Page 275 - Haematologica April 2020

Inflammation and TTP
thrombus formation in our study. In the absence of circu- lating VWF, thrombocytes fail to adhere to the fibrillar col- lagen surface under flow and thrombus formation is sig- nificantly impaired in the small vessels after injury. During the revision of this manuscript, similar hemostatic defects were reported to occur when vwf is independently knocked out.44 These findings support the crucial physio- logical function of zebrafish VWF in mediating thrombo- cyte adhesion/aggregation and thrombus formation.
The A13 protein in zebrafish is also conserved with a similar domain structure to that in mammals except for a longer signal peptide and propeptide. There is ∼52% sequence similarity at the A13 protein level between zebrafish and humans (or mice). Surprisingly, zebrafish plasma A13 is able to cleave the human FRETS-rVWF7345 at the similar rate as human plasma A13 does. This activ- ity is abrogated by EDTA, which chelates divalent metal ions, suggesting that zebrafish A13 is also a metallopro- tease. Additional evidence to support this notion is that plasma VWF multimers and thrombocyte-decorated VWF strings on the fibrillar collagen surface under flow are sig- nificantly increased in a13-/- zebrafish when compared with those in wt controls. The rate of thrombus formation in the caudal venules after oxidative injury is also dramat- ically accelerated in a13-/- zebrafish. More interestingly, a13-/- zebrafish develop a spontaneous but non-fatal TTP phenotype, similar to that in some patients with congeni-
tal TTP20,46 and in a baboon model of acquired TTP induced by passive immunization.25 Together, these results suggest that mild TTP may be compatible with life, but the affected individuals may have a significantly increased risk of developing catastrophic or potentially fatal TTP when an additional environmental trigger, such as infection or pregnancy, is present.
Infection or systemic inflammation often precedes the onset of TTP, which is thought to activate neutrophils, resulting in the release of their intracellular contents, including antimicrobial HNP1-3, proteolytic enzymes, nucleic acids, and histone-DNA complexes.29,42 Plasma lev- els of histone-DNA complexes are markedly elevated in patients with acute TTP.40 Although the causes of elevated levels of plasma histone-DNA are not fully understood. Histone-DNA has been shown to have deleterious effects with prothrombotic properties. Histones may activate platelets, enhance thrombin generation,47 induce exocyto- sis of endothelial Weibel-Palade bodies, and trigger the release of ULVWF from endothelium.42 Histones may also activate complement, resulting in the formation of a mem- brane attack complex that injures endothelium.48 Based on our results and those published in the literature, we hypothesize that histones, likely derived from NETosis and other cell death after injury, are potent triggers for TTP. When plasma A13 activity is present, the released ULVWF multimers are rapidly cleaved into smaller forms.
Figure 8. A proposed working model depicts how neutrophil activation and the release of histones induce microvascular thrombosis in a13-/- zebrafish. Infection and/or systemic inflammation may activate neutrophils and result in neutrophil death and tissue injury, leading to the release of their cellular contents including histone/DNA complexes or free histone. Histone/DNA complexes or free histone may then bind to endothelial receptors, which triggers exocytosis of Weibel-Palade bodies and the release of ultralarge multimers of von Willebrand factor (UL-VWF) from activated endothelium. These hyperactive UL-VWF multimers are rapidly removed by plasma ADAMTS13, so that normal hemostasis is maintained. However, when plasma ADAMTS13 is absent, as in cases of hereditary or acquired throm- botic thrombocytopenic purpura (TTP), the UL-VWF strings are not removed from the endothelial surface, resulting in an accumulation of platelets/thrombocytes and neutrophils, leading to thrombus formation and acute inflammation at sites of vascular injury. In the end, activation of the coagulation cascade, complement, and neutrophils may occur, resulting in further tissue damage and microvascular thrombosis, the characteristic pathological feature of TTP.
haematologica | 2020; 105(4)
1117