Species-dependent variability of ADAMTS13-mediated proteolysis of human recombinant von Willebrand factor
Species-dependent variability of ADAMTS13-mediated proteolysis of human recombinant von Willebrand factor
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DOI：
10.1111/j.1538-7836.2009.03453.x
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发表时间：
2009-07-01
影响因子：
10.4
通讯作者：
Schwarz, H. P.
中科院分区：
文献类型：
--
作者：
Varadi, K.;Rottensteiner, H.;Schwarz, H. P.
背景：血管性血友病因子（VWF）由一系列多聚体组成，其大小受血浆金属蛋白酶ADAMTS13调控。目的：研究不同物种血浆中ADAMTS13蛋白水解重组人VWF （rVWF）的效果和安全性，以作为临床前动物模型。方法：在体外适度变性条件下研究rVWF的降解，并采用多聚体分析、残余胶原结合和免疫印迹分析监测其降解情况。通过给食蟹猴、兔和vwf缺陷小鼠注射rVWF，并随后用免疫印迹法对血浆样品进行分析，来测定体内的卵裂。用合成人VWF肽（FRETS-VWF73）测定血浆ADAMTS13水平。结果：在实验动物中，只有兔血浆与人血浆具有相同的体外蛋白水解能力。小鼠血浆几乎不能分裂裂谷病毒。在不同剂量的裂谷病毒感染下，家兔和食蟹猴体内产生了adamts13特异性裂解产物，程度较小。在小鼠中几乎没有发生卵裂。兔和猴血浆中的ADAMTS13活性水平与人血浆中的相似，并且在rVWF输注到非常高剂量时没有显着改变，这表明rVWF不会导致两种动物内源性ADAMTS13的耗尽。结论：根据动物模型试验结果解释人类裂谷热生理时，需要考虑不同物种对裂谷热裂解的敏感性差异。
Background: von Willebrand factor (VWF) is composed of a series of multimers, the sizes of which are regulated by the plasma metalloprotease ADAMTS13. Objective: Proteolysis of human recombinant VWF (rVWF) by ADAMTS13 present in the plasma of different species typically used as preclinical animal models was investigated to evaluate the efficacy and safety of rVWF. Methods: Degradation of rVWF was studied in vitro under moderate denaturing conditions and was monitored by multimer analysis, residual collagen binding, and immunoblot analysis. In vivo cleavage was determined by administration of rVWF to cynomolgus monkeys, rabbits and VWF-deficient mice and subsequent analysis of plasma samples by immunoblot. Plasma ADAMTS13 levels were determined with a synthetic human VWF peptide (FRETS-VWF73). Results: From the animals tested, only rabbit plasma was as efficient as human plasma in proteolysing rVWF in vitro. Mouse plasma virtually failed to cleave rVWF. Administration of human rVWF resulted in ADAMTS13-specific cleavage products in rabbits and, to a lesser extent, in cynomolgus monkeys at various doses of rVWF. Virtually no cleavage occurred in mice. ADAMTS13 activity levels in rabbit and monkey plasma were similar to those in human plasma and were not significantly altered upon infusion of rVWF up to very high doses, indicating that rVWF did not lead to an exhaustion of endogenous ADAMTS13 in both species. Conclusions: The differences in susceptibility to cleavage of rVWF by different species need to be considered when interpreting the physiology of human rVWF from results of tests in animal models.