Introduction: Antibody mediated rejection (AMR) is a major cause of kidney allograft failure. APRIL (A Proliferation Inducing Ligand) and BLyS (B Lymphocyte Stimulator) are two critical survival factors for B lymphocytes and plasma cells, the main source of alloantibody. We generated rats deficient in APRIL and BLyS to characterize the effects of targeting these cytokines in our established rodent model of AMR in kidney transplant. Here we report our initial phenotyping and response to alloantigen in these novel rodents.

Methods: Using CRISPR/Cas9 we engineered APRIL^-/- and BLyS^-/- Lewis rats. The absence of APRIL and BLyS was determined using enzyme-linked immunosorbent assay (ELISA) and reverse transcription-polymerase chain reaction (RT-PCR). Spleen, bone marrow, blood, and lymph nodes were also analyzed using flow cytometry, enzyme-linked immunospot (ELISPOT), and immunohistochemistry. APRIL^-/- and BLyS^-/- rats were sensitized with Brown Norway blood (complete MHC mismatch). DSA was measured by flow cross match against Brown Norway splenocytes from sensitized wild type (WT), APRIL^-/- and BLyS^-/- rats.

Results: BLyS^-/- significantly decreased naïve B lymphocytes (CD45R+IgD+CD24-CD38+CD27+) but increased transitional B lymphocytes (CD45R+IgD+CD24++) when compared to WT and APRIL^-/- (p<0.03). BLyS^-/- were also found to decrease marginal zone B lymphocytes (CD45RA+HIS57+) compared to APRIL^-/- and WT, however this was not significant. Additionally, BLyS^-/- significantly depleted antibody secreting B cell production of IgM and IgG in all tissues compared to WT and APRIL (p<0.04). When challenged with alloantigen, sensitized BLyS^-/- had significant decreases in DSA for IgG1, IgG2a, and IgG2b when compared to WT. There was no statistical difference in DSA between APRIL^-/- and WT.

Conclusion: BLyS^-/- produced fewer naïve B lymphocytes and were found to increase transitional B lymphocytes when compared to WT and APRIL^-/-. Additionally, BLyS^-/- produced fewer alloantibodies when challenged with alloantigen. Antibody secreting B cells were also depleted in BLyS^-/-, which translates into a reduction of alloantibody production. Future studies will characterize the effect of BLyS^-/- to prevent AMR in a kidney transplant model.

CITATION INFORMATION: Bath N., Verhoven B., Wilson N., Reese S., Coons L., Panzer S., Djamali A., Redfield R. BLyS^-/- Rats to Prevent Alloantibody Production Am J Transplant. 2017;17 (suppl 3).

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