Skin xenotransplantation: Historical review and clinical potential

Abstract

Half a million patients in the USA alone require treatment for burns annually. Following an extensive burn, it may not be possible to provide sufficient autografts in a single setting. Pig skin xenografts may provide temporary coverage. However, preformed xenoreactive antibodies in the human recipient activate complement, and thus result in rapid rejection of the graft. Because burn patients usually have some degree of immune dysfunction and are therefore at increased risk of infection, immunosuppressive therapy is undesirable. Genetic engineering of the pig has increased the survival of pig heart, kidney, islet, and corneal grafts in immunosuppressed non-human primates from minutes to months or occasionally years. We summarize the current status of research into skin xenotransplantation for burns, with special emphasis on developments in genetic engineering of pigs to protect the graft from immunological injury. A genetically-engineered pig skin graft now survives as long as an allograft and, importantly, rejection of a skin xenograft is not detrimental to a subsequent allograft. Nevertheless, currently, systemic immunosuppressive therapy would still be required to inhibit a cellular response, and so we discuss what further genetic manipulations could be carried out to inhibit the cellular response.

Keywords: Burns; Genetic-engineering; Pig; Skin; Xenotransplantation.

Figure 1. Human IgM and IgG antibody binding to pig and human aortic endothelial cells (AECs) by flow cytometry

Human IgM (a) and IgG (b) binding to GTKO/hCD46 pig AECs was significantly decreased compared with wild-type (WT) pig AECs (*P<0.05), and was further decreased to GTKO/hCD46/NeuGcKO pig AECs (*P<0.05). There was significantly greater IgM binding to GTKO/hCD46/NeuGcKO pig AECs than human AECs (‡P<0.05), but there was no statistical significance of the extent of IgG binding between them. (MFI=mean fluorescence intensity; ns=not significant.) (Reproduced with permission from Wijkstrom M, et al. Kidney Int. 2017;91:790–796.)

Figure 2. (A) Human CD4+T cells and CD8+Tcells responses to WT and GTKO porcine aortic endothelial cells (pAECs) before and after activation by pIFN-γ, and (B) human peripheral blood mononuclear cell (PBMC) responses to WT and GTKO pAECs expressing hCD46

(A) The proliferative response of human CD4⁺T cells (n=3) and CD8⁺T cells (n=3) to WT and GTKO pAECs before and after activation by pIFN-γ. The response was significantly less to GTKO pAECs before (p<0.001) and after (p<0.05) activation. ³H incorporation values are presented as CPM. Data represent the mean (+/− SEM) and are representative of three different experiments. (Reproduced with permission from Wilhite T, et al. Xenotransplantation. 2012;19:56–63.) (B) Human cellular responses to pAECs expressing hCD46. Human PBMCs were co-cultured with pAECs. The mean of triplicate results was expressed as 3H-thymidine uptake. Data are representative of two different humans. PBMC proliferation is presented as counts per minute (CPM) for 3H incorporation. Data represent the mean (±SEM) and are shown. (*P<0.05). (Reproduced with permission from Ezzelarab M, et al. Xenotransplantation. 2015;22:487–489.)

Figure 3. Costimulation pathways in T cell regulation

Upon MHC-antigen interaction with the TCR, costimulation pathways can augment or suppress the activation of the T cell. From left to right: (i) CD28 is activated by CD80/CD86, after which CTLA-4 is upregulated and, with higher affinity than CD80/CD86, binds to CD28, inhibiting the signal. CTLA-4Ig and belatacept work by taking advantage of their higher affinity to CD28 over CD80/CD86, and thereby block CD80/CD86 activation of CD28. (ii) The CD40/CD154 pathway is another potent activator of T cells. Monoclonal antibodies against either of these surface proteins have potential for application in transplant immunosuppression. (iii) PD-1 is expressed on T cells, and its interaction with PD-1 Ligand (PD-L1) induces a suppressive signal to the T cell.

Figure 4. Significant reduction of the human CD4⁺T cell response to dominant-negative mutant class II transactivator (CIITA-DN) porcine aortic endothelial cells

Human (h) CD4⁺T cells were co-cultured with pAECs for 7 days (n=6). The response of hCD4⁺T cells was evaluated by [3H] thymidine incorporation. As a negative control, hCD4⁺T cells were cultured with culture medium (spont). There was a significantly lower hCD4⁺T cell response to CIITA-DN pAECs compared with WT pAECs (**P<0.01). (Reproduced with permission from Hara H, et al. Immunology. 2013;140:39–46.)