角蛋白及p38丝裂原活化的蛋白激酶磷酸化对角质形成细胞增殖抑制的调控

Regulation of phosphorylation keratin and p38 MAPK in inhibiting proliferation of keratinocytes

目的:研究抗角蛋白抗体对角蛋白及p38丝裂原活化的蛋白激酶(MAPK)磷酸化的影响。方法:提取抗体作用过的人舌鳞状细胞癌(Tca)细胞角蛋白,以未经抗体作用的细胞作阴性对照。角蛋白经十二烷基磺酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)分离、转膜,用抗酪氨酸磷酸化抗体作用和显色。超声裂解抗体作用细胞,设置阴性对照。经SDS-PAGE分离、转膜后用抗磷酸化p38MAPK抗体作用,化学发光剂发光显影。结果:角蛋白在分子质量大约为58ku处出现阳性条带;阴性对照未出现条带。抗磷酸化p38MAPK抗体作用后在分子质量大约44ku处有阳性条带;阴性对照在相应处未出现条带。结论:角蛋白及p38MAPK磷酸化是Tca细胞增殖抑制的重要调控方式。

Objective： To study the phosphorylation of keratin and p38 mitogen-activated protein kinases（MAPK） in Tca cells cultured＂ by antikeratin antibodies. Methods： The Tca cells were cultured with antikeratin antibodies. The cells were split by using of supersonic. Then the keratins and p38 MAPK were extracted and separated by SDS-PAGE and transfered to nitrocellulose membrane （NC）. Further, the NC was incubated with phosphotyrosine Ab and phosphor-p38 MAPK kinase （Thr180/ Tyr182） Ab respectively. As negative control, the keratins and p38 MAPK were taken from the Tca cells which were not treated with related antibodies. Finally, the specific proteins were indicated by chemiluminescence. Results： The phosphorylation of keratin （58 ku） and p38 MAPK （44 ku） were detected on the NC. But they were not detected in the controls. Conclusion： Phosphorylation of keratin and p38 MAPK is important to the regulation of inhibiting Tca cells proliferation.