摘要目的 探讨磷脂酰肌醇-3激酶(phosphatidylinositol 3-kinase,PI3K)-蛋白激酶B(protein kinase B,AKT)信号通路在调节哺乳期乳腺细胞钠碘转运体(Na+-I-symporter,NIS)表达中的作用,以及不同碘水平对该通路的影响.方法 将原代培养的小鼠哺乳期乳腺细胞分为3组:①对照组[0 μmol/PI3K抑制剂LY294002+0 μg/L胰岛素样生长因子Ⅰ(insulin-like growth factor Ⅰ,IGF-Ⅰ)];②刺激组(50 μg/L IGF-Ⅰ);③抑制组(40 Iμmol/L LY294002+ 50 μg/L IGF-Ⅰ).此外,将原代培养的小鼠哺乳期乳腺细胞按不同碘水平(0、5、50、1 000、3 000 μg/L)处理分为低碘1、2组,适碘组,高碘1、2组,并加入IGF-Ⅰ(50μg/L)刺激.采用实时荧光定量PCR及蛋白免疫印迹法检测上述各组细胞中AKT、NIS mRNA及蛋白表达水平.结果 刺激组AKT mRNA表达水平(1.497±0.550)高于抑制组(0.777±0.108,P＜0.05),而刺激组NIS mRNA及蛋白表达水平(0.783±0.187、0.618±0.103)均低于抑制组(2.430±1.423、1.417±0.250,P均＜0.05).随着碘水平的增加,除高碘1组(1.090±0.356)外,低碘1、2组,适碘组,高碘2组AKT mRNA表达水平(1.758±0.893、1.320±0.538、1.003±0.006、0.745±0.307)为逐渐下降趋势;低碘1、2组,适碘组,高碘1、2组NIS mRNA (2.259±0.682、1.823±0.332、1.409±0.366、1.321±0.405、1.150±0.454)及总AKT蛋白(0.640±0.106、0.601±0.081、0.583±0.089、0.555±0.097、0.532±0.023)表达水平均呈下降趋势;除低碘2组(0.484±0.179)外,NIS蛋白表达水平(0.556±0.199、0.502±0.179、0.455±0.126、0.435±0.138)呈下降趋势;除低碘2组(0.076±0.045)外,p-AKT蛋白表达水平(0.078±0.049、0.079±0.040、0.085±0.055、0.095±0.051)呈上升趋势.结论 PI3K-AKT信号通路在调节哺乳期乳腺细胞NIS表达方面起抑制作用.

abstractsObjective To elucidate the function of phosphatidylinositol 3-kinase (PI3K)-protein kinase B (AKT) signaling pathway underlying the regulation of Na+-I-symporter (NIS) and the influence of different levels of iodine on PI3K-AKT signaling pathway in lactating breast cells.Methods The primary cultured mammary gland cells were divided into three groups:①control group [0 μmol/L LY294002 + 0 μg/L insulin-like growth factor Ⅰ (IGF-Ⅰ)];②stimulation group (50 μg/L IGF-Ⅰ);③inhibition group (40 μmo]/L LY294002 + 50 μg/L IGF-Ⅰ).In addition,the cells were treated with different iodine contents (0,5,50,1 000,3 000 μg/L) for low iodine groups 1 and 2,iodine group,high iodine groups 1 and 2,and IGF-Ⅰ (50 μg/L) was used to stimulate PI3K-AKT signaling pathway.The expressions of AKT and NIS mRNA and protein were determined by real-time quantitative PCR and Western blotting,respectively.Results The expression of AKT mRNA (1.497 ± 0.550) in stimulation group was higher than that in inhibition group (0.777 ± 0.108,P ＜ 0.05),while the expression of NIS mRNA and protein in stimulation group (0.783 ± 0.187,0.618 ± 0.103) was lower than those in inhibition group (2.430 ± 1.423,1.417 ± 0.250,all P ＜ 0.05).With the iodine concentration increasing,except high iodine group 1 (1.090 ± 0.356),the expression of AKT mRNA in low iodine groups 1 and 2,iodine group,high iodine group 2 (1.758 ± 0.893,1.320 ± 0.538,1.003 ± 0.006,0.745 ± 0.307) tended to decline;total AKT protein (0.640 ± 0.106,0.601 ± 0.081,0.583 ± 0.089,0.555 ± 0.097,0.532 ± 0.023) and NIS mRNA (2.259 ± 0.682,1.823 ± 0.332,1.409 ± 0.366,1.321 ± 0.405,1.150 ± 0.454) tended to decline in low iodine groups 1 and 2,iodine group,high iodine groups 1 and 2;except low iodine group 2 (0.484 ± 0.179),NIS protein expression tended to decline (0.556 ± 0.199,0.502 ± 0.179,0.455 ± 0.126,0.435 ± 0.138);however,except low iodine group 2 (0.076 ± 0.045),the p-AKT protein expressions (0.078 ± 0.049,0.079 ± 0.040,0.085 ± 0.055,0.095 ± 0.051) were on the rise.Conclusion PI3K-AKT signaling pathway may play an inhibition role in the expression of NIS in lactating breast cells.