCR(reverse transcription-PCR,RT-PCR)和蛋白质印迹法(Western Blot)检测SCC-3中Fas L信使RNA(Messenger RNA,m RNA)和蛋白的表达;将转染后的SCC-3细胞在DDP(5ug/ml)处理下,在不同时间段通过四甲基偶氮唑蓝(MTT)法绘制细胞生长曲线,并通过针对Fas L的miR-590 inhibior从反面验证miR-590的作用。结果将miR-590 mimics转染至舌鳞癌细胞SCC-3,转染48小时后提取细胞总RNA,检测该组与对照组SCC-3细胞中Fas L m RNA和蛋白的表达情况后发现,miR-590可以在m RNA和蛋白水平上降低SCC-3细胞Fas L的表达。而将miR-590inhibitor转染至SCC-3,检测Fas L m RNA和蛋白的表达情况可发现,与对照组相比,表达miR-590 inhibitor的SCC-3细胞中Fas L m RNA和蛋白水平显著升高,增强了Fas L的转录。SCC-3细胞转染miR-590后按时间梯度加入DDP检测可发现,实验组细胞存活率低于对照组;转染miR-590 inhibitor反向验证可发现实验组细胞存活率高于对照组。结论miR-590可以有效抑制SCC-3细胞Fas L的表达,降低SCC-3的DDP的耐药性,从而可能降低和减少其对免疫活性细胞的死亡诱导和杀伤能力,并最终阻断了癌症细胞躲避免疫系统杀伤作用的途径,为舌鳞癌的生物治疗提供了新的靶点。
[Abstract]:Objective to predict and find out the specific Micro RNAs specific to Fas L by bioinformatics, to study its effect on the expression of SCC-3 Fas ligand FAS Lignadfas L in tongue squamous cell carcinoma, and to verify its effect on the immune escape of tongue squamous cell carcinoma.In order to understand the tumorigenesis, development and regulatory mechanisms.Methods miR-590 mimics or miR-590 inhibitor was transfected into SCC-3 cells.The expression of Fas L messenger RNA(Messenger RNAs and proteins in SCC-3 were detected by reverse transcription PCR(reverse transcription-PCRR-RT-PCRand Western blot.The transfected SCC-3 cells were treated with DDPn5ugrml. the growth curves of the transfected SCC-3 cells were plotted by tetramethyl azolium blue at different time intervals.The function of miR-590 is verified by miR-590 inhibior of Fas L.Results miR-590 mimics was transfected into tongue squamous cell SCC-3, and total RNAs were extracted 48 hours after transfection. The expression of Fas L m RNA and protein in SCC-3 cells was detected. It was found that miR-590 could decrease Fas L expression in SCC-3 cells at the level of m RNA and protein.When miR-590inhibitor was transfected into SCC-3, the expression of Fas LM RNA and protein was detected. Compared with the control group, the level of Fas LM RNA and protein in the SCC-3 cells expressing miR-590 inhibitor was significantly higher than that in the control group.After enhanced Fas L transcription. SCC-3 cells were transfected with miR-5 90. The survival rate of the experimental group was lower than that of the control group, and the survival rate of the experimental group was higher than that of the control group.Conclusion miR-590 can effectively inhibit the expression of Fas L in SCC-3 cells and reduce the drug resistance of SCC-3 DDP, which may reduce the death induction and killing ability of SCC-3 cells.Finally, it blocked the way of cancer cells avoiding the killing effect of immune system, and provided a new target for the biotherapy of tongue squamous cell carcinoma.
【学位授予单位】：安徽医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R739.86

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