Myosin light chain kinase 2 and Myomesin 2 are related to the stiffness of vocal fold lamina propria in aging rats and humans

Vocal fold fibroblasts play an important role in the production of the extracellular matrix in the vocal folds. Myofibroblasts increase in the aging vocal fold lamina propria with increased alpha-smooth muscle actin. In addition to alpha-smooth muscle actin, various sarcomeric genes are expressed in myofibroblasts. However, there have been no studies on sarcomeric genes with myofibroblast differentiation in aging vocal folds. The purpose of this study was to analyze the changes and functions of sarcomeric genes related to myofibroblast differentiation in aging vocal folds using next-generation sequencing (NGS). Young (6-month-old, 22 rats) and old (22-month-old, 22 rats) male Sprague–Dawley rats were used for this study. NGS was performed on the harvested lamina propria of the vocal folds in each group. NGS data were analyzed using functional annotation, gene ontology, network pathways, and network analysis methods. After identifying the increased expression of sarcomeric genes in aging vocal folds, we evaluated the expression of sarcomeric genes in the normal human vocal fold lamina propria removed after surgery for various vocal fold lesions. The functions of sarcomeric genes in fibroblast senescence, proliferation, differentiation, contraction, and stiffness were investigated. Among the four sarcomeric genes identified through network cluster analysis of the NGS results, the expression levels of myosin light chain kinase 2 (Mylk2) and myomesin 2 (Myom2) were significantly higher in the lamina propria of old rats than in young rats. The increase in Mylk2 and Myom2 expression was associated with cellular senescence but not with the proliferative ability of fibroblasts. However, the expression of Mylk2 and Myom2 increased with myofibroblast differentiation. Inhibition of Mylk2 and Myom2 affects cellular contraction, leading to reduced stiffness. Our results suggest that Mylk2 and Myom2 are novel biomarkers of vocal fold myofibroblasts and are involved in the regulation of vocal fold stiffness in aging rats and humans.