Purpose: To investigate the contribution and mechanism of miRNAs and autophagy in diabetic peripheral neuropathy. Methods: In this study, we used streptozotocin (STZ)-induced type I diabetes C57 mice as animal models, and we detected the expression of miR-34c and autophagic intensity in trigeminal ganglion (TG) tissue. The bioinformatics software was used to predict and analyze the potential targets of miR-34c. Primary trigeminal ganglion neurons were cultured in vitro to investigate the effect of miR-34c on axon growth and survival of TG cells. A corneal epithelial damage-healing model was established on the diabetic mice, then miR-34c antagomir was injected subconjunctivally. The condition of corneal epithelial healing was observed through sodium fluorescein staining, and the peripheral nerve degeneration of the cornea was evaluated by β-tublin corneal nerve staining. Results: The expression of miR-34c was significantly increased in TG tissue of type I diabetic mice by real-time PCR. Western blot showed that autophagy-related proteins Atg4B and LC3-II were significantly down-regulated in diabetes TG compared with normal control. Trigeminal neuron immunofluorescence showed that the length of the trigeminal ganglion cell synapses was significantly increased after miR-34c antagomir treatment compared with normal cultures. Subconjunctival injection of miR-34c antagomir can significantly promote corneal epithelium healing of diabetic mice and appreciably promote the regeneration of corneal nerve. At the same time, it can significantly increase the expression of autophagy in TG tissue of type I diabetic mice. Conclusions: In this study , miR-34c was found to affect the growth of trigeminal sensory neurons and the repair of diabetic corneal nerve endings by acting directly on Atg4B.

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.

Purpose: Our previous investigations revealed a significant role of methyltransferase-like 3 (METTL3)-mediated N6-methyladenosine (m⁶A) modification in the development of corneal inflammation in Fusarium infection, but the exact mechanism is unknown. Therefore, this research aimed to explore how METTL3 affects the inflammatory process of fungal keratitis (FK) in mice. Methods: We established in vitro and in vivo models by inoculating mice and primary corneal stromal cells with F. solani. METTL3 expression was confirmed by real-time quantitative polymerase chain reaction, immunofluorescence, and western blotting. After that, siRNAMETTL3 and AAV-sh-METTL3 were transfected into cells and mice to explore the role of METTL3 in the PI3K/AKT signaling pathway and inflammation. PI3K, p-PI3K, AKT, and p-AKT expression was analyzed by western blotting. Viability of corneal stromal cells was measured using a Cell Counting Kit-8 (CCK-8). Additionally, we detected interleukin (IL)-6, IL-1β, and tumor necrosis factor alpha (TNF-α) levels in corneal tissues and analyzed the role of METTL3 in inflammation in FK using slit-lamp biomicroscopy and hematoxylin and eosin staining. Results: Here, our results show that METTL3 increased in mouse FK, and the expression of p-PI3K and p-AKT decreased when METTL3 was downregulated. We also found that knockdown of METTL3 expression attenuated the inflammatory response and decreased TNF-α, IL-1β, and IL-6 expression in corneal-infected mice. Furthermore, inhibition of the PI3K/AKT pathway attenuated the inflammatory response of FK, decreased the expression of the above inflammatory factors, and enhanced the viability of corneal stromal cells. Conclusions: Based on the study results, METTL3 downregulation attenuates Fusarium-induced corneal inflammation via the PI3K/AKT signaling pathway.