B Cells Drive Chronic Lung Allograft Rejection after Ischemia-Reperfusion Injury in Mouse Lung Transplantation

Purpose Chronic lung allograft dysfunction (CLAD) is the primary obstacle to long-term survival after lung transplantation (LTX). Ischemia-reperfusion injury (IRI) increases the risk of CLAD in humans but the underlying mechanisms are not well understood. In a mouse minor alloantigen mismatched LTX model (C57BL/10 [B10] →C57BL/6 [B6]), we observed that IRI augments formation of B cell-rich tertiary lymphoid organs (TLOs) and chronic rejection in allografts but not isografts. We therefore hypothesized that B cells may link IRI with chronic rejection. Methods B10→B6 LTX with extended graft preservation (6 hours at 4°C followed by 45 minutes at 37°Cand 15 minutes at room temperature during anastomosis), which augments acute rejection, fibrosis, and TLO formation, was employed. Anti-CD20 antibody (aCD20: n=9) or isotype control (Iso: n=10) were given i.p. on days -2, +2, +7, +14, and +21. Peripheral blood was longitudinally assessed by flow cytometry after LTX. Grafts were assessed with histology, flow cytometry and immunofluorescence 28 days after LTX. Results aCD20 treatment effectively depleted CD19+ or B220+ B cells in peripheral blood (Figure A) and lung grafts (Figure B). Acute rejection (ISHLT A grade), peribronchial fibrosis, and airways with obliterative bronchiolitis (%OB) were lower in aCD20 mice (1.8, 1.4, and 13.1% respectively) than in Iso mice (3.1, 2.6, and 46.7% respectively) (Figure C). TLOs containing CD3 + , CD19 + , and peripheral node addressin (PNAd) + cells were also reduced (Figure D). Conclusion In this model of IRI-augmented chronic rejection, B cell depletion attenuated acute rejection, airway fibrosis, and lymphoid neogenesis. Our observations suggest that B cell activation and recruitment may be directly involved in the augmentation of CLAD in LTX recipients experiencing IRI.