TLR9-mediated activation of the NLRP3 inflammasome in Fusarium solani-induced keratitis.

Toll-like receptor 9 (TLR9) is a crucial innate immune receptor that recognises pathogenic DNA and initiates downstream inflammatory signaling, yet its function in fungal keratitis remains poorly defined. This study aimed to clarify the specific role of TLR9 in Fusarium solani (F. solani) -induced keratitis and to provide mechanistic insights for targeted therapy. A murine model of F. solani keratitis was established to examine the expression and regulation of TLR9 and NLRP3. RNA sequencing, quantitative real-time polymerase chain reaction (qRT-PCR), Western blotting, and immunofluorescence were used to assess gene and protein expression. The activities of TLR9 and NLRP3 were modulated by adeno-associated virus short hairpin RNA (AAV-shRNA), small interfering RNA (siRNA), and pharmacological agents (Nig and MCC950). Disease severity and histopathological alterations were evaluated by slit-lamp examination, clinical scoring, and hematoxylin-eosin staining, while protein interactions were analysed through molecular docking and immunocolocalization. TLR9 and NLRP3 were markedly up-regulated in F. solani-infected corneas. TLR9 silencing suppressed nuclear factor κB (NF-κB) and NLRP3 activation, thereby mitigating corneal inflammation, reducing stromal oedema, limiting inflammatory cell infiltration, and lowering IL-1β production. Pharmacological inhibition of NLRP3 with MCC950 produced similar protective effects, whereas activation by nigericin reversed these benefits. In conclusion, TLR9 functions as a pivotal upstream regulator of the NLRP3 inflammasome in F. solani keratitis. These findings uncover a mechanistic link between TLR9 signaling and inflammasome-mediated inflammation, highlighting TLR9 as a potential immunomodulatory target for fungal keratitis therapy.