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作 者:梁书卿 魏枫[1] 孙洪莉 郑朦 王晓艳 李冉浩 马玉博 Liang Shuqing;Wei Feng;Sun Hongli(Dept of Endocrinology,the First Affiliated Hospital of Baotou Medical College,Inner Mongolia University of Science and Technology,Baotou 014010)
机构地区:[1]内蒙古科技大学包头医学院第一附属医院内分泌科,包头014010
出 处:《安徽医科大学学报》2020年第12期1861-1865,共5页Acta Universitatis Medicinalis Anhui
基 金:内蒙古自治区科技计划项目(编号:201802128)。
摘 要:目的探讨DNA甲基化在甲状腺乳头状癌中的作用机制,使用去甲基化药物5-氮杂-2′-脱氧胞苷酸(5-Aza-CdR)对死亡相关蛋白激酶(DAPK)基因表达水平的影响。方法将10μmol/L的去甲基化制剂5-Aza-CdR分别应用于人甲状腺乳头状癌TPC-1细胞株及人正常甲状腺Nthy-ori-3细胞株,Real-timePCR和Westernblot法检测用药前后DAPK、DNA甲基转移酶(DNMTs)mRNA和蛋白的表达情况。结果Real-timePCR结果显示,与人正常甲状腺细胞相比,人甲状腺乳头状癌中DNMTsmRNA高表达;人甲状腺乳头状癌细胞应用5-Aza-CdR治疗后,DNMTsmRNA的表达量减少,而DAPK的mRNA表达量增加,差异有统计学意义(P<0.05);Westernblot结果显示,与人正常甲状腺细胞相比,甲状腺乳头状癌中DNMTs蛋白的表达高,DAPK蛋白表达少,经5-Aza-CdR作用48h后,DNMTs蛋白表达降低,DAPK蛋白表达升高(P<0.05)。甲状腺正常细胞中不论给药前后,DAPK及DNMTs的mRNA及蛋白表达均无差异。结论DNA甲基化在甲状腺乳头状癌中扮演重要的角色,5-Aza-CdR可能通过调节DNMTs的mRNA及蛋白的表达使沉默的DAPK基因恢复表达,从而发挥对甲状腺乳头状癌的防治作用。Objective To investigate the mechanism of DNA methylation in papillary thyroid carcinoma, effects of the demethylated drug 5-aza-2′-deoxycytidine(5-Aza-C dR) on the expression level of death-associated protein kinase(DAPK) genes.Methods Human thyroid papillary carcinoma cell line TPC-1 and human normal thyroid cell line Nthy-ori-3 were treated with 10 μmol/L 5-Aza-CdR. Real-Time PCR and Western blot were used to detect the expression levels of DAPK, DNA methyltransferases(DNMTs)mRNA and protein before and after treatment.Results Real-Time PCR results showed that DNMTs mRNA was highly expressed in human thyroid papillary carcinoma cells compared with normal thyroid cells. After 5-Aza-CdR treatment in human thyroid papillary cancer cells, the expression level of DNMTs mRNA decreased, while that of DAPK mRNA increased, and there was significant difference(P<0.05);Western blot results showed that the expression of DNMTs protein was higher in human thyroid papillary carcinoma and DAPK protein expression was less compared with normal thyroid cells, after 5-Aza-CdR was applied for 48 h, the expression of DNMTs protein decreased and the expression of DAPK protein increased(P<0.05). Before and after administration, mRNA and protein expressions of DAPK and DNMTs in normal thyroid cells were not different.Conclusion DNA methylation plays an important role in papillary thyroid carcinoma, 5-Aza-CdR may regulate the expressions of DNMTs mRNA and protein to restore the expression of silenced DAPK gene, which may play a preventive role in thyroid papillary carcinoma.
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