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B cell depletion synergizes with ECDI-fixed rat splenocyte infusions to induce concordant rat to mouse islet xenotransplantation tolerance

Shusen Wang, Taba Kheradmand, James Tasch, Jie Yang, Xunrong Luo

Division of Nephrology and Hypertension, Northwestern University Feinberg School of Medicine, Chicago, IL, United States

Background: Previously we demonstrated that infusions of ethylene carbodiimide (ECDI)-fixed donor splenocytes (SP) could only prolong concordant rat to mouse islet xenograft survival but fail to induce tolerance. The aim of this study was to determine whether combination of B cell depletion and rat ECDI-SP infusions could induce tolerance to islet xenografts in a rat-to-mouse transplant model.

Methods: Rat ECDI-SPs were infused i.v. to diabetic C57BL/6 (B6) mice at day -7 and day 1. 650 Lewis rat islets were transplanted into the renal subcapsular space of recipient B6 mice at day 0. Recipients were treated with 250 ug anti-CD20 mAb i.v. on day -10 and day 1.

Results: Mice receiving rat ECDI-SP infusions had prolonged islet xenograft survival (median of 48, range 27-61 days) compare with control group (median of 18, range 15-25, p=0.0026). In contrast, 100% of xenografts were indefinitely accepted (> 150 days post transplant) in mice treated simultaneously with anti-CD20 and rat ECDI-SP infusions (p=0.0018, compare with rat ECDI-SP infusion group). Moderate to high levels of mouse anti-rat antibodies could be detected in subtypes IgG1, IgG2a, IgG2b, and IgG3 in the serum at two weeks after infusion of rat ECDI-SP alone despite the observed graft prolongation. Conversely, with combination therapy, minimal levels of all subtypes of anti-rat IgG were detected in the serum, which correlated with indefinite graft survival. In long-term protected islet xenografts, histological examination revealed minimal deposition of IgG, IgM, or C3, in contrast to acutely rejected grafts. Furthermore, protected islet grafts harbored a significant number of Foxp3+ cells among peri-islet graft infiltrates, which was minimal in rejected islet xenografts. Furthermore, there was a significant decrease in spleen CD4 ⁺ effector memory T cell population (CD44^highCD62L^lowCD69^low) in long-term tolerized mice compared with controls.

Conclusions: Simultaneously with anti-CD20 and donor ECDI-SP infusions may be a promising regimen for tolerance induction in xenogeneic islet transplantation.

Keywords: Xenotransplantation; Islets; B cell; Tolerance