摘要
目的 探讨丝裂素活化蛋白激酶 (MAPK)激活、转核与血管紧张素II (AngII)刺激血管平滑肌细胞 (VSMC)增殖间的关系。方法 本实验采用培养大鼠胸主动脉VSMC。用 3 H 胸腺嘧啶核苷 (3 H TdR)掺入法测定DNA合成 ,用p42 /p44磷酸化抗MAPK抗体的蛋白免疫印迹法测定MAPK蛋白量 ,用免疫细胞化学技术观察MAPK活化并转位入细胞核的过程。结果 (1)AngII对于 3 H TdR掺入VSMC有显著刺激作用 ,该作用能被MAPK激酶的特异抑制剂PD980 5 9所抑制。AngII和PD980 5 9的上述作用都呈剂量依赖性。 (2 )AngII对MAPK蛋白表达有显著增强作用 ,此作用同样被PD980 5 9以剂量依赖方式抑制。 (3)AngII刺激 5min后 ,MAPK出现在VSMC的细胞浆中 ,30min时MAPK进入细胞核 ,3h后MAPK染色从核内消失。上述MAPK转核过程被PD980 5 9抑制。结论 本实验证实AngII能激活培养大鼠主动脉VSMC的MAPK 。
Objective To investigate the activation and translocation of mitogen activated protein kinase (MAPK) in the proliferation of vascular smooth muscle cell (VSMC) induced by angiotensin II (AngII). Methods Cultured VSMCs were obtained from rat thoracic aorta. DNA synthesis was determined by measuring the incorporation of 3?H thymidine into cells. The MAPK activity was evaluated by an immunobloting technique using anti p42/p44 phospho MAPK antibody and the presence of MAPK in the cytoplasm or nuclear was confirmed by immunocytochemistry using the specific antibody. Results (1) Ang II (10 -8 -10 -6 mol/L) dose dependently increased the incorporation of 3?H thymidine into VSMC, the increase in DNA synthesis by Ang II(10 -6 mol/L) was inhibited by PD98059 (1 50 μmol/L), a specific MAPK kinase antagonist, in a dose dependently manner. (2) The p42 p44 MAPK activities were significantly enhanced by Ang II, the activation could be also dose dependently inhibited by PD98059 (1 50 μmol/L). (3) The activated 42/p44 MAPK appeared in cytoplasm at 5 min, then translocated to nucleus at 30 min and finally disappeared from the nucleus at 3h after the stimulation of AngII. The translocation process of MAPK induced by 10 -6 mol/L AngII was blocked distinctly by PD98059 (50 μmol/L). Conclusion The results indicated that MAPK might be involved in cell proliferation of rat VSMC induced by Ang II through its activation, then translocation to nucleus.
出处
《中华心血管病杂志》
CAS
CSCD
北大核心
2001年第3期173-176,I001,共5页
Chinese Journal of Cardiology
基金
国家自然科学基金 (3 9670 3 5 6)
上海市教委项目 (97QB3 8)
中科院上海细胞生物学研究所分子细胞生物学开放研究实验室项目等资
