Ma Li-Ping, Wei Jing, Chang Jian-Xing, Zhang Cheng, Pei Zhi-Xin, Yang Qiu-Hong
Department of Hematology, The Second Hospital, Sun Yat-Sen University, Guangzhou 510120, Guangdong Province, China. mlp215yahoo.com.cn
Zhongguo Shi Yan Xue Ye Xue Za Zhi. 2009 Dec;17(6):1564-8.
The study was aimed to investigate the expression of Toll-like receptor 4 (TLR4) on platelets and to determine whether platelet TLR4 involves in its activation induced by lipopolysaccharide (LPS). Human platelet-rich plasma (PRP) and platelet suspension obtained from 15 healthy individuals pretreated with a concentration of 0.2 microg/ml of LPS in the presence or absence of thrombin (1 U/ml) for 1 hour. The expressions of TLR4, CD62P (P-select) and CD40L on platelets were detected by flow cytometry, and platelet TLR4 expression was further determined by Western blot analysis. The results indicated that the percentage of TLR4-positive platelets induced by thrombin was increased by 32.34% compared with the resting platelets (25.44%, p < 0.05). TLR4 expression on platelets treated with LPS was remarkably elevated in the presence or absence of thrombin. However, the expression level of the former was much higher than that of the latter and thrombin stimulation alone (p < 0.05). Moreover, the similar results were found in Western blot analysis. Synchronously, expressions of CD62P and CD40L on resting platelets were 6.39% and 2.45%, they were also markedly increased when treated with thrombin (42.68% and 14.8%) and LPS respectively, and the increases of expression of CD62P and CD40L were more significant when stimulated with both LPS and thrombin (63.03% and 13.94%). Although anti-TLR4 antibody inhibited significantly the increase of TLR4, CD62P and CD40L on platelets induced by LPS, which did not affect their increase induced by thrombin. In conclusion, the evidence has been shown that functional TLR4 can be expressed on human platelets. It may involve in platelet activation as an important mediator of LPS-induced CD62P and CD40L expressions on platelets.
本研究旨在探讨Toll样受体4(TLR4)在血小板上的表达,并确定血小板TLR4是否参与脂多糖(LPS)诱导的血小板激活。从15名健康个体获得富含血小板血浆(PRP)和血小板悬液,在有或无凝血酶(1 U/ml)存在的情况下,用浓度为0.2 μg/ml的LPS预处理1小时。通过流式细胞术检测血小板上TLR4、CD62P(P选择素)和CD40L的表达,并通过蛋白质印迹分析进一步确定血小板TLR4表达。结果表明,与静息血小板相比,凝血酶诱导的TLR4阳性血小板百分比增加了32.34%(25.44%,p<0.05)。在有或无凝血酶存在的情况下,用LPS处理的血小板上TLR4表达均显著升高。然而,前者的表达水平远高于后者及单独凝血酶刺激(p<0.05)。此外,蛋白质印迹分析也得到类似结果。同时,静息血小板上CD62P和CD40L的表达分别为6.39%和2.45%,在用凝血酶(42.68%和14.8%)和LPS处理时也显著增加,当同时用LPS和凝血酶刺激时,CD62P和CD40L表达的增加更显著(63.03%和13.94%)。虽然抗TLR4抗体显著抑制LPS诱导的血小板上TLR4、CD62P和CD40L的增加,但不影响凝血酶诱导的增加。总之,已有证据表明功能性TLR4可在人血小板上表达。它可能作为LPS诱导血小板上CD62P和CD40L表达的重要介质参与血小板激活。