DUSP22在血小板活化和动脉血栓形成中的作用研究

It has been recently discovered that protein tyrosine phosphatases play an important role in modulating the functions of platelet. We have been focused on studying the functions of the dual-specificity phosphatase 22 (DUSP22) and our preliminary data show that the inhibition of the DUSP22 activity selectively limits the collagen-induced platelet granule secretion and aggregation via the impediment to tyrosine phosphorylation of Syk, a key signaling protein downstream of GPVI. These results suggest that DUSP22 is implicated in regulation of the platelet activation and the arterial thrombosis. This work aims to understand the molecular functions of DUSP22 in regulating the collagen-induced platelet activation as well as to explore the functional relevance of DUSP22 in other agonist-induced platelet activation. We will focus on elucidating the mechanism by which the DUSP22 modulates the collagen-glycoprotein VI signaling pathway, using a set of molecular and biochemical methods such as western blot, immunoprecipitation, flow cytometry and confocal laser scanning microscopy. Moreover, we will employ the in vitro and in vivo arterial thrombosis models to explore and substantiate the causative role of DUSP22 in arterial thrombosis. This.work will contribute to unraveling the role and mechanism of DUSP22 in modulating the platelet functions and will provide insights into identifying the therapeutic targets against thrombosis.

近年来，人们发现蛋白酪氨酸磷酸酶在调节血小板功能方面起着重要作用。我们已通过前期实验发现抑制双特异性磷酸酶22（DUSP22）活性可以抑制“胶原-GPVI”通路下游重要信号分子Syk的酪氨酸磷酸化进而选择性抑制胶原诱导的血小板颗粒释放和聚集。据此，推测DUSP22特异性调控胶原诱导的血小板活化，且影响动脉血栓的形成。为验证此假设，本项目拟在前期工作基础上，进一步明确DUSP22是否调控除胶原外的多种激活剂引起的血小板激活，并利用免疫印迹、免疫共沉淀、流式细胞术和激光共聚焦显微技术等分子细胞生物学方法研究DUSP22调控“胶原-GPVI”信号通路的分子机制。此外，本项目还将利用体内外动脉血栓模型，明确DUSP22在动脉血栓形成过程中的作用。本项目的实施有助于阐释DUSP22在血小板中的生物学功能及相应分子机制，为我们提供新型、潜在的抗血栓药物靶点奠定基础。

近年来，人们发现蛋白酪氨酸磷酸酶在调节血小板功能方面起着重要作用。我们已通过前期实验发现抑制双特异性磷酸酶22（DUSP22）活性可以抑制“胶原-GPVI”通路下游重要信号分子Syk的酪氨酸磷酸化进而选择性抑制胶原诱导的血小板颗粒释放和聚集。DUSP22是否特异性调控胶原诱导的血小板活化，且影响动脉血栓的形成并不清楚。本项目在前期工作基础上，进一步确认BML-260可抑制胶原诱导的血小板颗粒释放、血小板聚集以及αIIbβ3“由内到外”信号的激活，但对TRAP诱导的凝血酶信号通路的激活没有明显影响。此外，为了确认通过小分子抑制剂获得的体外数据，本项目利用C57BL/6J小鼠构建了DUSP22基因敲除小鼠，研究发现DUSP22基因敲除的小鼠血小板对GPVI特异性配体胶原和Convulxin诱导的血小板颗粒释放和血小板聚集没有明显差异。小鼠断尾实验也确认DUSP22缺失并不影响小鼠正常的生理止血功能。最后，DUSP22的缺失也不能抑制血栓的形成。因此本项目最终确认DUSP22并不能调控血小板的活化，DUSP22不能作为潜在的抗血小板靶点，不具有抗血小板价值。