目的 探讨大剂量维生素 C 对肺癌 A549 细胞增殖及葡萄糖摄取的作用。 方法 应用 CCK8 法分析维生素 C 作用于 A549 细胞的 IC50 值;不同浓度的维生素 C(0~ 8 mmol / L)和葡萄糖(5 mmol / L 和 25 mmol / L)干预 A549 细胞 24 h,CCK8 法检 测细胞增殖;8 mmol / L 维生素 C 干预 A549 细胞 4 h,采用 FRASC 抗坏血酸试剂盒Ⅱ检测细胞内维生素 C 浓度,以人支气管上 皮样细胞(HBE 细胞)作为对照,分析细胞内维生素 C 浓度的时间变化;然后将 A549 细胞分为 5 mmol / L 葡萄糖、5 mmol / L 葡 萄糖 + 8 mmol / L 维生素 C、25 mmol / L 葡萄糖、25 mmol / L 葡萄糖 + 8 mmol / L 维生素 C 四个组干预 24 h,检测细胞内维生素 C 浓度,以荧光标记的 2-脱氧葡萄糖作为探针检测细胞内葡萄糖摄取量,qPCR 和蛋白质印迹法检测葡萄糖转运体 1 和 3 (GLUT1 和 GLUT3)的表达水平。 结果 维生素 C 作用于 A549 细胞 24 h 的 IC50 为 5. 192 mmol / L;维生素 C 抑制 A549 细胞增 殖的作用随剂量增加而增强(P<0. 05),且维生素 C 浓度为 4、8 mmol / L 时,5 mmol / L 葡萄糖干预后细胞活力低于 25 mmol / L 葡萄糖(P<0. 05);A549 细胞内维生素 C 浓度随时间而增加,且在 0. 5、1、2、4 h 时,均显著高于 HBE 细胞(P<0. 05);8 mmol / L 维生素 C 干预 24 h,A549 细胞内的葡萄糖摄取量降低(P<0. 05),且 25 mmol / L 葡萄糖干预后,细胞内维生素 C 浓度低于 5 mmol / L 葡萄糖(P<0. 05);葡萄糖浓度为 5 mmol / L 时,8 mmol / L 维生素 C 干预后 GLUT1、GLUT3 的 RNA 和蛋白表达水平均下降(P<0. 05),葡萄糖浓度为 25 mmol / L 时,GLUT3 的 RNA 水平和 GLUT1 的蛋白水平下降(P<0. 05)。 结论 低浓度葡萄糖时,高浓度维生素 C 可下调 GLUT1 和 GLUT3 表达,降低 A549 细胞内的葡萄糖摄取量,抑制 A549 细胞增殖。

Objective To investigate the effects of high-dose vitamin C on proliferation and glucose uptake in lung cancer A549 cells. Method The IC50 value of A549 cells treated with vitamin C was analyzed by CCK8 method. Different concentrations of vitamin C the final concentration was 0-8 mmol / L were applied to A549 cells for 24 hours and glucose concentrations were set at 5 mmol / L and 25 mmol / L respectively. Cell proliferation was detected by CCK8 method. A549 cells were interfered with 8 mmol / L vitamin C and intracellular vitamin C concentration was detected by FRASC Ascorbic acid kit Ⅱ. Human bronchial epithelioid cells HBE cell were used as a control to analyze the time change of intracellular vitamin C concentration for 4 h. A549 cells were then divided into four groups and interfered with 5 mmol / L glucose 5 mmol / L glucose + 8 mmol / L vitamin C 25 mmol / L glucose or 25 mmol / L glucose + 8 mmol / L vitamin C. Intracellular vitamin C concentration was detected and intracellular glucose intake was detected by using fluorescence-labeled 2-deoxyglucose as a probe. The expression levels of glucose transporters 1 and 3 GLUT1 and GLUT3 were detected by qPCR and Western Blot. Result The IC50 concentration of A549 cells treated with vitamin C for 24 h was 5. 192 mmol/ L. The inhibitory effect of vitamin C on A549 cell proliferation was enhanced with the increase of concentration P< 0. 05 and the cell viability of A549 cells after 5 mmol / L glucose intervention was lower than 25 mmol / L glucose when the concentration of vitamin C was 4 or 8 mmol / L P<0. 05 . Vitamin C concentration in A549 cells increased with time and was significantly higher than that in HBE cells at 0. 5 1 2 and 4 h P<0. 05 . After 8 mmol / L vitamin C intervention for 24 h the intracellular glucose intake of A549 cells was decreased P<0. 05 and after 25 mmol / L glucose intervention the intracellular vitamin C concentration was lower than 5 mmol/ L glucose P<0. 05 . When glucose concentration was 5 mmol / L the expression levels of RNA and protein of GLUT1 and GLUT3 were decreased after 8 mmol / L vitamin C intervention P< 0. 05 while the RNA level of GLUT3 and the protein level of GLUT1 were decreased when glucose concentration was 25 mmol / L P< 0. 05 . Conclusion At low glucose concentration high-dose vitamin C down-regulated the expression of GLUT1 and GLUT3 decreased glucose uptake in A549 cells and inhibited the proliferation of A549 cells.