Colloidal Confinement of Polyphosphate on Gold Nanoparticles Robustly Activates the Contact Pathway of Blood Coagulation

Abstract

Platelet-sized polyphosphate (polyP) was functionalized on the surface of gold nanoparticles (GNPs) via a facile conjugation scheme entailing EDAC (N-(3-(dimethylamino)propyl)-N'-ethylcarbodiimide hydrochloride)-catalyzed phosphoramidation of the terminal phosphate of polyP to cystamine. Subsequent reduction of the disulfide moiety allowed for anchoring to the colloidal surface. The ability of the synthesized polyP-GNPs to initiate the contact pathway of clotting in human pooled normal plasma (PNP) was then assayed by quantifying changes in viscous, mechanical, and optical properties upon coagulation. It is revealed that the polyP-GNPs are markedly superior contact activators compared to molecularly dissolved, platelet-sized polyP (of equivalent polymer chain length). Moreover, the particles' capacity to mobilize Factor XII (FXII) and its coactivating proteins appear to be identical to very-long-chain polyP typically found in bacteria. These data imply that nanolocalization of anionic procoagulants on colloidal surfaces, achieved through covalent anchoring, may yield a robust contact surface with the ability to sufficiently cluster active clotting factors together above their threshold concentrations to cease bleeding. The polyP-GNPs therefore serve as a promising foundation in the development of a nanoparticle hemostat to treat a range of hemorrhagic scenarios.

Neutralizing blood-borne polyphosphate in vivo provides safe thromboprotection

“…PolyP nanoparticles are stable in physiologic buffers for several hours 15 and may precipitate on the platelet surface to provide the ideal substrate for efficient FXII contact activation. Similar to these polyP aggregates, SC polyP conjugated with colloidal gold nanoparticles activates FXII with potency equivalent to that of LC polyP 47 . In sum, the data indicate that the localization, composition and functions of platelet polyP are regulated on various levels that warrant further in vivo analysis.…”

Factor XII Contact Activation

“…Supporting the hypothesis that aggregate formation is critical for polyP activity, short-chain polyP conjugated to colloidal gold nanoparticles is as potent as long-chain polyP for FXII contact activation. 31 Platelets store polyP in their dense granules and these organelles are Ca 2þ -rich (molar range). Platelet-derived polyP binds to Ca 2þ and Ca 2þ -polyP readily forms stable nanoparticles, suggesting that polyP mostly operates as particles on platelet surfaces rather than in soluble form in the supernatant.…”

Inorganic Polyphosphates Are Important for Cell Survival and Motility of Human Skin Keratinocytes and Play a Role in Wound Healing

“…Inorganic polyP shows promise in different phases of wound healing, including hemostasis and re-epithelialization, as polyP is a normal component of different cells that play a role in this process, including platelets, dermal fibroblasts, and keratinocytes. The use of polyP as a therapeutic for acute and chronic wounding has begun to garner interest, in part because of experiments elucidating its role in wound healing [33], as well as in hemostasis [69,[94][95][96][97][98][99]. We have recently shown a role for polyP in the response to UV survival, cell motility, and wound healing [33].…”