淋巴细胞特异蛋白质酪氨酸激酶/P38信号传递途径在小鼠肾小管上皮细胞的作用

目的 探讨小鼠肾上管上皮细胞是否存在淋巴细胞特异蛋白质酪氨酸激酶（lck）基因表达及lck/p38细胞分裂原活化蛋白信号传递分子经IL-12、IL-2、脂多糖（LPS）等刺激时在肾小管上皮细胞的变化。方法 无菌分离培养BALB/C小鼠肾小管上皮细胞，采用地高辛标记的lck寡核苷酸探针原位杂交检测肾小管上皮细胞中lck基因表达；应用放射自显影方法观察lck活性变化，利用免疫印迹分析lck蛋白表达及p38磷酸化。结果 IL-12、IL-2、LPS刺激肾小管上皮细胞活化，lck mRNA表达较对照组明显增强；经上述物质刺激后lck活性、p38磷酸化分别于5min、15min最强，其中LPS刺激最显著而lck、p38蛋白水平则无明显变化。加入lck抑制剂PP1，IL-12刺激时未发现p38磷酸化，而IL-2、LPS刺激只有轻度抑制。结论 IL-12、IL-2、LPS促进肾小管上皮细胞中lck基因表达，并只有IL-12唯一通过lck诱导p38磷酸化，它们皆可通过lck/p38信号传递途径参与对肾小皮细胞发挥生物作用。

Role of Ick/ p38 signaling pathways in murine renal tubular epithelial cells

Objective To investigate whether lymphocyte-specific protein-tyrosine kinase (lck) expression exists in murine renal tubular epithelial cells and to observe the variations of lck/p38 mitogen-activated protein kinase signaling pathways molecules in renal tubular epithelial cells stimulated with IL-12, IL-2 and LPS, respectively. Methods Renal tubular epithelial cells were obtained by separation from BALB/C mice and cultured in conditioned medium. The mRNA expression of lck in renal tubular epithelial cells was measured by in situ hybridyzation with digoxin-labeled lck oligonucleotide prob. Lck activity was determined by autoradiography, and expression of lck protein and phosphorylation of p38 by western blotting analysis. Results The level of lck mRNA was significantly enhanced in renal tubular epithelial cells stimulated with IL-12, IL-2 or LPS compared to control groups. The maximal effect on lck activity and p38 phosphorylation were present 5 and 15 minutes after stimulation as indicated above. In addition, the effect of LPS was the most obvious among the three stimulations, while the expression level of lck and p38 protein was not increased. IL-12 did not stimulate p38 phosphorylation in renal tubular epithelial cells pre-incubated with lck inhibitor PP1, while the inhibitory effects were mild in renal tubular epithelial cells stimulated with IL-2 or LPS. Conclusion IL-12, IL-2 and LPS may promote lck gene express in renal tubular epithelial cells and only IL-12 can induce p38 phosphorylation through lck uniquely, and they can play biologic role in renal tubular epithelial cells via lck/ p38 signaling pathways.