Modulation of Pepsin-Mediated Inflammatory Responses in Vocal Cord Epithelial Cells by Amprenavir.

Abstract

Pepsin is a key driver of epithelial injury and inflammation in laryngopharyngeal reflux (LPR). Prior work in esophageal cells implicates hypoxia-inducible factor-2α (HIF-2α) in refluxate-mediated cytokine secretion. We investigated whether pepsin increases HIF-2α under neutral (pH 7) conditions in immortalized human true vocal cord (TVC) cells and whether amprenavir, a pepsin inhibitor, can reverse this response. TVCs were treated with 0.1-1 mg/mL pepsin and/or 10 μM amprenavir at pH 7 for 2, 4, 6, and 24 h. HIF-2α protein levels were evaluated by Western blot. Based on the most inflammatory condition, expression of proinflammatory genes was assessed with or without a HIF-2α transcriptional inhibitor. The 6-h 1 mg/mL pepsin treatment induced the highest HIF-2α levels and was selected for further analysis. Pepsin significantly increased HIF-2α expression (p < 0.01), and amprenavir reduced it numerically (p = 0.056). Pepsin significantly upregulated ICAM1, IL1B, and IL8 (p < 0.05), while amprenavir significantly reduced ICAM1, IL8, and TNF (p < 0.05). Inhibition of HIF-2α selectively reduced IL1B expression (p < 0.05). Pepsin exposure increased HIF-2α and upregulated key inflammatory genes. While amprenavir significantly reduced cytokine expression, HIF-2α inhibition alone had only a modest effect, suggesting additional pathways contribute to pepsin-induced inflammation. Amprenavir's ability to attenuate this response supports its therapeutic potential in LPR. N/A.