IL-36α Exerts Proinflammatory Effects in Aspergillus fumigatus Keratitis of Mice Through the Pathway of IL-36α/IL-36R/NF-κB

PURPOSE: To explore the role of IL-36α in corneas infected by Aspergillus fumigatus. METHODS: The experimental group was comprised of 15 corneas with fungal keratitis, and 15 healthy donor corneas were included in the control group. IL-36α was detected in normal and infected corneas of humans and C57BL/6 mice. Mice corneas were infected with A. fumigatus with or without pretreatment of recombinant mouse (rm) IL-36α and IL-36α neutralizing antibody (Ab). Primary macrophages were stimulated with 75% ethanol-killed A. fumigatus with or without pretreatment of rmIL-36α. The severity of the disease was documented by clinical score and photographs with a slit lamp. PCR, western blot, and immunostaining were used to determine the expression of IL-36α, IL-1β, IL-6, and TNF-α. Polymorphonuclear neutrophilic leukocyte infiltration was assessed by myeloperoxidase (MPO) assay and flow cytometry. Macrophage infiltration was tested by immunofluorescent staining and flow cytometry. RESULTS: IL-36α mRNA and protein were significantly elevated in human and mice corneas after infection. The rmIL-36α treatment of C57BL/6 mice increased clinical score, MPO levels, macrophage infiltration, and expression of the proinflammatory cytokines IL-1β, IL-6, and TNF-α compared with the infected controls, which showed a decrease due to IL-36α Ab treatment. In primary macrophages, IL-36α expression was also significantly increased by A. fumigatus. The rmIL-36α treatment upregulated IL-1β, IL-6, and phosphorylated nuclear factor (NF)-κB expression, which was significantly inhibited by rmIL-36Ra. CONCLUSIONS: IL-36α act as a proinflammatory cytokine in A. fumigatus keratitis by promoting the infiltration of neutrophils and macrophages and increasing the secretion of IL-1β, IL-6, and TNF-α, in addition to regulating expression of phosphorylated NF-κB.