The Role of Aerobic Glycolysis in TGF-β-induced Epithelial-to-mesenchymal Transition in Lung Epithelial Cells

Chronic respiratory diseases (CRDs) are a growing global health concern, characterized by persistent inflammation, airway remodeling, and progressive airflow limitation. Elevated levels of the pro-inflammatory cytokine TGF-β promote epithelial-to-mesenchymal transition (EMT), a central process in CRD pathogenesis. However, its impact on cellular energy metabolism, particularly glycolysis, in lung epithelial cells remains unclear. We investigated the role of glycolysis in TGF-β-induced EMT using A549 and BEAS-2B lung epithelial cells. Immunoblotting, RT-qPCR, migration assays, ChIP, Seahorse analysis, lactate assays, glucose uptake, and siRNA transfection were used to dissect the link between glycolysis and EMT. TGF-β-induced EMT was reflected by enhanced motility and changes in epithelial and mesenchymal markers, alongside Smad3 phosphorylation and recruitment to the Snail promoter. It also upregulated glycolytic enzymes and lactate production. Inhibition of glycolysis with 2-deoxy-D-glucose (2-DG) suppressed TGF-β-induced Smad3 phosphorylation and EMT. Smad3 siRNA decreased TGF-β-driven glycolytic enzyme expression, including 6-phosphofructo-2-kinase/fructose 2,6-bisphosphatase 3 (PFKFB3). Moreover, TGF-β-induced glycolytic lactate further promoted Smad3 phosphorylation, suggesting a positive feedback loop. In vivo, decreased E-cadherin and increased Snail expression correlated with elevated GLUT1 and PFKFB3 in the lung epithelium of house dust mite (HDM)-induced allergic asthma and bleomycin (BLM)-induced idiopathic pulmonary fibrosis (IPF) models. Our findings reveal a vicious TGF-β-lactate-TGF-β cycle in which TGF-β enhances glycolysis, elevates lactate, and amplifies Smad3 signaling to promote EMT. Targeting glycolytic reprogramming may provide novel therapeutic strategies for TGF-β-associated CRDs such as asthma and pulmonary fibrosis.