T cell responses to corneal xenogeneic grafts using genetically-engineered pig corneas

Hidetaka Hara (1), Whayoung Lee (1), David Ayares (2), David K. C. Cooper (1).

(1)Thomas E. Starzl Transplantation Institute, University of Pittsburgh, Pittsburgh, PA, USA, (2) Revivicor, Blacksburg, VA, USA

The immune-privileged environment provides a corneal xenograft with some protection and, as the cornea is avascular, hyperacute rejection is not seen. The results of wild-type (WT) pig corneal transplantation (Tx) in monkeys (reported from Korea and China) have been encouraging.

The T cell response to WT pig corneas

Our own in vitro studies, however, indicate there is a significant human T cell response to WT pig corneal endothelial cells (CECs), with a proliferative response on MLR.

Rodent studies from several groups have demonstrated that rejection is related primarily to CD4⁺T cells and macrophages. However, in pig-to-monkey corneal Tx, CD8⁺T cells appear to be more prominent, with CD20⁺B cells also playing a role. Graft infiltration was predominantly with CD8⁺T lymphocytes and macrophages, with minimal CD4⁺T and CD20⁺B cells. When the graft was rejected, an increase in CD8⁺ effector memory T cells was documented in the blood. Anti-donor and anti-Gal antibodies were increased. 

As CECs are a target for the host T cell response, anterior lamellar keratoplasty (ALK) reduces the immune response. Survival after pig ALK in monkeys was longer (>3 months) than that after full-thickness corneal grafts (PK) (+/-2 weeks). With corticosteroid therapy, PK grafts from WT pigs survived in monkeys for >6 months. Our own data in monkeys treated with corticosteroids, however, indicate that there is an elicited IgG response within days of Tx, suggesting sensitization, which would be expected to result in rapid graft failure.

Corneal decellularization reduces antigenicity by removing stromal cells, which play a role in sensitizing the host, and reducing the expression of pig antigens (e.g., Gal, NeuGc). Using decellularized WT pig grafts, ALK has significantly longer graft survival than untreated corneas.  

The T cell response to genetically-engineered pig corneas

In view of the strength of the immune response to WT pig corneas, corneas from genetically-engineered pigs are likely to be necessary to protect them from rejection, particularly as long-term immunosuppressive therapy is contraindicated. 

            In vitro, the human CD4⁺T cell response to CECs from α1,3-galactosyltransferase gene-knockout (GTKO) or GTKO/hCD46 pigs is significantly weaker than to WT CECs. Furthermore, CECs from human mutant dominant-negative class II transactivator (CIITA) transgenic pigs can suppress swine leukocyte antigen (SLA) class II expression even when the CECs are activated, resulting in reduced human T cell proliferation in vitro on MLR. Corneal Tx from GTKO/CD46/CIITA-DN pigs should therefore be associated with a reduced primate cellular response.

            In conclusion, WT pig corneas will induce both humoral and cellular immune responses, particularly in high-risk patients with inflamed and/or vascularized corneal beds. In our opinion, genetically-engineered pigs will be essential if clinical corneal Tx is to prove fully successful.

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Sources of funding: NIH, Revivicor.