Stem Cell-Based Regeneration of Atrophied Vocal Folds after Recurrent Laryngeal Nerve Injury

This study evaluated the therapeutic potential of calcium hydroxylapatite (CaHA) and tonsil-derived mesenchymal stem cells (TMSCs) suspended in hyaluronic acid (HA) for regenerating atrophied vocal fold in a rabbit model of recurrent laryngeal nerve (RLN) injury. Vocal fold atrophy was induced by transecting the unilateral RLN in rabbits. Histological changes in muscle fiber (H&E), cartilage (Safranin O), and collagen deposition (Masson's trichrome), were assessed at 8, 12, 16, and 26 weeks post-injury to confirm the establishment of vocal fold atrophy model. CaHA or TMSC＋HA was injected into the atrophied vocal folds. After 8 weeks, laryngeal tissues were harvested to evaluate tissue morphology and the engraftment of transplanted cells using histological staining (H&E, Safranin O, Masson's trichrome, Alizarin Red S) and immunofluorescence analysis for human mitochondria. At 8 weeks after RLN transection, CaHA injection-maintained medialization of the atrophied vocal fold but failed to promote tissue regeneration and induced localized foreign body reaction characterized by multinucleated giant cells and granular calcium deposit. In contrast, TMSC＋HA injection markedly enhanced muscle fiber regeneration, reduced interstitial spaces between muscle fibers, and did not form fibrous capsule. Moreover, TMSC＋HA did not elicit immune reactivity or tissue calcification, suggesting superior biocompatibility and tissue integration. These findings indicate that while CaHA acts as a durable filler regenerating potential, TMSC＋HA offers distinct advantages by promoting muscle regeneration, and extracellular matrix remodeling. TMSC＋HA hydrogel presents a promising regenerative strategy for restoring atrophied vocal folds following RLN injury, demonstrating excellent biocompatibility and effective tissue integration.