Background: Uncontrolled complement activation causes exacerbated inflammatory responses and tissue damage in both allo- and xenotransplantation. Therefore, fine-regulation of the complement system is the key to achieving long-lasting graft transplantation. To date, few therapies have targeted the regulation of complement. Here, we investigated the effect of a lyophilized, gamma-irradiated, viral cleared, apoptotic peripheral blood mononuclear-cell secretome (APOSEC) on complement activation and deposition on endothelial cells in an in vitro xenotransplantation model.
Method: Complement activation was achieved by co-treating wild-type porcine aortic endothelial cells (wt-PAECs) with 10% pooled normal human serum. Cells were co-treated either with buffer (as control) or with three concentrations of APOSEC (1, 2.5, or 10 U/ml). Complement deposition was then assessed by immunofluorescence staining and quantified by cell ELISA for C3b/c, C4b/c and C5b-9. The effect of APOSEC on the formation of the anaphylatoxins C3a and C5a was measured by supernatant ELISA. The impact of APOSEC on cell viability was analysed by flow cytometry. 
Results: Co-incubation of human serum treated wt-PAECs with 10 U/ml APOSEC significantly reduced complement deposition. The binding of C3b/c, C4b/c, and C5b-9 to the endothelial cells was reduced by up to 80%. Moreover, lower levels of C3a (36%) and C5a (76%) anaphylatoxins in the cell culture supernatant were detected. Co-incubation of cells with 10 U/ml of APOSEC increased cell viability by more than 50%.
Conclusion: Our data indicate that APOSEC can reduce complement activation, leading to a reduction in cellular deposition and an increase in endothelial cell viability. The observed complement modulation suggests that APOSEC could be a novel, clinically available and relatively cheap therapy for complement-related graft rejection events both early (ischemia/reperfusion injury) and later in allo- and xenotransplantation (vascular rejection, thrombotic microvascular angiopathy).