趋化因子CCL2对血小板内p38MAPK-HSP27通路的活化作用

The chemokine CCL2 regulates platelet activation through P38MAPK-HSP27 signaling pathway

目的探讨趋化因子CCL2调控血小板活化的机制。方法抽取20位健康志愿者静脉血,分离血小板,采用蛋白芯片筛选及Western blotting验证,获得人血小板中经外源性CCL2(浓度1000ng/ml)刺激后磷酸化表达发生变化的激酶。选取野生型C57和CCL2–/–小鼠各10只,通过Western blotting检测经血小板诱导剂胶原刺激后小鼠血小板中P38丝裂原活化蛋白激酶(P38MAPK)和热休克蛋白27(HSP27)的表达水平。结果蛋白芯片检测结果显示,人血小板经CCL2因子体外刺激后,共有12种激酶磷酸化位点的磷酸化表达升高,其中P38MAPK(T180/Y182)与HSP27(S78/S82)的磷酸化表达经Western blotting验证明显升高(P<0.05)。应用胶原体外刺激,野生型C57小鼠血小板内P38MAPK(T180/Y182)、HSP27(S78/S82)的磷酸化表达明显高于未经刺激的小鼠血小板,差异有统计学意义(P<0.01);而经胶原刺激的CCL2–/–小鼠血小板内P38MAPK(T180/Y182)、HSP27(S78/S82)的磷酸化表达则明显低于经胶原刺激的野生型C57小鼠血小板,差异有统计学意义(P<0.05)。结论趋化因子CCL2可能通过P38MAPK-HSP27通路来调控血小板的活化。

Objective To investigate the mechanism of chemokine CCL2 in the regulation of platelets activation.Methods A total of 20 healthy volunteers was enrolled.The expression of the phosphorylated kinases in platelets was detected by the protein chip and was validated by Western blotting after stimulation by CCL2(1000 ng/ml).The platelets of 10 wild type(WT)mice and 10 CCL2^-/-mice were isolated.After activated with collagen,the phosphorylation of P38 mitogen-activated protein kinase(P38 MAPK)(T180/Y182)and heat shock protein 27(HSP27)(S78/S82)was examined by western blotting.Results A total of 12 upregulated phosphorylated kinases was identified using the protein chip screening(P<0.05).The phosphorylation of P38 MAPK(T180/Y182)and HSP27(S78/S82)was significantly increased after stimulation with CCL2(P<0.01).The phosphorylation of P38 MAPK(T180/Y182)and HSP27(S78/S82)was remarkably increased in WT mouse platelets after stimulation with collagen(P<0.01).The phosphorylation of P38 MAPK(T180/Y182)and HSP27(S78/S82)was significantly reduced in the absence of CCL2(P<0.05).Conclusions CCL2 might affect the functions of platelets through P38 MAPK-HSP27 signal pathway.