S1PR2介导血小板焦亡并参与脓毒症发生发展的机制研究

Activation of inflammasome has been described as the main pathway involved in platelet pyroptosis, counts decrease and inflammatory cytokine release during sepsis. So, it is very emergent to illustrate the molecular mechanism underlying inflammasome activation in the pathogenesis of sepsis. S1PR2 is highly expressed on platelet. Our previous studies found that activation of the S1PR2 signaling deteriorated the adverse outcome of sepsis, in which the detailed molecular mechanism remains unknown. Our preliminary experiments also found that S1PR2 gene deficiency inhibited the activation of casepase-11 inflamasome. Furthermore, decreased platelets pyroptosis was observed in S1pr2-/- mice after sepsis challenge. It has been demonstrated that RhoA-ROCK signaling induced aberrant actin polymerization and initiated inflammasome activation. We also found S1PR2 gene caused RhoA activiation.Thus, we speculate that S1PR2 signaling induces the activation of caspase-11 inflammasome mediated by RhoA-ROCK pathway in platelets, which promotes sepsis development. Using genetic murine and cellular models, adoptive cellular therapy and the techniques such as laser confocal microscopy etc., the current project will elaborate the hypothesis. These will enrich the inflammasome activation theory of sepsis, and also be meaningful to the exploration of novel goal-directed therapeutic strategies for sepsis in the near future.

血小板通过炎性体途径诱导自身焦亡，造成血小板数量急剧下降，炎症介质大量释放，在脓毒症发生发展中发挥重要作用，因此阐明调控血小板炎性体活化的分子机制对于防治脓毒症尤为迫切。S1PR2高表达于血小板，本团队曾报道活化S1PR2加重脓毒症不良预后，但机制不明。我们预实验发现：S1PR2功能缺失导致caspase-11炎性体活化障碍；S1pr2-/-小鼠脓毒症后血小板焦亡水平降低。文献证实RhoA-ROCK信号启动炎性体活化，我们发现血小板S1PR2活化促进RhoA激活。因此推测：S1PR2通过RhoA-ROCK信号通路启动caspase-11炎性体活化，促进血小板焦亡，介导脓毒症发生发展。本项目拟运用遗传动物/细胞模型、细胞过继回输、激光共聚焦成像等技术阐明上述假说，完善脓毒症病理过程中的炎性体活化理论，为脓毒症防治提供新靶标。

脓毒症是感染引起的机体反应异常导致危及生命的器官功能障碍，进一步可发展为脓毒症休克，死亡率高达40%，已成为威胁人类健康的难治性疾病。近年来研究发现，脓毒症患者机体存在极为复杂的免疫功能紊乱状态：一方面，表现为以炎症介质过度释放为代表的过度炎症反应；另一方面，表现为吞噬杀菌活性减弱，抗原呈递功能受抑等抗感染免疫防御能力降低。近来研究发现而血小板在脓毒症免疫调控中扮演着重要角色，其主要参与炎症介质释放和免疫细胞调节，血小板减少已成为脓毒症患者机体炎症反应增强和预后不良的标志。因此，如何控制血小板炎症因子释放，预防血小板数量减少，从而改善脓毒症机体炎症免疫反应，已成为脓毒症研究热点。研究表明，炎性体活化诱导血小板焦亡和炎症介质释放在脓毒症的发生发展过程中发挥重要作用。一方面，血小板焦亡促进炎症介质如IL-1β、IL-18等的大量释放，引起组织损伤；另一方面，血小板焦亡导致血小板死亡，表现为数量显著下降，引起机体免疫功能低下，最终促进脓毒症和感染性休克的发生发展。因此，解析血小板介导脓毒症发生发展的炎性体信号分子机制对丰富脓毒症病理生理机制具有重要意义。本项目利用基因敲除小鼠（S1pr2-/-、和Casp11-/-），综合运用siRNA和腺病毒转染以及免疫荧光等技术，从整体动物到细胞和分子水平，系统研究S1PR2对血小板RhoA-ROCK信号通路、Caspase-11炎性体活化及其介导血小板焦亡和IL-1β、IL-18等成熟释放的影响和作用，分析其与器官功能损伤的相关性，并探讨干预S1PR2介导的血小板焦亡对脓毒症小鼠的保护作用，从而阐明了S1PR2通过RhoA-ROCK信号通路调控caspase-11炎性体活化信号分子机制，完善了S1PR2在脓毒症发病中发挥重要作用的理论体系，并为探寻脓毒症预警标志和治疗靶标提供了科学依据。