凝血酶对U_(937)细胞uPAR mRNA表达的影响

Effects of Thrombin on uPAR mRNA Expression in Cultured U_（937） Cells

目的研究凝血酶通过凝血酶受体 (TR)对细胞uPARmRNA表达的影响。 方法在凝血酶、灭活凝血酶、和抗TR小肽多抗等作用下 ,以RT -PCR法测定培养U937细胞uPARmRNA的表达量。 结果凝血酶增强uPARmRNA表达呈现凝血酶剂量和作用时间的依赖性。其作用机制与凝血酶和细胞膜上TR间的结合、以及凝血酶的蛋白水解活性有关。高浓度凝血酶或中等浓度凝血酶在延长作用时间时 ,对U937细胞uPARmRNA表达有明显抑制效应。 结论凝血酶对U937细胞uPARmRNA表达的影响 ,除通过活化TR外可能还存在其它的途径 。

Objective To study the role of thrombin in uPAR mRNA expression in cultured U 937 cells. Methods The method of reverse transcriptional PCR (RT-PCR)for monitoring urokinase type plasminogen activator receptor (uPAR) mRNA has been used to evaluate uPAR mRNA expressive changing in U 937 cells by adding different concentrations of thrombin, thrombin plus rabbit anti-thrombin receptor 14 peptides (KYEPFWEDEEKNS)-antibody, or DFP-thrombin added into the experimental cells cultures. Results The effect of thrombin on increasing uPAR mRNA expression was blocked by the rabbit antibody, which depended upon the antibody concentration used. No effects were shown for DFP-thrombin on the uPAR mRNA expression. Under certain concentration of antibody, the blockage effect was more strong in the higher concentration of thrombin (40 nmol/L)than in the lower one (10 nmol/L). As compared with the changes of uPAR mRNA level by the increasing doses these two kinds of thrombin in the controls and from tested samples. When U 937 cells cultured in the medium with rather high of thrombin up to 24h, the uPAR mRNA levels actually dropped about 50% for 40 nmol/L thrombin treated cultures as compared to the controls, and uPAR mRNA levels also decreased with thrombin concentration (from 2.5nmol/L to 40 nmol/L). Conclusion It is clarified that the action of thrombin on the cells to increase uPAR mRNA expression was mediated by the thrombin receptor, and thrombin played a suppressive role (may be not through the thrombin receptor) as well as on the experimental cells during its up regulation for the uPAR mRNA expression.