Cyclosporine A reprograms corneal epithelial TLR2 signaling to suppress dendritic cell maturation in dry eye inflammation

This study aimed to elucidate the immunomodulatory mechanism of Cyclosporine A (CsA) in dry eye disease (DED), focusing on its action on corneal epithelial cells (CECs) and the subsequent regulation of dendritic cell (DC) maturation and T-cell activation via the TLR2-MyD88/NF-κB signaling pathway. In vitro experiments utilized an immortalized mouse CECs and bone marrow-derived DCs. CECs were treated with a range of CsA concentrations (0-120 mg/L), followed by for TLR2 pathway analysis. TLR2-knockdown CECs were co-cultured with immature DCs to assess DC maturation markers (flow cytometry for CD80, CD86, MHC-II), cytokine secretion, and DC-mediated allogeneic T-cell proliferation. CsA exposure dose-dependently and potently suppressed TLR2-MyD88/NF-κB signaling in CECs. CsA-treated CECs suppressed DC maturation and reduced the secretion of pro-inflammatory cytokines (IL-6, IL-12, TNF-α, IFN-γ). Consequently, these DCs exhibited a diminished capacity to stimulate T-cell proliferation. Knockdown of TLR2 showed the similar results. However, the inhibitory effect of CsA on DC maturation markers was lost and potentially reversed to a stimulatory phenotype upon TLR2 knockdown in CECs, indicating CEC TLR2 is a primary target for CsA's immunomodulatory effect. Our findings unveil a novel innate immunomodulatory mechanism for CsA in DED, whereby it acts on CECs to suppress TLR2-MyD88/NF-κB signaling, thereby inhibiting DC maturation and downstream T-cell activation. This pathway complements CsA's canonical T-cell suppression and provides a mechanistic basis for its immunomodulatory efficacy.