In vitro bio-functional profiles and in vivo outcome of α1,3-galactosyltransferase gene-knockout miniature pig-to-nonhuman primate corneal xenotransplantation

Abstract

Purpose : To investigate bio-functional corneal profiles and outcome of α1,3-galactosyltransferase gene-knockout miniature (GTKOm) pig-to-nonhuman primate (NHP) corneal xenotransplantation.

Methods : We used 14 eyes from 3 types of 7 GTKO pigs (GTKOm, n = 4; GTKO/hCD39 knock-in miniature (KIm), n = 2; and GTKO/CMAH/iGB3s triple KO miniature (TKOm), n = 1). For in vitro study, endothelial cell density (ECD) changes and Na-and K-dependent ATPase after 7 days of preservation were evaluated. Endothelial cell proliferation was also measured. For in vivo study, 9 rhesus macaques underwent full thickness corneal xenotransplantation. Four were transplanted 2 GTKOm and 2 TKOm pig corneas and administered full dose immunosuppression (full dose group) including anti-CD20 antibody (Ab), tacrolimus, and basiliximab, which had already showed long-term graft survival in wild type miniature pig cornea. The other five were transplanted 4 GTKOm and 1 GTKO/hCD39 KIm pig corneas and administered low dose immunosuppression including tacrolimus and basiliximab (without anti-CD20 Ab). Changes in T and B cell subsets, anti-αGal, and non-αGal Abs and C3a were evaluated.

Results : ECD of all corneas maintained at more than > 2800/mm² through 7 days. ATPase pumps were well stained. Doubling time was 30.41 hours. Full dose group showed long-term graft survival (>277, >187, >187, >83 days). However, median graft survival of low dose group was 72 days (165, 91, 72, 55, 37 days) (Figure 1). T cell subsets were not changed in both groups and were not different from each other. Activated B cells in high dose group at last examination were lower than that in low dose group (P = 0.026). Anti-αGal Abs were not changed in both groups. At 4 weeks and last examination, anti-non-αGal IgG was increased from baseline (all P = 0.043) in low dose group, but not changed in high dose group. Aqueous C3a concentration at last examination in low dose group was increased from baseline (P = 0.043), and was higher than that in high dose group (P = 0.014).

Conclusions : Profiles of GTKOm pig cornea is feasible for corneal xenotransplantation. It is still important to inhibition of B cells and complement activations as well as T cell suppression even using GTKOm and/or hCD39 KI pig corneas in pig-to-NHP corneal xenotransplantation.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.

 

Representative photographs of corneal grafts after transplantation.