Aggregation of human platelets in plasma by porcine blood cells in vitro is probably mediated by thrombin generation.

The infusion of pig progenitor cells into baboons is associated with a thrombotic microangiopathy probably related to the interaction of these cells with the baboon endothelial cells and platelets. We have shown previously that pig peripheral blood mononuclear cells (p-PBMC), are able to activate the human coagulation cascade as they are able to generate thrombin when added to defibrinated plasma. In this work, we have tested the interaction of p-PBMC with human platelets to assess the capacity of p-PBMC to cause platelet aggregation and the possible role of complement activation in this aggregation. Human platelet aggregation assays, using collagen (1 or 2 microg/ml), were performed with platelets in platelet-rich plasma (PRP) or platelets washed by filtration. PRP or washed platelets were also incubated with p-PBMC or human PBMC (h-PBMC) at several concentrations and aggregation was measured. The effect of Dansylarginine N-(3-ethyl-1,5-pentanediyl)amide (DAPA), an inhibitor of thrombin, was studied on platelet aggregation caused by the pig cells. Complement activation was measured by deposition of fragment c derived from C3 splitting (C3c) on pig cells incubated with citrated platelet poor plasma (PPP). When human PRP was incubated with p-PBMC, aggregation was a consistent event quantitatively similar to that induced by collagen. No aggregation of washed platelets was observed when these were incubated with p-PBMC or h-PBMC. Aggregation of human platelets in PRP, induced by p-PBMC, was inhibited when DAPA (100 microm) was added to the incubation mixture (23%), indicating that the thrombin inhibitor blocked the capacity of p-PBMC to aggregate human platelets. No deposition of C3c fragments on p-PBMC was detected when the porcine cells were incubated for up to 20 min with citrated PPP. The fact is that p-PBMC induces human platelet aggregation in plasma being thrombin generation a likely explanation for this observation. Our data suggest that, in the system assayed, complement activation is not a cause of platelet aggregation. These findings are relevant for the clarification of the reported thrombotic microangiopathy complicating the intravenous infusion of pig cells in primates in attempts to induce pig tolerance in baboons.