miR-139-5p靶向调控DNMT1抑制胶质母细胞瘤的生长和转移

miR-139-5p targeting DNMT1 inhibits the growth and metastasis of glioblastoma

目的考察miR-139-5p在胶质母细胞瘤(glioblastoma, GBM)中的表达及生物学作用,以及miR-139-5p对DNA甲基转移酶1(DNMT1)的调控作用。方法通过qRT-PCR检测GBM肿瘤组织、配对癌旁组织、人正常神经胶质细胞系HEB和人神经胶质瘤细胞系U251中miR-139-5p的表达。通过转染miR-139-5p模拟物来上调U251细胞中miR-139-5p的表达,通过转染靶向DNMT1的siRNA(DNMT1-siRNA)来下调DNMT1的表达。qRT-PCR、Western blotting、免疫组织化学和免疫荧光检测组织和细胞中DNMT1和2型神经纤维瘤病(NF2)的表达。细胞计数试剂盒-8(CCK-8)、流式细胞术和Matrigel Transwell评估U251细胞的增殖、凋亡和侵袭能力。结果与癌旁组织或HEB细胞相比,GBM组织和U251细胞中的miR-139-5p表达被抑制(P<0.05)。与对照细胞相比,转染miR-139-5p模拟物明显下调了DNMT1的表达并上调了NF2的表达(P<0.05)。与对照细胞相比,转染DNMT1-siRNA明显促进了NF2的表达(P<0.05)。转染miR-139-5p模拟物或DNMT1-siRNA均可诱导U251细胞凋亡并抑制细胞侵袭(P<0.05)。结论 miR-139-5p在GBM中发挥抗癌作用,miR-139-5p通过靶向负调控DNMT1来抑制肿瘤的增殖和转移。

Objective To investigate the expression and biological role of miR-139-5 p in glioblastoma and the regulatory effect of miR-139-5 p on DNA methyltransferase 1(DNMT1). Methods qRT-PCR was used to detect the expression of miR-139-5 p in glioblastoma tumor tissue, paired paracancerous tissue, human normal glioma cell line HEB, and human glioma cell line U251. The expression of miR-139-5 p in U251 cells was up-regulated by transfection of miR-139-5 p mimetics, and the expression of DNMT1 was down-regulated by transfection of DNMT1-targeted siRNA(DNMT1-siRNA). The expression of DNMT1 and neurofibromatosis type 2(NF2) in tissues and cells was detected by qRT-PCR, Western blotting, immunohistochemistry and immunofluorescence. The cell counting kit-8(CCK-8), flow cytometry and Matrigel Transwell were used to evaluate the proliferation, apoptosis and invasion ability of U251 cells. Results Compared with paracancerous tissues or HEB cells, miR-139-5 p expression in glioblastoma tissues and U251 cells was suppressed(P<0.05). Compared with control cells, transfection of miR-139-5 p mimic significantly down-regulated the expression of DNMT1 and up-regulated the expression of NF2(P<0.05). Compared with control cells, transfection of DNMT1-siRNA significantly promoted the expression of NF2(P<0.05). Transfection of miR-139-5 p mimetics or DNMT1-siRNA significantly induced U251 cell apoptosis and inhibited cell invasion(P <0.05).Conclusion miR-139-5 p plays an anti-cancer role in glioblastoma,and it inhibits tumor proliferation and metastasis by targeting negative regulation of DNMT1.