Comparative Response of Platelet fV and Plasma fV to Activated Protein C and Relevance to a Model of Acute Traumatic Coagulopathy

Abstract

BackgroundAcute traumatic coagulopathy (ATC) has been linked to an increase in activated protein C (aPC) from 40 pM in healthy individuals to 175 pM. aPC exerts its activity primarily through cleavage of active coagulation factor Va (fVa). Platelets reportedly possess fVa which is more resistant to aPC cleavage than plasma fVa; this work examines the hypothesis that normal platelets are sufficient to maintain coagulation in the presence of elevated aPC.MethodsCoagulation responses of normal plasma, fV deficient plasma (fVdp), and isolated normal platelets in fVdp were conducted: prothrombin (PT) tests, turbidimetry, and thromboelastography (TEG), including the dose response of aPC on the samples.ResultsPT and turbidimetric assays demonstrate that normal plasma is resistant to aPC at doses much higher than those found in ATC. Additionally, an average physiological number of washed normal platelets (200,000 platelets/mm3) was sufficient to eliminate the anti-coagulant effects of aPC up to 10 nM, nearly two orders of magnitude above the ATC concentration and even the steady-state pharmacological concentration of human recombinant aPC, as measured by TEG. aPC also demonstrated no significant effect on clot lysis in normal plasma samples with or without platelets.ConclusionsAlthough platelet fVa shows slightly superior resistance to aPC's effects compared to plasma fVa in static models, neither fVa is sufficiently cleaved in simulations of ATC or pharmacologically-delivered aPC to diminish coagulation parameters. aPC is likely a correlative indicator of ATC or may play a cooperative role with other activity altering products generated in ATC.

The pathogenesis of traumatic coagulopathy

“…Tissue damage and shock are associated with platelet release of soluble CD40-ligand, a potent immune activator that itself can cause further ECA and platelet activation, and is known to be necessary in order to stabilise thrombi [29]. Immune stimulation, including complement activation, is associated with release of damageassociated molecular patterns (DAMPs), such as mitochondrial DAMPs and histone-complexed DNA [30,31]. Immune activation can aggravate tissue damage through mechanisms including proteolytic degradation and oxidative stress, thus amplifying haemostatic activation.…”

“…This mechanism is plausible but problematic due to the kinetics of the reactions. Platelets and plasma Factor Va are resistant to aPC cleavage at concentrations of aPC seen in ATC or even therapeutic use of recombinant human aPC in sepsis [31]. As a normal platelet count of 200 9 10 9 l À1 overcame aPC anticoagulant effects even at very high concentrations of aPC, and there was no detectable effect on fibrinolysis with or without platelets [31], it is difficult to envisage how aPC could drive the phenotype described as ATC.…”

“…Platelets and plasma Factor Va are resistant to aPC cleavage at concentrations of aPC seen in ATC or even therapeutic use of recombinant human aPC in sepsis [31]. As a normal platelet count of 200 9 10 9 l À1 overcame aPC anticoagulant effects even at very high concentrations of aPC, and there was no detectable effect on fibrinolysis with or without platelets [31], it is difficult to envisage how aPC could drive the phenotype described as ATC. Furthermore, though factor V is depleted and PC converted to aPC in ATC, it has been amply demonstrated that thrombin generation potential is dramatically elevated in trauma patients; this is surely inconsistent with the notion that aPC is inhibiting thrombin generation by inactivating factor V [32].…”

Validation of acute traumatic coagulopathy in an ovine model of trauma and haemorrhage

“…However the circulating concentration of PAI-1 is roughly ten times that of protein C, raising doubts about the ability of activated protein C (aPC) to deplete PAI-1 levels to the extent required to accelerate fibrinolysis (41). This has led to suggestions that ATC is a fibrinolytic form of disseminated intravascular coagulation (DIC) driven by increased tissue plasminogen activator (tPA) release from the site of injury (21,(42)(43)(44). There is also emerging evidence to implicate platelet dysfunction and endothelial glycocalyx degradation in the development of ATC (45)(46)(47)(48), although the exact contributions of these factors remains unclear.…”

“…The protein C hypothesis therefore provides an explanation for both systemic anticoagulation and hyperfibrinolysis, although the magnitude of these effects in the development of ATC has not been quantified. (98) The significance of the protein C pathway in ATC has recently been questioned by Campbell and colleagues (44) who have shown that platelet and plasma factor Va pools are resistant to cleavage by aPC in vitro, both at concentrations observed in trauma patients and following the therapeutic doses of recombinant human aPC used in sepsis. In addition they found no evidence of fibrinolysis, arguing that the high circulating levels of PAI-1 are unlikely to be inactivated to the extent required to drive the fibrinolytic picture seen with ATC.…”