Anti–Interleukin 6 Receptor Antibodies Attenuate Antibody Recall Responses in a Mouse Model of Allosensitization

BACKGROUND: Interleukin (IL)-6 is a regulatory cytokine for T helper type 17 (Th17) and Treg cells and a potent stimulus for B/plasma cells. The current study evaluated the effect of IL-6 receptor (IL-6R) blockade with an antiYIL-6R monoclonal (mMR16-1) in alloantibody recall responses. METHODS: A mouse model of human leukocyte antigen (HLA).A2 sensitization was used for studies to evaluate the efficacy of antiYIL-6R on alloantibody recall responses and to examine the impact of IL-6R blockade on Th17, Treg, follicular T helper (Tfh) and plasma cells using multiparameter flow cytometry, flow antibody binding, and enzyme-linked immunospot (ELISpot) assay. RESULTS: Re-exposure of C57BL/6 mice to HLA.A2(+) skin allografts resulted in a surge of donor-specific (antiYHLA.A2) immunoglobulin (Ig)G antibodies. AntiYIL-6R treatment significantly decreased but did not eliminate alloantibody responses (IgG mean fluorescence intensity, 486 T 153 vs. control 792 T 193, P = 0.0076). Flow cytometry analysis showed that antiYIL-6R treatment resulted in reduction of IL-21+CD4+ (Th17) cells (P = 0.006 vs. control) and CXCR5(+)CD4(+) Tfh cells (P = 0.04), but increased foxp3(+)CD4(+) (Treg) cells in the CD4(+) population (P =0.04 vs. control). The IgG ELISpot experiments showed a significant reduction of IgG spots in the bone marrow and the spleen cells from the antiYIL-6RYtreated mice. In vitro treatment of mouse hybridoma (PA2.1) cultures with antiYIL-6R decreased IgG spot formation but had limited effect on cell proliferation. CONCLUSION: The data indicate that antiYIL-6R therapy attenuates alloantibody recall responses by modulating a number of immune regulatory and effector cells, including Th17, Tfh, Treg, and importantly, the long-lived plasma cells in the bone marrow.