Robust thrombolytic and anti-inflammatory action of a constitutively active ADAMTS13 variant in murine stroke models
Robust thrombolytic and anti-inflammatory action of a constitutively active ADAMTS13 variant in murine stroke models
复制标题
DOI：
10.1182/blood.2021012787
复制
发表时间：
2022-03-10
期刊：
影响因子：
20.3
通讯作者：
Allan, Stuart M.
中科院分区：
文献类型：
--
作者：
South, Kieron;Saleh, Ohud;Allan, Stuart M.
我们对ADAMTS13结构的理解的进步，以及充分活性所需的构象变化，重新激活了其作为溶栓治疗的可能性。我们测试了一种新的Ala1144Val ADAMTS13变体（组成活性[ca] ADAMTS13），它具有组成活性，通过ADAMTS13活性的体外测定来表征，并在2种缺血性卒中小鼠模型、远端FeCl3大脑中动脉闭塞（MCAo）模型和伴有全身炎症和缺血/再灌注损伤的短暂性大脑中动脉闭塞（tMCAO）模型中大大增强了溶栓活性。在两种模型中，疗效的主要测量是恢复区域脑血流（rCBF）到MCA区域，这是用激光散斑对比成像确定的。与野生型（wt） ADAMTS13相比，caADAMTS13变体具有组成活性构象和抗荧光共振能量转移底物von Willebrand factor 73 （FRETS-VWF73）的活性增强了5倍。此外，caADAMTS13在亚生理浓度下抑制vwf介导的血小板捕获，并增强t-PA/纤溶蛋白（原）的溶解，而wtADAMTS13没有观察到这两种情况。在caADAMTS13治疗的动物中，观察到rCBF的显著恢复和病变体积的减少。在FeCl3 MCAo 1小时后，caADAMTS13变体显著减少了MCA中残留的VWF和纤维蛋白沉积、血小板聚集形成和中性粒细胞募集。在tMCAo模型再灌注4小时后给药，caADAMTS13变体诱导血小板聚集体的显著溶解和由此导致的组织灌注不足的减少。caADAMTS13变体代表了治疗急性缺血性卒中的潜在可行治疗选择，在其他血栓适应症中，由于其组成活性构象增强了体外和体内活性。
Advances in our understanding of ADAMTS13 structure, and the conformation changes required for full activity, have rejuvenated the possibility of its use as a thrombolytic therapy. We have tested a novel Ala1144Val ADAMTS13 variant (constitutively active [ca] ADAMTS13) that exhibits constitutive activity, characterized using in vitro assays of ADAMTS13 activity, and greatly enhanced thrombolytic activity in 2 murine models of ischemic stroke, the distal FeCl3 middle cerebral artery occlusion (MCAo) model and transient middle cerebral artery occlusion (tMCAO) with systemic inflammation and ischemia/reperfusion injury. The primary measure of efficacy in both models was restoration of regional cerebral blood flow (rCBF) to the MCA territory, which was determined using laser speckle contrast imaging. The caADAMTS13 variant exhibited a constitutively active conformation and a fivefold enhanced activity against fluorescence resonance energy transfer substrate von Willebrand factor 73 (FRETS-VWF73) compared with wild-type (wt) ADAMTS13. Moreover, caADAMTS13 inhibited VWF-mediated platelet capture at subphysiological concentrations and enhanced t-PA/plasmin lysis of fibrin(ogen), neither of which were observed with wtADAMTS13. Significant restoration of rCBF and reduced lesion volume was observed in animals treated with caADAMTS13. When administered 1 hour after FeCl3 MCAo, the caADAMTS13 variant significantly reduced residual VWF and fibrin deposits in the MCA, platelet aggregate formation, and neutrophil recruitment. When administered 4 hours after reperfusion in the tMCAo model, the caADAMTS13 variant induced a significant dissolution of platelet aggregates and a reduction in the resulting tissue hypoperfusion. The caADAMTS13 variant represents a potentially viable therapeutic option for the treatment of acute ischemic stroke, among other thrombotic indications, due to its enhanced in vitro and in vivo activities that result from its constitutively active conformation.