Proteomic Stratification Across Human Male Vocal Fold Tissue Layers

To compare relative proteomic abundance across the human true vocal fold cover (epithelium and superficial lamina propria; TVFC), vocal ligament (intermediate and deep lamina propria; VL), and thyroarytenoid muscle (TA), and to characterize shared and distinct molecular pathways.

Cross-sectional exploratory proteomic study using tandem mass tag-based quantitative mass spectrometry on tissue from five human male cadaveric larynges (mean age 56.6 years) obtained from a national tissue bank. Differential protein abundance was assessed using linear mixed modeling, and Gene Ontology (GO) enrichment analysis characterized functional specialization across layers.

Of 895 quantified proteins, 554 were detected in all three layers, with 181 jointly detected in the TVFC and VL, and 160 detected only in the TA. Differential expression analysis revealed minimal divergence between TVFC and VL (all FDR > 0.10) but robust separation of TA from both the TVFC and VL (FDR < 0.05). GO enrichment demonstrated that TVFC was enriched for inflammatory and immune processes, VL occupied an intermediate position sharing features with both TVFC and TA, and TA demonstrated contractile specialization.

This exploratory study provides an initial descriptive comparison of protein expression across human vocal fold layers, with VL serving as a potential transitional zone between the immunomodulatory cover and contractile muscle. These findings confirm the well-established histological, cellular, and functional differences among these layers, and provide preliminary data to inform future studies of laryngeal biology, pathology, and intervention.