(240) Type 2 conventional dendritic cells mediate FVIII recognition in FVIII immune response

Introduction/Rationale: Hemophilia A is an X-linked bleeding disorder caused by mutations in the F8 gene, leading to a deficiency of coagulation factor VIII (FVIII). However, FVIII infusions used for the treatment of bleeding can trigger the development of neutralizing FVIII antibodies. Although several studies have demonstrated that various immune cells, such conventional dendritic cells (cDCs), macrophages, monocytes, and marginal zone B cells, co-localize or interact with FVIII, the primary immune cell population mediating FVIII immune recognition remains poorly explored. We hypothesize that type 2 cDCs (cDC2) mediate FVIII recognition and CD4+ T cell activation that enhances FVIII immunity.

Methods: Batf3⁻/⁻ mice (cDC1 knockout [KO]), Zeb2-Delta3⁻/⁻ mice (cDC2 KO), and Zbtb46-DTR mice without diphtheria toxin depletion (WT) were utilized to isolate bone marrow-derived dendritic cells (BMDC) for FVIII uptake. cDC1 ko mice received five doses of recombinant human FVIII or normal saline via retro-orbital (RO) injection. CD4⁺ T cells from immunized mice were co-cultured with BMDCs isolated from naïve cDC1 KO, cDC2 KO, and WT mice and labeled with CellTrace Violet (CTV) to assess CD4+ T cell proliferation. Plasma samples were collected to measure anti-FVIII antibody levels by ELISA.

Results: Immature BMDC endocytosed more FVIII than mature and intermediate-mature BMDC as expected. However, immature BMDC from cDC2 KO mice exhibited reduced FVIII uptake compared to those from WT and cDC1 KO mice. Although BMDC from cDC2 KO mice enhanced CD4+ T cell proliferation compared to WT mice and cDC1 KO mice, cDC2 KO mice treated with FVIII exhibited reduced anti-FVIII antibodies than WT mice and cDC1 KO mice.

Conclusion: Together, these data suggest that cDC2 predominantly contributes to FVIII-specific adaptive immunity through promotion of FVIII uptake and anti-FVIII antibodies formation.