RNA-Sequencing Reveals the Modulation of the NLRP3 Inflammasome by miR-223-3p in Extracellular Vesicles in Bacterial Endophthalmitis.

Abstract

Purpose: Extracellular vesicles (EVs) are critical mediators of cell-cell communication via transfer of molecular cargo including miRNAs and regulate transcription in various physiological and pathological conditions. This study aimed to investigate the role of EV-derived-microRNAs (EV-miRNAs) in bacterial endophthalmitis, focusing on their regulatory impact on inflammation and host immune responses.

Methods: C57BL/6 mice were infected with Pseudomonas aeruginosa (P. aeruginosa) and Staphylococcus aureus (S. aureus) to induce endophthalmitis. EVs were isolated and characterized followed by miRNA profiling to identify differentially expressed miRNAs. The miRNet platform was used to elucidate potential interactions between exosomal miRNA and retinal mRNA, followed by in vivo and in vitro validation of key miRNAs and their target genes. Additionally, EVs were extracted from vitreous fluid samples of patients with endophthalmitis, and miR-223-3p and NLRP3 expressions were assessed by qPCR.

Results: Bacterial endophthalmitis led to pronounced neutrophil infiltration in the retina of mice. The miRNA profiling identified 651 differentially expressed miRNAs in P. aeruginosa and 29 in S. aureus, with 10 miRNAs shared between both infections. The miR-223-3p, miR-467a-3p, and miR-467d-3p emerged as major regulators of inflammatory pathways, targeting genes such as NLRP3, CXCL5, and IKKα. In patient vitreous samples, miR-223-3p was upregulated in culture-positive samples, correlating with reduced NLRP3 expression. The miRNAs, particularly miR-223-3p, play a critical role in modulating the immune response in bacterial endophthalmitis, largely through the regulation of the NLRP3 inflammasome.

Conclusions: The findings suggest that miR-223-3p could serve as biomarkers in culture-negative cases and therapeutic targets for managing inflammation in bacterial endophthalmitis, potentially guiding treatments aimed at preserving retinal integrity.