BC200通过介导脑胶质瘤细胞干性的维持抑制替莫唑胺的促凋亡作用

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摘要目的：探讨血清长链非编码RNA BC200在脑胶质瘤干细胞（GSCs）中的作用及相关作用机制。方法：神经干细胞培养液培养脑胶质瘤U87 MG细胞，并提取成球生长的GSCs；采用免疫荧光法检测GSCs干细胞标志性分子CD133和Nestin；小分子RNA（siRNA）转染沉默GSCs中BC200表达；替莫唑胺（TMZ）200 μmol·L-1处理GSCs 24 h，光镜下观察TMZ对GSCs悬浮细胞球形态的影响；采用Western blot方法检测CD133、Bax、Bcl-2和cleaved caspase-3的表达变化。结果：分离提取的GSCs球状生长，细胞球表面CD133和Nestin呈阳性表达。TMZ处理GSCs 24 h，GSCs成球能力略微下降，未见细胞球状状态明显离散，Bax、cleaved cas-pase-3、Bcl-2未有明显变化（均P＞0.05）。沉默BC200后GSCs CD133表达显著下降（P＜0.05）。BC200沉默后，TMZ处理GSCs发现其成球能力明显减弱、细胞分散；cleaved caspase-3和Bax分别上调（45.36%±4.25%）和（63.23%±5.12%），Bcl-2下调（31.22%±3.80%），均P＜0.01。结论：BC200能够维持脑胶质瘤U87 MG细胞的干性，该效应可促进细胞对TMZ诱导凋亡的抵抗。

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关键词 ： BC200, 脑胶质瘤, 干性, 替莫唑胺, 干细胞

Abstract：Objective: To investigate the role of serum long-chain non-coding RNA BC200 (lncRNA BC200) in glioma stem cells (GSCs) and related mechanism. Methods: Glioma U87 MG cells were cultured in neural stem cell culture medium and extracted into spherical growth GSCs. Immunofluorescence was used to detect the marker molecules CD133 and nestin of GSCs stem cells. Small RNA (siRNA) transfection silenced BC200 expression in GSCs. The GSCs were treated with Temozolomide (TMZ, 200 μmol·L-1) for 24 h, and the effect of TMZ on the morphology of suspended cells in GSCs was observed under light microscope. The expression of CD133, Bax, Bcl-2 and cleaved caspase-3 was detected by Western blot. Results: The extracted GSCs were isolated for spheroid growth, and the cell surface CD133 and nestin were positively expressed. After TMZ treatment of GSCs for 24 h, the ability of GSCs to form spheres did not change significantly, and the Bax, cleaved caspase-3 and Bcl-2 did not change significantly at 24 h. The expression of CD133 in GSCs was significantly decreased after silencing BC200. After BC200 silencing, TMZ treatment of GSCs showed that its ability to form a ball was significantly weakened, and the cells were dispersed. At 24 h, cleaved caspase-3 and Bax were up-regulated (45.36±4.25)% and (63.23±5.12)% (P<0.01). Bcl-2 was down-regulated (31.22±3.80)% (P<0.01). Conclusion: BC200 is able to maintain the dryness of glioma U87 MG cells, which promotes cell resistance to TMZ-induced apoptosis.

Key words： BC200 glioma stem cell characteristics temozolomide stem cell

收稿日期: 2018-11-12

基金资助:浙江省科技计划项目（2013C33103）。

作者简介: 童民锋（1977-），男，浙江金华人，副主任医师。

链接本文:

https://xb.wmu.edu.cn/CN/ 或 https://xb.wmu.edu.cn/CN/Y2019/V49/I7/529

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