Porcine PD-L1: cloning, characterization, and implications during xenotransplantation

Abstract

BACKGROUND: Effective intervention achieved by manipulating cell-mediated xenogeneic immune responses would critically increase the clinical feasibility of xenotransplantation as immediate hyperacute rejections become controllable through genetic modulations of donor organs. Endogenous negative regulatory signals like the programmed death 1 (PD-1)-programmed death ligand 1 (PD-L1) system are candidate targets for the control of cell-mediated xenogeneic immune response. METHODS: A porcine PD-L1 molecule was cloned using RACE (rapid amplification of cDNA ends) technology based on the human PD-L1 sequence. The functional effects of cloned porcine PD-L1 were tested on human CD4(+) T cell activation using porcine PD-L1-transfected bystander cells. Cellular proliferation was monitored by [3H] thymidine incorporation, and human T cell apoptosis was measured by flow cytometry. RESULTS: Porcine PD-L1 (GenBank accession number AY837780) was found to have 73.8% sequence homology with human PD-L1 and to contain two immunoglobulin domains in its extracellular region. Moreover, porcine PD-L1 expressed on Chinese hamster ovary (CHO) cells inhibited human CD4(+) T cell proliferation stimulated with anti-CD3 only or anti-CD3 plus anti-CD28. Percentages of apoptotic activated human T cells increased by over 30% in the presence of porcine PD-L1/CHO cells, and the addition of recombinant human PD-1-Fc fusion proteins during human T cell activation reversed the inhibitory effects of porcine PD-L1. CONCLUSIONS: Cloned porcine PD-L1 showed high sequence homology with human PD-L1 and a similar molecular structure. Moreover, porcine PD-L1 inhibited human CD4(+) T cell activation in human PD-1-dependent manner, and this involved activated T cell apoptosis. The authors suggest that PD-1-PD-L1 might play an important endogenous immune regulatory role during xenogeneic transplantation, and that the effective application of this system would improve transplanted xenogeneic organ survival.