Autoantibodies targeting the C-terminal fragment of perlecan (LG3) were identified in patients with kidney transplantation and are associated with rejection episodes. High anti-LG3 titers were also observed in patients awaiting a renal transplantation and their presence correlates with an increased risk of allograft rejection and dysfunction. The mechanisms responsible for immunization to LG3 remain largely uncharacterized.

To evaluate the importance of inflammation for anti-LG3 production prior to transplantation, wild type (WT) C57BL/6 mice and CD4+ T cell depleted mice (CD4 dep) were immunized with recombinant murine LG3 (0.05 mg/s.c.), mouse serum albumin (MSA) or PBS every 2 weeks for a total of 4 immunizations in the presence or absence of incomplete Freund's adjuvant (IFA). Anti-LG3, total IgG and anti-nuclear antibodies (ANA) titers were assessed by ELISA. The presence of memory B cells was evaluated in WT and CD4 KO mice by ELISPOT.

LG3 immunization triggered anti-LG3 production even in the absence of IFA (LG3+IFA 25/25; 100%, LG3 w/o IFA: 14/34; 41%). LG3 immunization did not modulate total IgG levels (PBS: 210+/-46 [mu]g/ml, LG3: 250+/-32 [mu]g/ml) nor ANA concentration (PBS: 44+/-12 [mu]g/ml, LG3: 57+/-15 [mu]g/ml) indicating that anti-LG3 production is not the consequence of a broad autoimmune response. MSA immunization did not induce anti-LG3 production showing that break in tolerance to LG3 is specific to LG3 immunization. CD4 dep mice immunized with LG3 failed to develop anti-LG3 antibodies. Additionally, we identified IgG+ memory B cells specific to LG3 in the spleen, bone marrow and peritoneal cavity of non-immunized WT and CD4 KO mice.

Collectively, our results demonstrate that LG3 fuels anti-LG3 production even in the absence of inflammation. The break in tolerance to LG3 is dependent on CD4+ T cells and LG3-specific memory B cells. This humoral memory is present in the normal repertoire of WT mice and is helper T cell-independent. This study helps better understand the mechanisms implicated in the presence of anti-LG3 antibodies prior to transplantation.

CITATION INFORMATION: Padet L, Dieude M, Yang B, Turgeon J, Cardinal H, Hebert M.-J. Detection of Specific B Cell Memory to the Perlecan LG3 Antigen in the Normal Immune Repertoire. Am J Transplant. 2017;17 (suppl 3).

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