Introduction: An increased risk of acute antibody-mediated rejection (ABMR) has been described after blood group incompatible (ABOi) renal transplantation. To detect long-term effects of rituximab induction on graft outcome in ABOi renal transplantation, we analyzed HLA antibody formation, clinically relevant immune parameters and protocol biopsies in a prospective renal transplant study up to 5 years posttransplant.
Methods:  Mononuclear cell subsets (peripheral blood; protocol biopsies (n=58, 3 months; n=34, 1 year)) and in-vitro T and B cell responses were assessed up to 5 years posttransplant in 85 renal transplant recipients (living donation: n=25 ABO incompatible (ABOi; with rituximab induction) and n=30 ABO compatible (ABOc); deceased donation (DD): n=30, ABO compatible). IgG anti-HLA antibodies were assessed by single antigen assay in ABOc and ABOi recipients.
Results: In ABOi patients, an increased frequency of biopsy-proven acute rejection was found only in the 3-12 month posttransplant period (6/24 (25%) versus 1/30 (3%), ABOc and 0/30 (0%), DD; P=0.003). A significantly increased frequency of ABMR but not of acute cellular rejection was detected at 5 years in ABOi (4/24, 17%) compared to non-rituximab treated ABOc (0/25) and DD patients (0/27; P=0.008).      
After rituximab induction in ABOi recipients, peripheral blood B cell subsets were profoundly downregulated for at least 3 years, most pronounced with regard to memory B cells, together with impaired in-vitro B cell responses (P=0.010, T-dependent; P=0.053, T-independent), but increased CD4 helper activity compared to ABOc patients at 2 years (P=0.019).
Cell subset analysis in protocol biopsies showed rituximab-induced B cell depletion in ABOi patients at 3 months (P<0.001 vs. ABOi and DD), but comparable B cell counts and even enhanced counts of CD3+ T cells (P=0.041), CD68+ macrophages (P=0.021) and CD138+ plasma cells (P=0.033) at 1 year. The degree of interstitial fibrosis and tubular atrophy was significantly enhanced in ABOi and DD compared to ABOc patients (26±6%, ABOi, 8±3%, ABOc, 22±4%, DD; P=0.009).
IgG anti-HLA antibody formation was not significantly different between ABOi and ABOc patients up to 5 years posttransplant. De novo formation of donor-specific HLA antibodies (DSA) occurred in 1/30 (3%) ABOc and 2/23 (9%) ABOi patients (P=0.0573).  
Conclusion: Rituximab induction induced a long-term profoundly delayed B cell repopulation together with compromised B cell responses in ABOi patients. This resulted in complete B cell depletion in  protocol graft biopsies at 3 months only but even upregulated T cell, macrophage und plasma cell counts at 1 year coinciding with an increased acute rejection frequency between 3 and 12 months posttransplant. Rituximab induction in ABOi patients did not affect IgG anti-HLA antibody formation.