PRMT2高表达对人甲状腺癌TPC-1细胞增殖的影响及机制研究
发布时间：2018-11-07 17:58
【摘要】：目的:探讨蛋白精氨酸甲基转移酶2(protein arginine methyltransferase 2,PRMT2)高表达对甲状腺癌TPC-1细胞增殖的影响,并初步探讨其调控细胞周期蛋白D1(Cyclin D1,CCND1)的分子机制。方法:将已构建好的PRMT2慢病毒载体转染TPC-1细胞,构建TPC-1-PRMT2稳定细胞株,采用蛋白免疫印迹(Western Blot)鉴定甲状腺癌TPC-1-PRMT2稳定细胞株;设立实验组即四环素诱导组(+Dox),阴性对照组即未加四环素诱导组(-Dox)。以上述两组细胞为实验对象,分别采用MTT检测方法、克隆形成实验检测各组细胞的增殖及克隆形成能力;Western Blot检测甲状腺癌中高表达PRMT2后对p-Akt、p-GSK-3β、CCND1蛋白表达的影响,并对其影响甲状腺癌细胞增殖的分子机制进行初步探讨。免疫共沉淀实验判断在普通甲状腺癌TPC-1细胞中,PRMT2与TRβ的相互作用。结果:1.转染TPC-1细胞后获得TPC-1-PRMT2稳定细胞株,经四环素诱导后Western Blot检测到PRMT2-3Flag蛋白的表达;2.MTT检测及克隆形成实验表明,PRMT2高表达后可降低TPC-1细胞的增殖能力;3.Western Blot检测提示,PRMT2高表达后p-Akt、p-GSK-3β、CCND1的表达均减少。而p-ERK蛋白的表达未见明显减少。且不影响TRβ的表达。4.免疫共沉淀实验表明,在普通TPC-1细胞内,PRMT2与TRβ存在相互作用。结论:1.PRMT2高表达后抑制甲状腺癌TPC-1细胞的增殖。2.PRMT2高表达后抑制甲状腺癌TPC-1细胞的增殖,可能通过调控Akt/GSK 3β信号通路,下调细胞周期蛋白CCND1表达水平有关;3.在普通TPC-1细胞内,PRMT2能和TRβ发生相互作用。
[Abstract]:Aim: to investigate the effect of the overexpression of arginine methyltransferase 2 (protein arginine methyltransferase 2 (PRMT2) on the proliferation of thyroid carcinoma TPC-1 cells and the molecular mechanism of its regulation of cyclin D1 (Cyclin D1). Methods: the constructed PRMT2 lentivirus vector was transfected into TPC-1 cells to construct TPC-1-PRMT2 stable cell line. Western blot (Western Blot) was used to identify TPC-1-PRMT2 stable cell line of thyroid carcinoma. Establish experimental group, tetracycline induced group, (Dox), negative control group, that is, no tetracycline induced group (- Dox).) The above two groups of cells were used as experimental objects. The proliferation and clone forming ability of each group were detected by MTT assay and clone formation assay respectively. The effects of high expression of PRMT2 on the expression of p-Akttnp-GSK-3 尾 and CCND1 protein in thyroid carcinoma were detected by Western Blot, and the molecular mechanism of its influence on the proliferation of thyroid cancer cells was discussed. The interaction of PRMT2 and TR 尾 in normal thyroid carcinoma TPC-1 cells was determined by immunoprecipitation assay. The result is 1: 1. After transfection of TPC-1 cells, TPC-1-PRMT2 stable cell lines were obtained, and the expression of PRMT2-3Flag protein was detected by Western Blot induced by tetracycline, 2.MTT detection and clone formation assay showed that high expression of PRMT2 could reduce the proliferation ability of TPC-1 cells. The results of 3.Western Blot showed that the expression of p-Aktbumin p-GSK-3 尾 and CCND1 decreased after high expression of PRMT2. However, the expression of p-ERK protein was not significantly decreased. The expression of TR 尾 was not affected. 4. Immunoprecipitation assay showed that PRMT2 interacted with TR 尾 in normal TPC-1 cells. Conclusion: the high expression of 1.PRMT2 inhibits the proliferation of TPC-1 cells in thyroid carcinoma, and the high expression of 2.PRMT2 inhibits the proliferation of TPC-1 cells of thyroid carcinoma, which may be through the regulation of Akt/GSK 3 尾 signaling pathway. The down-regulation of cyclin CCND1 expression was related to the expression of cyclin CCND1. 3. In normal TPC-1 cells, PRMT2 can interact with TR 尾.
【学位授予单位】：南华大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R736.1
本文编号：2317133
[Abstract]:Aim: to investigate the effect of the overexpression of arginine methyltransferase 2 (protein arginine methyltransferase 2 (PRMT2) on the proliferation of thyroid carcinoma TPC-1 cells and the molecular mechanism of its regulation of cyclin D1 (Cyclin D1). Methods: the constructed PRMT2 lentivirus vector was transfected into TPC-1 cells to construct TPC-1-PRMT2 stable cell line. Western blot (Western Blot) was used to identify TPC-1-PRMT2 stable cell line of thyroid carcinoma. Establish experimental group, tetracycline induced group, (Dox), negative control group, that is, no tetracycline induced group (- Dox).) The above two groups of cells were used as experimental objects. The proliferation and clone forming ability of each group were detected by MTT assay and clone formation assay respectively. The effects of high expression of PRMT2 on the expression of p-Akttnp-GSK-3 尾 and CCND1 protein in thyroid carcinoma were detected by Western Blot, and the molecular mechanism of its influence on the proliferation of thyroid cancer cells was discussed. The interaction of PRMT2 and TR 尾 in normal thyroid carcinoma TPC-1 cells was determined by immunoprecipitation assay. The result is 1: 1. After transfection of TPC-1 cells, TPC-1-PRMT2 stable cell lines were obtained, and the expression of PRMT2-3Flag protein was detected by Western Blot induced by tetracycline, 2.MTT detection and clone formation assay showed that high expression of PRMT2 could reduce the proliferation ability of TPC-1 cells. The results of 3.Western Blot showed that the expression of p-Aktbumin p-GSK-3 尾 and CCND1 decreased after high expression of PRMT2. However, the expression of p-ERK protein was not significantly decreased. The expression of TR 尾 was not affected. 4. Immunoprecipitation assay showed that PRMT2 interacted with TR 尾 in normal TPC-1 cells. Conclusion: the high expression of 1.PRMT2 inhibits the proliferation of TPC-1 cells in thyroid carcinoma, and the high expression of 2.PRMT2 inhibits the proliferation of TPC-1 cells of thyroid carcinoma, which may be through the regulation of Akt/GSK 3 尾 signaling pathway. The down-regulation of cyclin CCND1 expression was related to the expression of cyclin CCND1. 3. In normal TPC-1 cells, PRMT2 can interact with TR 尾.
【学位授予单位】：南华大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R736.1
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相关硕士学位论文 前2条
1 卢甜;PRMT2高表达对人甲状腺癌TPC-1细胞增殖的影响及机制研究[D];南华大学;2015年
2 刘美萍;PRMT2下调NF-κB对甲状腺癌细胞增殖及凋亡的影响[D];南华大学;2011年
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