Effects of vitamin C and heat treatment on the proliferation of lung cancer A549 cells via PI3K/AKT/HIF-1α pathway
Objective To investigate the effects of vitamin C and heat treatment on lung cancer A549 cells and the related mechanisms.Method Different concentrations of vitamin C(The final concentration is 0,1,2,4,8 mmol/L 0,1,2,4,8 mmol/L)and different temperatures(37℃and 41℃)were applied to A549 cells.CCK8 assay was used to detect the cell activity,and FRASC Vitamin C assay Kit II was used to detect the concentration of vitamin C in the cells.Then A549 cells were divided into control group(37℃,0 mmol/L vitamin C),vitamin C group(37℃,8 mmol/L vitamin C),heat treatment group(41℃,0 mmol/L vitamin C)and combined group(41℃,8 mmol/L vitamin C)after 24 hours of intervention,2-NBDG was used to detect glucose uptake of A549 cells.The expression levels of GLUT1,GLUT3,PI3K,AKT,p-PI3K,p-AKT and HIF-1α were detected by Western Blot assay.The results were statistically analyzed by ANOVA and LST-t test.Result The cell viability of A549 decreased with the increase of vitamin C intervention concentration(P<0.05),and the cell viability of 41℃heat treated cells was lower than 37℃after vitamin C intervention.(P<0.05).The intracellular vitamin C level increased with the increase of intervention concentration,and the intracellular vitamin C level of 41℃heat treated cells was lower than 37℃after 2,4 and 8 mmol/L vitamin C intervention.(P<0.05).After the intervention,compared to the control group,the intracellular glucose intake of vitamin C group,heat treatment group and combined group was decreased(P<0.05),and the heat treatment group was lower than the vitamin C group(P<0.05),and the combined group was lower than the vitamin C group and heat treatment group(P<0.05).Western Blot results showed that the expression levels of GLUT1,PI3K,P-PI3K,p-AKT in the three intervention groups,and GLUT3,HIF-1α in the vitamin C group,and GLUT3,HIF-1α and AKT in the combined group were lower than those in control group(P<0.05).The protein expression levels of GLUT1,PI3K,p-PI3K in the combined group were lower than those in the vitamin C group(P<0.05),and GLUT3,AKT,P-PI3K and HIF-1α in the combined group were lower than those in the vitamin C group and the heat treatment group(P<0.05).Conclusion Compared with vitamin C or 41℃heat treatment,vitamin C combined heat treatment significantly inhibited the PI3K/AKT/HIF-1α pathway of A549 cells,down-regulated GLUT1 and GLUT3,reduced intracellular glucose uptake,and inhibited A549 cell proliferation.
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