PURPOSE: Acute Respiratory Distress Syndrome (ARDS) remains a critical health threat with limited pharmacological treatments. This study investigates the role of the N-methyladenosine (mA) demethylase AlkB homolog 5 (ALKBH5) in alveolar macrophages and its subsequent impact on inflammation and autophagy during ARDS pathogenesis. 6 6
METHODS: Primary mouse alveolar macrophages were stimulated with Lipopolysaccharide (LPS) and transfected with ALKBH5 knockdown or overexpression plasmids. Global m6A levels were assessed via Dot Blot, while mA modification of the target gene ULK1 was analyzed using MeRIP-qPCR. Macrophage polarization, migration, and autophagy (LC3-II/p62 flux) were evaluated in vitro. The therapeutic potential of ALKBH5 downregulation was validated in an LPS-induced murine ARDS model through lung histology, cytokine analysis (ELISA), and microvascular permeability assessments. 6
RESULTS: ALKBH5 was found to be a critical regulator of mA methylation in alveolar macrophages. LPS stimulation decreased mA modification of ULK1 mRNA, a process reversed by ALKBH5 downregulation. ALKBH5 knockdown significantly suppressed LPS-induced autophagy by reducing autophagosome formation and inhibited M1 pro-inflammatory polarization. In vivo, ALKBH5 downregulation significantly mitigated lung tissue damage, reduced pulmonary edema, and lowered levels of pro-inflammatory cytokines, including IL-1β, TNF-α, and IL-17. 6 6
CONCLUSIONS: Our findings demonstrate that ALKBH5-driven mA demethylation of ULK1 exacerbates ARDS by promoting macrophage autophagy and pro-inflammatory responses. These results suggest that targeting ALKBH5 may disrupt pathogenic m6A demethylation, offering a novel therapeutic strategy for mitigating ARDS progression. 6
