Shigatoxin Triggers Thrombotic Thrombocytopenic Purpura in Genetically Susceptible ADAMTS13-Deficient Mice.

Abstract

The life threatening disease TTP is associated with ultra-large von Willebrand Factor multimers (UL-VWF) in the circulation due to inherited or acquired deficiency of the ADAMTS13 metalloprotease. Here we show that ADAMTS13-deficient mice generated by gene targeting are viable and exhibit normal survival through 2 years of age. Despite the absence of VWF-cleaving protease activity (<1% of normal), wild-type and ADAMTS13-deficient plasma exhibit identical VWF multimer distributions, and Adamts13−/− mice develop spontaneous TTP at an extremely low rate (2 cases out of 358 mice). However, intravital microscopy demonstrated that VWF-mediated platelet-endothelial interactions are significantly prolonged in Adamts13−/− mice. These observations suggested that additional environmental triggers and/or genetic modifying factors may be required to bring about TTP in the setting of ADAMTS13 deficiency. To address the effect of VWF level on development of TTP, Adamts13−/− mice were crossed to mice of the CASA/Rk strain which exhibit markedly elevated plasma VWF levels. Resulting CASA/Adamts13−/− mice demonstrated plasma VWF ranging from 150% to 600% of C57BL/6 controls, and we found that 21% of these mice were thrombocytopenic at baseline (vs. 0% of controls). Introduction of the CASA/Rk genetic background also resulted in the appearance of UL-VWF in CASA/Adamts13−/− mice, further prolonged VWF-mediated platelet-endothelial cell interactions, increased the rate of spontaneous TTP, and markedly decreased survival. Challenge of CASA/Adamts13−/− mice with shigatoxin (derived from bacterial pathogens associated with the related human disease hemolytic uremic syndrome) resulted in a striking syndrome closely resembling human TTP, with thrombocytopenia, profound microangiopathic hemolytic anemia, and platelet- and VWF-thrombi seen in multiple organs. Surprisingly, we observed no correlation between plasma VWF level and severity of TTP, implying the existence of TTP-modifying genes distinct from VWF. Our laboratory is pursuing the identification of these genes which may provide insight into the pathogenesis and treatment of TTP in humans. Finally, our data also suggest that microbial-derived toxins, or possibly other sources of endothelial injury, may be among the key factors required to trigger acute TTP in the setting of ADAMTS13 deficiency.

Shiga toxin-2 results in renal tubular injury but not thrombotic microangiopathy in heterozygous factor H-deficient mice

“…Consistent with our observations, many studies in which mice have either been inoculated with Stx-producing E. coli strains [18,29,30] or injected with purified Stx with [20,21,[25][26][27]31] or without LPS [22][23][24]32,33] have not developed evidence of glomerular thrombosis. We have summarized these studies in Table 2, where we list the experimental insult (e.g.…”

“…We have summarized these studies in Table 2, where we list the experimental insult (e.g. Stx-producing bacterial infection, purified Stx) together with the reported renal phenotype [18,[20][21][22][23][24][25][26][27][29][30][31][32][33]. The lack of typical glomerular pathological changes of TMA in these studies is striking.…”

“…Do the typical glomerular lesions seen in human TMA ever develop in Stx-treated mice? Only a few studies have reported glomerular lesions that reflect some of the typical changes seen in human renal TMA [24][25][26][27]33]. Importantly, some of these were provoked under special circumstances [24,33] and in none did the experimental animals develop the complete triad of thrombocytopenia, glomerular TMA and MAHA that comprise HUS.…”

“…Furthermore, the toxic effects of Stx are enhanced by concomitant administration of lipopolysaccharide (LPS) [19]. Hence, purified Stx-2, sometimes in combination with LPS, has been used by several groups in an attempt to induce HUS in mice [20][21][22][23][24][25][26][27]. Therefore, to test our hypothesis, we chose to treat Cfh +/mice with purified Stx-2 in the presence of LPS.…”

Advances in understanding stroke risk in children – a geneticist's view

“…Translating genetic knowledge to clinical practice: ADAMTS13 as an example ADAMTS13, also known as von-Willebrand-factor (VWF) cleaving protease, is an established factor in blood coagulation and stroke (Chauhan et al, 2006). ADAMTS13 reduces adhesion and aggregation of platelets (Motto et al, 2005;Chauhan et al, 2008) and down regulates thrombus formation and inflammation (Motto et al, 2005). Reduction or absence of ADAMTS13 activity results in impaired degradation of ultra-large VWF multimers and leads to excessive VWF-induced platelet aggregation on the endothelium.…”