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目的 探讨蛋白酪氨酸激酶 (PTKs)活性在人肾小管上皮细胞 (HKC)转分化中的作用。方法 将传代的HKC细胞分为 :(1)无血清对照组 (C) ;(2 )酸性成纤维细胞生长因子 (aFGF)组 (A) ;(3) genistein组 (G) ,加入 80ng/ml重组人aFGF条件下 ,分别加入不同浓度的 genistein ;培养 72h。应用免疫组化检测HKC细胞表达α SMA(α smoothmuscleactin ,α SMA)的变化 ;对转分化过程中PTKs的活性用流式细胞仪加以分析。结果 aFGF(A)组加入 80ng/mlaFGF后a 平滑肌动蛋白 (a SMA)表达增强 ,细胞a SMA阳性率 (6 0 .6 0± 2 .70 ) %较对照组 (C) (3.35± 1.4 5 ) %有非常显著性差异 (P <0 .0 1)。当G4,5组介入genistein浓度 4 8、96 μg/ml时 ,细胞α SMA阳性率为 (3.90± 2 .0 9) % ,(3.98± 1.75 ) % ,较aFGF组 (A)明显降低 (P <0 .0 1)。PTKs的活性随加入aFGF的浓度增加而增高 ,当aFGF的浓度为 2 0、4 0、80ng/ml时 ,分别比对照组 (C)PTKs的活性增加 16 %、17%、2 4 .6 % ,当介入genistein剂量为 6、12、2 4、4 8μg/ml时 ,分别比aFGF组 (A)PTKs的活性下降 4 5 .6 %、5 3.5 %、6 6 %、81.9% ,干预因素作用下HKC细胞PTKs活性与genistein剂量呈负相关 (r =- 0 .987,P <0 .0 5 )。结论 HKC转分化的机制可能与PTKs活性失控
Objective To investigate the role of protein tyrosine kinases (PTKs) in the transdifferentiation of human renal tubular epithelial cells (HKC). Methods The subcultured HKC cells were divided into: (1) Serum-free control group (C); (2) Acid fibroblast growth factor (AFGF) group (A); (3) Genistein group Recombinant human aFGF conditions, were added to different concentrations of genistein; cultured 72h. The expression of α-smooth muscle actin (α SMA) in HKC cells was detected by immunohistochemistry. The activity of PTKs in transdifferentiation was analyzed by flow cytometry. Results The a SMA positive expression of aFGF (A) group was significantly higher than that of control group (C) (3.35 ± 1.4 5) )% Has a very significant difference (P <0. The positive rate of α SMA was (3.90 ± 2.09)% and (3.98 ± 1.75)%, respectively, when compared with that of aFGF group (A) when the concentration of genistein was 48 and 86 μg / ml <0 .0 1). The activity of PTKs increased with the increase of aFGF concentration. When the concentration of aFGF was 20, 40, 80 ng / ml, the activity of PTKs in control group increased by 16%, 17%, 24.6% , The activity of PTKs decreased by 45.6%, 53.5%, 66%, 81.9% compared with aFGF group (A) when the dose of genistein was 6,12,2 and 4,48μg / ml respectively The activity of PTKs in HKC cells was negatively correlated with the dosage of genistein (r = - 0.987, P <0.05). Conclusions The mechanism of HKC transdifferentiation may be out of control with PTKs activity