The effect of Gal expression on pig cells on the human T‐cell xenoresponse

Abstract

Background Lack of Gal expression on pig cells is associated with a reduced primate humoral immune response as well as a reduction in cytokine production by human cells in vitro. We investigated whether lack of Gal expression is associated with reduced human T-cell response in vitro. Methods Peripheral blood mononuclear cells (PBMCs) were obtained from healthy humans and naïve baboons. Human CD4+ and CD8+ T cells were isolated. Porcine aortic endothelial cells (pAECs) were isolated from wild-type (WT) and α1,3-galactosyltransferase gene-knockout (GTKO) pigs. WT pAECs were treated with α-galactosidase, reducing Gal expression. Swine leukocyte antigen (SLA) class I and II expression on pAECs was measured, as was T-cell proliferation and cytokine production in response to pAECs. Results Reduced Gal expression on WT pAECs after α-galactosidase treatment was associated with reduced human PBMC proliferation (P < 0.005). SLA class I and II expression on WT and GTKO pAECs was comparable. Human CD4+ and CD8+ T-cell proliferation was less against GTKO pAECs before (P < 0.001) and after (P < 0.01 and P < 0.05, respectively) activation. Human and baboon PBMC proliferation was less against GTKO pAECs before (P < 0.05) and after (P < 0.01 and P < 0.05, respectively) activation. Human PBMCs produced a comparable cytokine/chemokine response to WT and GTKO pAECs. However, there was less production of IFN-γ/TNF-α by CD4+ and IFN-γ/granzyme B/IP-10 by CD8+ T cells in response to GTKO pAECs. Conclusions The absence of Gal on pig cells is associated with reduced human T-cell proliferation (and possibly selected cytokine production). Adaptive primate T-cell responses are likely to be reduced in GTKO xenograft recipients.

Immunological Challenges and Therapies in Xenotransplantation

“…In particular, porcine endothelial cells constitutively express CD80/CD86 (Koshika et al 2011) and SLA class I, and possibly class II molecules as well (Choo et al 1997). Porcine endothelial cells also trigger a direct, MHC-restricted CD8 þ T-and CD4 þ T-cell activation (Yamada et al 1995;Dorling and Lechler 1998;Kim et al 2010;Koshika et al 2011;Wilhite et al 2012). High levels of cytotoxicity have also been detected, mediated mainly by CD4 þ T cells, but also by CD8 þ T cells.…”

“…Analyzing in vitro lymphocyte proliferation induced by wild-type or GalT-KO endothelial cell shows that aGal epitope expression on the endothelium is associated with a greater proliferation of CD4 þ and CD8 þ T cells (Lin et al 2008;Wilhite et al 2012), suggesting an as-yetunidentified role of aGal epitopes in sustaining T cell response. The absence of aGal is also associated with a significantly reduced secretion of INF-g, TNF-a, IL-17A by CD4 þ T cells, and of INF-g, granzyme-B, and the chemokine IP-10 by CD8 þ T cells (Wilhite et al 2012), partially confirming earlier findings reported by Saethre et al (2008), who showed that exposure to Gal þ/þ endothelial cells was associated with a significant release of human INF-g, human and porcine proinflammatory IL-6 and IL-8, and several human b chemokines, whereas this picture was not seen after exposure to Gal 2/2 cells (Saethre et al 2008). Unlike the report from Wilhite et al (2012), however, the T-cell-recruiting a-chemokine IP-10 in the Saethre study was induced in cells lacking the aGal epitopes.…”

Recent Progress in Xenotransplantation, with Emphasis on Virological Safety

“…A comprehensive overview of the experimental transplantations in the years until 2013 is given by Cooper et al [35]. To demonstrate the development in the field, the longest survival times listed by Cooper et al [35] and survival times published later [6,[36][37][38][39][40][41][42][43] are summarized in Tables 1 and 2. The ability to modify pigs genetically to protect the donor organs from the primate's immune response has resulted in survival of heterotopic pig hearts in baboons for longer than 2.5 years [39-41,44,45] ( Table 1).…”

“…Animals expressing different combinations of the complement-regulatory proteins have been created (for overview, see Table 2 in [5]). In addition, immune T cell responses were reduced by simultaneous expression of CTLA4-IgG and mutant MHC class II transactivator [6][7][8][9]. a-gal epitopes and 2 N-glycoylneuraminic acid-terminated (Neu5Gc) gangliosides are targets for pre-existing antibodies in human sera [10].…”

Absence of Immune Responses with Xenogeneic Collagen and Elastin

“…Depletion of alpha-gal epitopes by decellularization or by alpha-galactosidase treatment might reduce the humoral-and cellular-mediated immune responses. 8,52,53 Therefore, appropriate test systems for these epitopes should be applied in addition to the tests described in our study for a proper selection of suitable ECM components.…”