血小板PI3Kγ在血管损伤修复中的作用机制

Vascular inflammation and neoinitma formation after arterial injury are the main causes of restenosis following balloon angioplasty and stent implantation. Platelet adhesion, activation and aggregation play an essential role in inflammatory cell infiltration and neoinitma formation in the response to vascular injury. Increasing evidences reveal that PI3Kγ is critically involved in a number of inflammatory and autoimmune diseases. Our preliminary studies showed that PI3Kγ play a critical role in inflammatory cells recruitment at sites of injured tissue. Activated platelets promoted recruitment of inflammatory cells by PI3K/NF-κB signalling pathway. Neoinitma formation after vascular injury was significantly reduced in PI3Kγ-/- mouse indicating a key role of PI3Kγ in vascular response to injury. This study is proposed to investigate the effect of platelets PI3Kγ on neoinitma formation and vascular remodeling after vascular injury. In vivo, mouse carotid wire injury is used as models with adoptive transfer of activated, washed platelets to verify the role of PI3Kγ in the platelet-leukocyte aggregation, inflammatory cell infiltration and neoinitma formation. In vitro PI3Kγ-/- platelets are used to verify the effect of PI3Kγ on platelet activation, adhesion, aggregation and reactive oxygen species generation and its interaction with inflammatory cells and vascular resident cells. We aim to clarify the role and mechanism of platelet PI3Kγ in vascular injury and repair.

血管损伤后炎症及新生内膜过度增生是介入治疗术后再狭窄主要原因。研究提示：血小板粘附、激活、聚集是血管损伤介导的炎症细胞浸润、新生内膜形成关键环节；而PI3Kγ在炎症、氧化应激及组织损伤中起重要作用。我们前期发现：PI3Kγ在炎症细胞向损伤处聚集过程中起决定性作用，活化血小板通过PI3K/NF-KB信号通路招募炎症细胞，并且PI3Kγ基因敲除鼠（PI3Kγ-/-）血管损伤后新生内膜形成明显减少，提示：血小板PI3Kγ在血管损伤修复中起关键作用。本研究拟在前期研究基础上探讨血小板PI3Kγ在血管损伤修复中的作用。体内实验在颈动脉内膜损伤模型上静脉注射活化的血小板，观察血管损伤后血小板PI3Kγ如何调控炎症细胞浸润及新生内膜形成，体外以PI3Kγ-/-小鼠血小板为模型，观察血小板PI3Kγ对血小板功能及与炎症细胞、血管壁细胞的作用,以期阐明血小板PI3Kγ在血管损伤修复中的作用机制。

越来越多的研究表明PI3Kγ在炎症、氧化应激、心血管疾病及组织损伤、修复中起重要作用。血管损伤所致的血管疾患目前是全球范围内发病率及死亡率最高的疾病之一，血小板参与血管炎症的发生发展。本项目旨在探讨血小板PI3Kγ在小鼠血管损伤中的作用及机制。.在小鼠颈动脉结扎模型基础上发现：血管损伤后3天、7天白细胞浸润明显，其中第3天以中性粒细胞为主，第7天以单核巨噬细胞为主，而PI3Kγ基因敲除小鼠损伤血管局部炎症细胞浸润均明显减少；PI3Kγ基因敲除小鼠损伤血管局部的细胞因子(TNF-α 和IL-6)及粘附分子(ICAM-1和VCAM-1)、NF-κB表达减少，PI3Kγ基因敲除小鼠活性氧的生成较WT小鼠明显降低。血管损伤后PI3Kγ基因敲除小鼠新生内膜形成较WT小鼠明显减少，血管平滑肌细胞向内膜的增殖迁移减少；体外实验显示PI3Kγ-/-血管平滑肌增殖及迁移能力降低。.另外，我们在小鼠颈动脉部分结扎模型上反复静脉注射活化的血小板，研究发现小鼠反复注射ADP活化的血小板后血管内膜-中膜增厚血管重塑明显，而注射PI3Kγ-/-血小板后此作用明显减弱。WT血小板注射小鼠损伤血管的炎症细胞浸润、炎症介质表达、MMP-9活性及活性氧生成明显增多；而PI3Kγ-/-血小板注射小鼠血管炎症细胞浸润、炎症介质表达、MMP-9活性及活性氧生成增多不明显。.我们在体外研究发现ADP不能诱导PI3Kγ-/-血小板Akt、p-38活化，不能诱导PI3Kγ-/-血小板活化。 ADP不能诱导PI3Kγ-/-血小板NADPH酶活化及活性氧生成。ADP诱导的血小板-白细胞聚集物形成及血小板-内皮细胞粘附中PI3Kγ-/-血小板较WT 血小板明显减弱。.研究结果阐明了PI3Kγ及血小板PI3Kγ在血管损伤修复过程中作用及作用机制，共发表论文7篇，其中SCI论文5篇。培养博士研究生1名，协助培养硕士研究生3名。