MiR-223-3p Regulates Autophagy and Inflammation by Targeting ATG16L1 in Fusarium solani–Induced Keratitis

Abstract

Purpose Increasing evidence suggested that microRNAs (miRs) are implicated in the regulation of the inflammatory response and autophagy in multiple diseases. The present study aimed to explore the effect of miR-223-3p on inflammation and autophagy in fungal keratitis (FK). Methods An FK mouse model was established, and primary corneal stromal cells were isolated by inoculation with Fusarium solani . The expression of miR-223-3p was determined by quantitative RT-PCR. Subsequently, the target gene of miR-223-3p was identified by a dual-luciferase reporter assay. The levels of miR-223-3p were altered by transfecting miR agomir/antagomir to evaluate its effects. Slit-lamp biomicroscopy and hematoxylin and eosin staining were employed to detect corneal damage. The levels of autophagy were assessed by immunofluorescence, Western blotting, mRFP-GFP-LC3 fluorescence microscopy, and electron microscopy. In addition, inflammation was demonstrated by determining the proinflammatory mediators IL-1β and TNF-ɑ. Results Our data suggested that miR-223-3p was increased and that autophagic flux was impaired in mouse FK. Then, we confirmed that autophagy-related gene 16L1 (ATG16L1) was a potential target of miR-223-3p and that this miR negatively regulated the expression of ATG16L1. The inhibition of miR-223-3p attenuated inflammation in FK, reduced P62 expression, and increased the ratio of LC3-II/LC3-I, whereas the overexpression of miR-223-3p displayed the opposite results. Conclusions Taken together, miR-223-3p might regulate autophagy via targeting ATG16L1 in experimental F. solani keratitis and is associated with the inflammatory response. MiR-223-3p might be a potential therapeutic target for FK.

Inhibition of the m6A Methyltransferase METTL3 Attenuates the Inflammatory Response inFusarium solani-Induced Keratitis via the NF-κB Signaling Pathway

“…After activation of autophagy, corneal stromal cells are more activated and the clearance of fungus is accelerated, which is helpful for the repair of keratitis. 17 , 24 , 45 METTL3-mediated m6A methylation can negatively regulate autophagy, and downregulation of METTL3 can promote the expression of autophagy-related genes and enhance autophagy. 46 , 47 Therefore, we speculate that downregulation of METTL3 may promote fungal clearance by increasing autophagy flux, and further experimental validation is needed.…”

“…After 2 weeks, the FK models were established as described in our previous study. 17 In brief, intraperitoneal injection with 0.6% pentobarbital sodium was applied for general anesthesia, and 0.5% proparacaine hydrochloride was used for topical anesthesia in mice. Subsequently, the corneal epithelium (2.5 mm in diameter) was removed with an electric scraper.…”

Tear Film MicroRNAs as Potential Biomarkers: A Review

“…Increased expression of tear miR-223-3p in HEK patients has also been reported in the corneal stromal cells of fungal keratitis murine models [ 57 ], associating it with keratitis pathophysiology. Studies have shown upregulation of miR-155 and miR-132 in association with HSV keratitis murine models, specifically in angiogenesis of the cornea [ 70 , 71 ].…”

“…Furthermore, the miR-16, miR-142, and miR-223 families are crucial during hematopoietic lineage differentiation and are known to participate in the development and differentiation of immune cells [ 19 , 56 ]. Upregulation of miR-223-3p leads to increased detection of pro-inflammatory cytokines TNF-α and IL-1β in the corneas of mouse models of fungal keratitis [ 57 ]. Cortes-Troncoso et al found that miR-142-3p, which was expressed in the salivary glands of SS patients but not healthy controls, is transported from activated T-cells into glandular cells where it is capable of restricting cAMP production and altering intracellular Ca 2+ signaling [ 58 ].…”

Exosomes and autophagy in ocular surface and retinal diseases: new insights into pathophysiology and treatment

“…As one of infectious corneal diseases, fungal keratitis may result in blindness [ 69 ]. In Fusarium solani fungal keratitis (FK) mouse models, miR-223-3p might regulate autophagy via targeting ATG16L1 and was associated with the inflammatory response, which might be a potential therapeutic target [ 70 ]. Another study has demonstrated that the expression of miR-665-3p in the cornea was upregulated and autophagy flux was impaired when infected with Fusarium solani, thus miR-665-3p inhibitors might activate autophagic pathways and inhibit inflammation to cure FK [ 71 ].…”