C-terminus of HSC70-Interacting Protein-Mediated Transforming Growth Factor-β 1 Protein Degradation Underlies the Anti-fibrotic Effects of Verbascoside in Pulmonary Fibrosis.

Background

Pulmonary fibrosis (PF) is a progressive and fatal interstitial lung disease with limited therapeutic options. Transforming growth factor-β1 (TGF-β1) is a master driver of fibrogenesis, but strategies to effectively degrade this protein remain underexplored. Verbascoside, a natural phenylethanoid glycoside, exhibits anti-inflammatory and anti-fibrotic activities. However, its molecular mechanism in PF is unclear.

Methods

A BLM-induced mouse model was used to evaluate VBS efficacy in vivo. Human lung fibroblasts (IMR-90) were treated with BLM or TGF-β1 to assess fibroblast activation, migration, and fibrotic marker expression. Mechanistically, we examined the C-terminus of HSC70-interacting protein (CHIP)-mediated ubiquitination and protein kinase A (PKA) signaling, using pharmacological inhibitors (MG132, H89) and CHIP knockdown.

Results

VBS significantly reduced BLM-induced pulmonary fibrosis in mice, as shown by decreased hydroxyproline, collagen deposition, and fibrotic markers. In vitro, VBS suppressed BLM- or TGF-β1-induced fibroblast migration and activation. Notably, VBS reduced TGF-β1 protein levels without altering its mRNA expression. Mechanistically, VBS increased intracellular cAMP levels and PKA activity, promoted the interaction between CHIP and TGF-β1, and enhanced TGF-β1 ubiquitination. Furthermore, we demonstrate that PKA directly phosphorylates CHIP. These effects were abolished by PKA inhibition (H89) or CHIP knockdown, confirming that VBS drives CHIP-mediated TGF-β1 degradation via PKA activation. VBS also exhibited a favorable safety profile in mice.

Conclusion

VBS activates the PKA/CHIP axis to promote TGF-β1 ubiquitination and degradation, thereby alleviating PF. These findings position VBS as a promising PF therapeutic and support targeting TGF-β1 stability.