Zhang Chun-zhi, Kang Chun-sheng, Pu Pei-yu, Wang Guang-xiu, Jia Zhi-fan, Zhang An-ling, Han Lei, Xu Peng
Department of Neurosurgery, Tianjin Medical University General Hospital and Laboratory of Neuro-Oncology, Tianjin Neurological Institute, Tianjin 300052, China.
Zhonghua Zhong Liu Za Zhi. 2009 Oct;31(10):721-6.
To study the inhibitory effect of knocking down microRNA(miR)-221 and miR-222 on human glioma cell growth and its possible mechanism.
miRNA-221/222 antisense oligonucleotides (antisense miR221/222) were transfected into human glioma U251 cells by lipofectamine. Northern blot analysis was conducted to detect the mRNA expression of miR-221/222 in the control and transfected cell groups. The proliferation activity of cells was determined by MTT assay. Cell invasion ability was examined by transwell assay, and cell cycle kinetics and apoptosis were detected with flow cytometry. The expression of relevant proteins was analyzed by Western blotting. The therapeutic efficacy of antisense miR221/222 on the growth of xenograft tumors in nude mice were also observed.
In the antisense miR-221/222-transfected cells, the expression of miR-221/222 was significantly reduced; the cell invasion ability was suppressed, cell cycle was blocked at G(0)/G(1) phase, and apoptotic cells were increased. The growth of xenograft tumors treated with antisense miR-221/222 was also inhibited. In antisense miR-221/222 treated tumor cells, the expression of bcl-2 was down-regulated while connexin43, p27, PUMA, caspase-3, PTEN, TIMP3 and Bax up-regulated, and p53 expression not changed.
There is a significant inhibitory effect of antisense miR-221/222 on the growth of human glioma U251 cells. miR-221/222 may be considered as a candidate target for gene therapy of human gliomas.
研究敲低微小RNA（miR）-221和miR-222对人胶质瘤细胞生长的抑制作用及其可能机制。
采用脂质体法将miRNA-221/222反义寡核苷酸（反义miR221/222）转染到人胶质瘤U251细胞中。通过Northern印迹分析检测对照组和转染细胞组中miR-221/222的mRNA表达。采用MTT法测定细胞增殖活性。通过Transwell法检测细胞侵袭能力，用流式细胞术检测细胞周期动力学和凋亡情况。通过蛋白质印迹法分析相关蛋白的表达。还观察了反义miR221/222对裸鼠移植瘤生长的治疗效果。
在转染反义miR-221/222的细胞中，miR-221/222的表达显著降低；细胞侵袭能力受到抑制，细胞周期阻滞于G(0)/G(1)期，凋亡细胞增加。反义miR-221/222处理的移植瘤生长也受到抑制。在反义miR-221/222处理的肿瘤细胞中，bcl-2的表达下调，而连接蛋白43、p27、PUMA、半胱天冬酶-3、PTEN、TIMP3和Bax上调，p53表达未改变。
反义miR-221/222对人胶质瘤U251细胞的生长有显著抑制作用。miR-221/222可被视为人类胶质瘤基因治疗的候选靶点。
