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目的探讨抑制α-烯醇化酶(ENO1)表达对胶质瘤细胞糖代谢和细胞生长的影响。方法在胶质瘤细胞U251中,利用siRNA抑制ENO1表达后,利用葡萄糖摄取和乳酸生成实验检测糖代谢水平的改变;利用3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide(MTT)和5-Ethynyl-2’-deoxyuridine(EDU)着色的方法检测细胞增殖和细胞周期的改变。Western blot活性分析抑制ENO1表达糖代谢标志基因Hexokinase 2(HK2)和Lactate dehydrogenase A(LDHA)基因的表达水平改变。结果成功筛选了抑制ENO1的siRNA片段。在抑制ENO1表达,胶质瘤U251细胞摄取葡萄糖的能力明显降低(P=0.023)以及乳酸生成能力明显下降(P=0.007)。进一步,MTT和EDU着色分析显示,在抑制ENO1表达后,细胞的增殖能力从第2天开始明显降低(P<0.05);除此之外,细胞周期G1/S转化能力明显抑制(P=0.0425)。机制分析显示,ENO1下调后糖代谢标志基因HK2和LDHA表达也明显下降。结论 ENO1作为候选癌基因,通过正调控葡萄糖代谢从而促进了胶质瘤细胞的生长。
Objective To investigate the effect of inhibiting the expression of ENO1 on the glucose metabolism and cell growth of glioma cells. Methods The inhibitory effect of ENO1 on the expression of ENO1 in glioma cell line U251 was investigated by using glucose uptake and lactate production to detect the changes of glucose metabolism. Using the method of 3- (4,5-dimethyl-2-thiazolyl) -2,5-diphenyl -2-H-tetrazolium bromide (MTT) and 5-Ethynyl-2’-deoxyuridine (EDU) staining were used to detect cell proliferation and cell cycle changes. Western blot analysis showed that the expression of Hexokinase 2 (HK2) and Lactate dehydrogenase A (LDHA) genes in ENO1 expression were inhibited. Results The siRNA fragment that inhibits ENO1 was successfully screened. Inhibition of ENO1 expression significantly decreased glucose uptake in glioma U251 cells (P = 0.023) and lactate production ability (P = 0.007). Further, the MTT and EDU staining analysis showed that after the inhibition of ENO1 expression, cell proliferation decreased significantly from day 2 (P <0.05); in addition, cell cycle G1 / S conversion was significantly inhibited (P = 0.0425 ). Mechanism analysis showed that ENO1 downregulated the expression of glucose metabolism marker genes HK2 and LDHA also significantly decreased. Conclusions ENO1, as a candidate oncogene, promotes glioma cell growth by positively regulating glucose metabolism.