HOXA4基因抑制对胶质瘤细胞U251的影响

The influence of HOXA4 gene inhibition in U251 glioma cells

目的探讨HOXA4在人胶质瘤中的表达水平以及对胶质瘤细胞增殖、侵袭、凋亡的影响。方法通过生物信息学分析HOXA4在人胶质瘤中表达情况。选取胶质瘤细胞U251转染HOXA4 siR NA作为si-HOXA4组,抑制HOXA4基因的表达,同时转染阴性对照慢病毒载体作为si-NC组。q RT-PCR和Western blot检测转染后HOXA4基因和蛋白的表达水平。MTT实验、细胞克隆实验、Transwell实验和流式细胞术分别检测抑制HOXA4基因表达对细胞增殖、侵袭、周期和凋亡的影响。Western blot分析HOXA4下游凋亡蛋白P53、Bcl-2、Cytochrome C、Caspase-3的表达。结果与正常脑组织比较,人胶质瘤中HOXA4表达水平明显升高(P<0.001)。与si-NC组比较,si-HOXA4组U251细胞的OD值从转染后第4天明显下降(P<0.01),第7天下降更为显著(P<0.001)。si-HOXA4组U251细胞较si-NC组侵袭细胞数明显减少(P<0.01),G0/G1期比例显著升高(P<0.05),凋亡率明显升高(P<0.05),凋亡蛋白P53、Cytochrome C、Caspase-3表达明显升高,Bcl-2明显下降(P<0.05)。结论HOXA4基因在人胶质瘤中高表达,可能通过影响细胞增殖、侵袭、凋亡等生物功能对胶质瘤的发生发展起重要作用。

Objective To explore the expression of HOXA4 in human glioma and the influence of HOXA4 on proliferation, invasion and apoptosis of glioma cells. Methods Bioinformatic analysis was performed to investigate the expression of HOXA4 in human glioma. Glioma cell line U251 was transfected with HOXA4 siR NA lentiviral vectors as si-HOXA4 group to inhibit the expression of HOXA4 and with negative control siR NA lentiviral vectors as si-NC group. The expressions of HOXA4 mR NA and protein were detected by q RT-PCR and Western blot in the two groups. The effects of HOXA4 on proliferation, invasion, apoptosis and cell cycle were analyzed by MTT assay, colony formation assay, Transwell assay and flow cytometry assay. The expression of apoptosis-related proteins including P53, Bcl-2, Cytochrome C, Caspase-3 were analyzed by Western blot. Results Compared with normal brain tissue,HOXA4 expression was obviously elevated in human glioma（P 〈 0.001）. Compared with si-NC group, the OD value of si-HOXA4 group decreased 4 d after transfection（P 〈 0.01） and more significantly decreased 7 d after transfection（P 〈 0.001）, the number of invasive cells was decreased（P 〈 0.01）, the G0/G1 ratio was increased（P 〈 0.05）, apoptosis rate was higher（P 〈 0.05）, the expressions of P53, Cytochrome C and Caspase-3 were significantly increased and Bcl-2 was decreased significantly（ P 〈 0.05）. Conclusion HOXA4 expression is significantly higher in human glioma, and may play important role in the development and progression of glioma cells by affecting cell proliferation, invasion and apoptosis.