ATF4 Involvement in TLR4 and LOX-1-Induced Host Inflammatory Response to Aspergillus fumigatus Keratitis

PURPOSE: Activating transcription factor 4 (ATF4) is induced by various stressors. Here, we investigated the expression of ATF4 in the host inflammatory response to Aspergillus fumigatus (A. fumigatus) keratitis. METHODS: A. fumigatus keratitis mouse models developed by intrastromal injection as well as corneal epithelium scratching were examined daily with a slit lamp microscope for corneal opacification and ulceration. Subsequent in vitro experimentation was carried out in human corneal epithelial cells (HCECs) as well as THP-1 macrophages infected with A. fumigatus. Inhibitors, including CLI-095, Poly (I), SCH772984, and SP600125, were used to assess the role of proteins like toll-like receptor 4 (TLR4), lectin-type oxidized LDL receptor 1 (LOX-1), extracellular signal-regulated kinases (ERK1/2), and c-Jun N-terminal kinase (JNK) in ATF4 expression as a response to A. fumigatus infection. This assessment was made in both mouse models and HCECs using western blot. RESULTS: Compared to the controls, ATF4 was increased in corneas from two kinds of A. fumigatus keratitis models at 3 days after infection. ATF4 expression was upregulated with A. fumigatus conidia both in HCECs and THP-1 macrophages 16 hours after stimulation. Furthermore, ATF4 expression in response to A. fumigatus infection was shown to be dependent on TLR4 and LOX-1 expression, and ERK1/2 and JNK contributed to the expression of ATF4 in response to A. fumigatus. CONCLUSION: Our results clearly indicate that ATF4 was involved in the host antifungal immune response to A. fumigatus keratitis; expression was found to be dependent on TLR4, LOX-1 expression, and MAPKs pathway.