Pharmacological activation of TRPML1 enhances autophagy regulating hypertonicity and TGF-β-induced EMT in proximal tubular epithelial cells

Proximal tubular epithelial cells (PTECs) are central to maintaining kidney homeostasis. Under pathological conditions, such as ischemia or inflammation, PTECs promote profibrotic signals, including transforming growth factor (TGF)-β, and undergo epithelial-mesenchymal transition (EMT). EMT is characterized by decreased epithelial markers (e.g., E-cadherin) and increased mesenchymal markers (e.g., α-smooth muscle actin [α-SMA]), which promote myofibroblast activation and fibrosis progression. We previously demonstrated that hyperosmotic stress, characterized by elevated extracellular solute concentrations, induces EMT in PTECs. However, we observed that hyperosmotic stress simultaneously activates autophagy, a cellular process that has antagonistic effects on EMT, primarily mediated by transient receptor potential mucolipin 1 (TRPML1). However, the interplay between hyperosmotic stress-induced EMT and autophagy remains unclear. This study examined whether enhancing autophagy via TRPML1 activation could modulate EMT under hyperosmotic stress. Using the TRPML1 agonist ML-SA1, we observed a significantly increased autophagic flux, indicated by elevated LC3-II levels, without cytotoxic effects. Under hyperosmotic conditions, ML-SA1 further amplified autophagic flux in PTECs compared to hyperosmotic stress alone. Notably, this enhanced autophagy suppressed EMT by maintaining E-cadherin expression and reducing α-SMA levels. Furthermore, the ML-SA1-mediated autophagy enhancement attenuated EMT and profibrotic factor production in TGF-β-treated cells, suggesting a broader protective role beyond hyperosmotic stress. These findings reveal a novel interaction between hyperosmotic stress-induced autophagy and EMT, emphasizing TRPML1 activation's therapeutic potential to mitigate PTEC injury and fibrosis progression.