MicroRNA miR-212-5p Regulates the MEK/ERK Signaling Pathway by Targeting A-Raf proto-oncogene serine/threonine-protein kinase (ARAF) to Regulate Cowshed PM2.5-Induced NR8383 Apoptosis.

ABSTRACT

Objective: To investigate the role of miR-212-5p-targeted ARAF during the apoptosis of rat alveolar macrophages induced by cowshed PM2.5. Methods: miRNA and related target genes and pathways were predicted using the KEGG, TargetScan, and other prediction websites. NR8383 macrophages were treated with cowshed PM2.5 to establish an in vitro lung injury model in rats; meanwhile, for the assessment of cell viability, apoptosis, intracellular calcium ions, and mitochondrial membrane potential in NR8383 cells, RT-qPCR was used to detect the expression of miR-212-5p and the target gene ARAF. Results: The bioinformatic analyses showed that miR-212-5p and ARAF were involved in PM2.5-associated cellular damage. Exposure to different concentrations (0 µg/mL, 60 µg/mL, 180 µg/mL, 300 µg/mL) with different durations (0 h, 12 h, 24 h, 48 h) of cowshed PM2.5 resulted in apoptosis, increased intracellular calcium ions, and decreased mitochondrial membrane potential. The miR-212-5p mimic group showed an up-regulation of Bax and cleaved Caspase 3 expression but decreased Bcl2 expression compared to the NC group, and overexpression of ARAF up-regulated the expression of p-MEK1/2 and p-ERK1/2 and simultaneously reversed the above phenomena. Conclusions: miR-212-5p targets ARAF to affect the cowshed PM2.5-induced apoptosis through the MEK/ERK signaling pathway, providing a potential target for relevant farming industry and pathology studies.