Evaluation of Dying Vocal Fold Epithelial Cells by Ultrastructural Features and TUNEL Method

Abstract

Cell death is a regulated mechanism of eliminating cells to maintain tissue homeostasis. This study described 2 methodological procedures for evaluating cell death in the epithelium of immobilized, approximated and vibrated vocal folds from 12 New Zealand white breeder rabbits. The gold standard technique of transmission electron microscopy evaluated high-quality ultrastructural criteria of cell death and a common immunohistochemical marker, the terminal deoxynucleotidyl transferase dUTP nick end labeling method, to confirm cell death signaling. Results revealed that ultrastructural characteristics of apoptotic cell death, specifically condensed chromatin and apoptotic bodies, were observed after vocal fold vibration and approximation. Although episodes of necrosis were rare, few enlarged cell nuclei were present after vibration and approximation. The vocal fold expresses an immunohistochemical marker for apoptosis along the apical surface of the epithelium. This study provides a solid foundation for future investigations regarding the role of cell death in vocal fold health and disease.

Acute exposure to vibration is an apoptosis‐inducing stimulus in the vocal fold epithelium

“…Rather, vocal folds that underwent approximation demonstrated the smallest epithelial cell nuclei area. These findings are in agreement with previous studies reporting the expression of TUNEL-labeled cells, condensed chromatin, and apoptotic bodies in the vocal folds after vibration and approximation (11). The present study adds to the body of literature objective quantification of TUNEL staining and the measurement of epithelial cell nuclei area.…”

“…Apoptosis, the most commonly studied type of programmed cell death, is the physiological elimination of cells (8–10). Several studies have reported that apoptosis occurs in the vocal folds in response to biomechanical trauma (11–14). Following in vivo vocal fold vibration and vocal fold movement alone using a rabbit model, our laboratory observed morphological features of condensed chromatin and apoptotic bodies in vocal fold epithelial cells (11).…”

“…Several studies have reported that apoptosis occurs in the vocal folds in response to biomechanical trauma (11–14). Following in vivo vocal fold vibration and vocal fold movement alone using a rabbit model, our laboratory observed morphological features of condensed chromatin and apoptotic bodies in vocal fold epithelial cells (11). Although rare, ultrastructural features of enlarged cell nuclei, which is suggestive of necrosis, were also present.…”

“…Although rare, ultrastructural features of enlarged cell nuclei, which is suggestive of necrosis, were also present. To confirm that cell death occurred, we found that the vocal fold epithelium expressed an immunohistochemical marker for apoptosis using terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining (11). These findings are in agreement with the previously reported effects of mechanical trauma on cell death in vocal fold fibroblasts (12–14).…”

Singer’s Nodules: Investigating the Etiopathogenetic Markers Progressing Their Pathogenesis and Clinical Manifestations

“…The most common of such epithelium-associated apoptotic processes are described in the context of laryngeal squamous cell carcinoma, leucoplakia, keratosis, epithelial hyperplasia, human papilloma virus (HPV) papilloma and few other benign lesions. Hence, this leads us to speculate that apoptotic cell death may at least partly contribute to the pathogenesis of diseases affecting the vocal fold epithelium [ 10 ]. Additionally, apoptosis has been described in the laryngeal epithelium in response to the physical stressors of abduction, adduction, and biochemical vibrations.…”