Putting polyphosphates to the test: evidence against platelet-induced activation of factor XII

Abstract

Key Points• Coagulation factor XII is not activated by platelets.The recent claim that stimulated platelets activate the intrinsic pathway of coagulation by the release of polyphosphates has been considered a breakthrough in hemostasis research. In little more than 3 years, the original publication by Müller et al has been cited >100 times. However, none of the citing articles has sought to independently validate this potentially paradigm-shifting concept. To this end, we performed extensive experimentation in vitro and in vivo in an attempt to verify the claim that factor XII (FXII) is primarily activated by stimulated platelets. In contrast to the original assertion, platelet-derived polyphosphates were found to be weak activators of FXII, with a FXIIa-generating activity of <10% compared with equivalent concentrations of kaolin. Using different coagulation assays, it was shown that platelet contribution to whole blood coagulation was unrelated to the generation of activated FXII in vitro. Additionally, key results used to verify the hypothesis in the original study in vivo were found to be irreproducible. We conclude that plateletderived polyphosphates are not physiologically relevant activators of FXII. (Blood. 2013;122(23):3818-3824) IntroductionThe enigmatic role of the intrinsic pathway of coagulation has been studied with renewed interest ever since it was shown that 1 strain of factor XII (FXII)-deficient mice are protected from thrombosis in various in vivo models, eg, in ferric chloride-induced thrombosis in mesenteric vessels, carotid vessels and inferior vena cava, and collagen-induced pulmonary embolism, [1][2][3] and that FXII inhibitors protect from arterial thrombosis in various animal models. [4][5][6] These studies provide strong evidence for a role for FXII in pathological thrombosis in mice, although clinical evidence for an analogous role in humans is still lacking.One central question that has yet to be conclusively answered in this context is how FXII is activated during thrombosis in vivo. 7 Several activators of FXII have been proposed, most notably fibrin clot surfaces, 8 collagen exposed by damaged endothelium, 9 misfolded or denatured proteins, 10 and neutrophil extracellular traps. 11 However, none of these potential activators have been validated conclusively in vivo.Recently, Müller et al 12 proposed that FXII activation in vivo is caused by negatively charged polyphosphates (polyP) secreted from platelet dense granules on platelet activation (hereinafter designated paper A). In a very recent review in Blood elaborating on these findings, 13 one of the authors of paper A asserts that "the identification of polyP as a platelet-derived procoagulant agent provides a longsought link between primary and secondary hemostasis and may represent a new paradigm for the treatment of thromboembolic and inflammatory diseases." As the implications of this claim are considerable, it is imperative to independently verify the experimental results.In the present study, we tested the following c...

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Synthetic polyphosphate inhibits endogenous coagulation and platelet aggregation in vitro

“…Previous studies have revealed that platelet-derived polyphosphate promotes blood coagulation (5,6). However, the effects obtained when adding synthetic polyphosphate have been conflicting.…”

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