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目的 miR-449b在多种肿瘤细胞中都处于低表达状态,是潜在的肿瘤抑制因子。本课题主要探究miR-449b表达对GBM细胞替莫唑胺(TMZ)治疗敏感性的的影响及内在的调控机制。方法选用两株GBM细胞,T98G和LN229,将miR-449b模拟剂转入这两种细胞中,采用MTT法、克隆集落法和免疫印迹法检测miR-449b表达对不同剂量TMZ处理后的GBM细胞的存活、增殖能力和凋亡水平的影响;同时检测miR-449b表达对替莫唑胺治疗下GBM细胞ATP水平的影响,以及将外源ATP脂质体转入后对细胞活力的影响。结果 MTT研究表明,增加miR-449b表达可降低替莫唑胺治疗下GBM细胞的存活能力,表现为肿瘤细胞活力下调、克隆集落形成能力减弱及Cleaved-PARP的蛋白水平增加;ATP检测结果发现,miR-449b表达可降低替莫唑胺应激下肿瘤细胞ATP水平,并且外源性ATP补充可明显增强替莫唑胺治疗下肿瘤细胞的活力。结论以上研究结果提示,miR-449b可通过加剧替莫唑胺治疗下GBM细胞ATP水平消耗,进而促进GBM细胞对替莫唑胺治疗的敏感性,有利于提高替莫唑胺对GBM的治疗效果及其在临床上的应用。
The purpose of miR-449b in a variety of tumor cells in a low expression state, is a potential tumor suppressor. This topic mainly explores the influence of miR-449b expression on the sensitivity of TMZ treatment in GBM cells and the underlying regulatory mechanism. Methods Two GBM cells, T98G and LN229, were selected and transfected into these two cell lines. MTT, cloning and Western blotting were used to detect the expression of miR-449b in TMZ-treated GBM cells The effect of miR-449b expression on the ATP level of GBM cells treated with temozolomide and the effect of transfection of exogenous ATP liposomes on cell viability were also examined. Results The results of MTT showed that the increase of miR-449b expression could reduce the viability of GBM cells treated with temozolomide. The results showed that the viability of GBM cells was down-regulated, the colony-forming ability and the protein level of Cleaved-PARP were increased. ATP- Expression decreased the ATP level in tumor cells under temozolomide stress, and exogenous ATP supplementation significantly enhanced the viability of tumor cells treated with temozolomide. Conclusions The above results suggest that miR-449b can enhance the sensitivity of GBM cells to temozolomide treatment by exacerbating the consumption of ATP in GBM cells treated with temozolomide and improve the therapeutic effect of temozolomide on GBM and its clinical application.