Adjuvant effects of peptidoglycan on CD4 +T cell activation by MHC-II +fibroblast-like synoviocytes

Abstract

Abstract Lyme arthritis (LA) is the predominant late-stage manifestation of Lyme disease, caused by tick-borne spirochete Borrelia burgdorferi. LA can resolve with antibiotic therapy or persist after treatment in a subset of patients, known as post-antibiotic LA. Post-antibiotic LA is characterized by proliferative synovitis, dysregulated Th1/ IFNɣ responses and MHC-II+ fibroblast-like synoviocytes (FLS), a common cell type in inflamed joints. High levels of B. burgdorferi peptidoglycan (PG) have been found in LA patient synovia, yet the effects of PG on host immune responses are not well characterized. We hypothesize that PG acts as an adjuvant to enhance antigen presentation and CD4 +T cell activation in the joint. To test this, primary murine macrophages and FLS, both abundant in LA synovia, were treated in vitro with IFNɣ, PG, or both. Stimulation of macrophages led to upregulation of MHC-II in an IFNɣ-dependent manner, as expected. FLS stimulated with IFNɣ led to upregulation of MHC-II and production of proinflammatory cytokines, which was enhanced by PG as well as the muramyl-dipeptide subunit of PG, a known NOD2 ligand. To examine T cell responses, IFNɣ-primed MHC-II +FLS were co-cultured with OT-II mouse T cells (all with identical antigen-specific T cell receptors) and supplemented with the cognate OVA 323–339peptide. MHC-II +FLS activated naïve CD4 +T cell proliferation, which was enhanced by PG stimulation. Co-culture supernatants showed upregulated IFNɣ and IL-12 production, indicating a Th1 response. Negative controls using an alternative OVA 257–264peptide or NOD2-deficient FLS failed to induce T cell responses. These data show that PG acts as an immune adjuvant enhancing peptide-specific CD4 +T cell activation by MHC-II +FLS.