Department of Hematology, The Third Xiangya Hospital, Central South University, No. 138 Tongzi Road, Changsha, Hunan, PR China.
Cell Immunol. 2018 Jun;328:86-92. doi: 10.1016/j.cellimm.2018.04.002. Epub 2018 Apr 7.
Platelet is an important cell contributing to hemostasis and immunity. Bacterial lipopolysaccharide (LPS), mainly functioning by stimulating toll-like receptor 4 (TLR4), mediates platelet activation and sepsis. However, the inter-relationship between these players in sepsis remains unknown. We found that the aggregation of platelets was enhanced in complete blood of sepsis patients than that of healthy donors. PRP isolated from complete blood of healthy donors was used in the following study to filter out the interference of irrelevant cells. The results shown that the maximum aggregation rate (MAR) was significantly higher in LPS-challenged PRP model than that of controls, and administration of the specific TLR4 inhibitor, TAK242, reduced the MAR in this model. LPS promoted P-selectin expression and intracellular ROS production, and both TAK242 and N-acetyl-L-cysteine (NAC) could depressed the LPS-induced increase of P-selectin and intracellular ROS. HO administration increased P-selectin expression partially but had little effect on intracellular ROS, thought it increased mitochondrial damage. In vivo, LPS increased both intracellular ROS and CD62P comparing with that of controls, effects that were prevented by TAK242. Furthermore, platelet aggregation through LPS-TLR4 pathway was involved in AKT, PKC and p38 phosphorylation but not cGMP/cAMP pathway. In conclusion, this study shows that intracellular ROS, not extracellular ROS such as HO, plays a crucial role in facilitating platelet aggregation via LPS/TLR4 pathway, and this process was involved in AKT, PKC and p38 phosphorylation but not cGMP/cAMP pathway. The results would helpful for understanding the role of intracellular ROS and LPS-TLR4 pathway in platelet aggregation.
血小板是参与止血和免疫的重要细胞。细菌脂多糖（LPS）主要通过刺激 toll 样受体 4（TLR4）来介导血小板激活和脓毒症。然而，在脓毒症中，这些因素之间的相互关系尚不清楚。我们发现脓毒症患者的全血中血小板聚集增强，高于健康供体。从健康供体的全血中分离的 PRP 用于以下研究，以过滤掉无关细胞的干扰。结果表明，LPS 刺激的 PRP 模型中的最大聚集率（MAR）明显高于对照，并且特异性 TLR4 抑制剂 TAK242 的给药降低了该模型中的 MAR。LPS 促进 P-选择素表达和细胞内 ROS 产生，TAK242 和 N-乙酰-L-半胱氨酸（NAC）均可抑制 LPS 诱导的 P-选择素和细胞内 ROS 增加。HO 给药部分增加 P-选择素表达，但对细胞内 ROS 影响不大，尽管它增加了线粒体损伤。在体内，LPS 增加了细胞内 ROS 和 CD62P，与对照相比，这一作用被 TAK242 阻止。此外，通过 LPS-TLR4 途径的血小板聚集涉及 AKT、PKC 和 p38 磷酸化，但不涉及 cGMP/cAMP 途径。总之，这项研究表明，细胞内 ROS（而非 HO 等细胞外 ROS）在通过 LPS/TLR4 途径促进血小板聚集方面起着至关重要的作用，这一过程涉及 AKT、PKC 和 p38 磷酸化，但不涉及 cGMP/cAMP 途径。这些结果有助于了解细胞内 ROS 和 LPS-TLR4 途径在血小板聚集中的作用。
