Inhibition of B-cell activation and antibody production by triggering inhibitory signals via the PD-1/PD-ligand pathway

Background: The development of donor-reactive antibodies is regarded to be an-important barrier limiting long-term outcome of allo-and xenografts. We asked whether enhanced signaling via the co-inhibitory receptor programmed cell death-1 (PD-1; CD279) can downregulate human B-cell activation. Methods: Proliferation of human purified CD19(+) B cells was induced by in vitro stimulation with CpG oligodeoxynucleotides (CpG-B). To induce antibody production, peripheral blood mononuclear cells were co-cultured with the porcine B-cell line L23. Triggering of inhibitory signals via the PD-1 receptor was obtained either using a -recombinant agonistic soluble ligand (PD-L1.Ig) or L23 transfectants overexpressing membrane-bound human PD-L1 (CD274; L23-PD-L1 cells). Results: Stimulation of purified CD19+ B cells with CpG--B resulted in upregulation of PD-1 and strong proliferation. Addition of PD-L1. Ig significantly reduced B-cell proliferation in a dose-dependent manner. A great proportion (similar to 1%) of human circulating B cells recognizes the epitope galactose-alpha 1,3-galactose-beta 1,4-N-acetylglucosamine-R (alpha-gal). Thus, when B cells-in the presence of T cell help-were cocultured with alpha-gal-expressing L23 cells, anti-gal and anti-L23 antibodies could readily be detected in the culture supernatant. The level of induced antibodies was significantly reduced when stimulation was performed by L23-PD-L1 cells. Conclusions: Enhancing inhibitory signals may be part of future protocols to better control humoral immunity to allo- and xenografts.