Immunomodulatory Strategies Directed Toward Tolerance of Vascularized Composite Allografts

Abstract

Background: Achieving tolerance of vascularized composite allografts (VCAs) would improve the risk-to-benefit ratio in patients who undergo this life-enhancing, though not lifesaving, transplant. Kidney cotransplantation along with a short course of high-dose immunosuppression enables tolerance of heart allografts across a full major histocompatibility complex (MHC) mismatch. In this study, we investigated whether tolerance of VCAs across full MHC disparities could be achieved in animals already tolerant of heart and kidney allografts.

Methods: Miniature swine that were tolerant of heart and/or kidney allografts long term underwent transplantation of myocutaneous VCA across the same MHC barrier. Before VCA transplant, group 1 (n = 3) underwent class I-mismatched kidney transplantation; group 2 (n = 3) underwent 2 sequential class I-mismatched kidney transplantations; group 3 (n = 2) underwent haploidentical MHC-mismatched heart/kidney transplantation; and group 4 (n = 2) underwent full MHC-mismatched heart/kidney transplantation.

Results: All 3 animals in group 1 and 2 of 3 animals in group 2 showed skin rejection within 85 days; 1 animal in group 2 showed prolonged skin survival longer than 200 days. Animals in groups 3 and 4 showed skin rejection within 30 days and regained in vitro evidence of donor responsiveness.

Conclusions: This is the first preclinical study in which hearts, kidneys, and VCAs have been transplanted into the same recipient. Despite VCA rejection, tolerance of heart and kidney allografts was maintained. These results suggest that regulatory tolerance of skin is possible but not generally achieved by the same level of immunomodulation that is capable of inducing tolerance of heart and kidney allografts. Achieving tolerance of skin may require additional immunomodulatory therapies.

Figure 1

Gross appearance of VCA. Representative clinical images of VCAs from Group 3 animal #21517 (top row) and Group 4 animal #22026 (bottom row) by postoperative day. Animals in both groups showed patchy areas of necrosis starting on postoperative day 7 that progressed to epidermal sloughing.

Figure 2

Histology from representative VCA biopsies taken on postoperative day 30-31. VCA biopsy from Group 3 animal #21517 (top row) shows grade 4 rejection of the epidermis and acute rejection of the muscle with endarteritis. VCA biopsy from Group 4 animal #22025 (bottom row) shows grade 4 rejection of the epidermis and acute rejection of the muscle with endarteritis.

Figure 3

MLR assays from VCA recipients. Stimulation indices to self, donor-type (SLA^ad or SLA^dd), and third-party (YO) peripheral blood mononuclear cells before VCA transplant and 30-31 days after VCA transplant. MLR, mixed-lymphocyte reaction. YO, York.

Figure 4

CML assays from VCA recipients. Percent specific lysis is plotted as a function of effector:target ratio. Response against donor-type (SLA^ad or SLA^dd) targets before VCA transplant and 30-31 days after VCA transplant. CML, cell-mediated lympholysis.

Figure 5

Alloantibody response. Levels of circulating IgM and IgG alloantibody were measured by flow cytometry in recipients in Groups 3 and 4. Data were normalized to the mean fluorescence intensity of negative control values to plot normalized mean fluorescence intensity as a function of postoperative day (POD). HK tx, heart kidney transplant.

Figure 6

Histology from representative kidney samples taken prior to VCA transplantation and after VCA rejection. Group 3 animal #21517 (top row) and Group 4 animal #22025 (bottom row) demonstrate the presence of organized lymphoid structures before and after VCA rejection.