Transcriptome Analysis of Left Versus Right Intrinsic Laryngeal Muscles Associated with Innervation.

OBJECTIVES/HYPOTHESIS: Recurrent laryngeal nerve injury diagnosed as idiopathic or due to short-term surgery-related intubation exhibits a higher incidence of left-sided paralysis. While this is often attributed to nerve length, it is hypothesized there are asymmetric differences in the expression of genes related to neuromuscular function that may impact reinnervation and contribute to this laterality phenomenon. To test this hypothesis, this study analyzes the transcriptome profiles of the intrinsic laryngeal muscles (ILMs), comparing gene expression in the left versus right, with particular attention to genetic pathways associated with neuromuscular function. STUDY DESIGN: Laboratory experiment. METHODS: RNA was extracted from the left and right sides of the rat posterior cricoarytenoid (PCA), lateral thyroarytenoid (LTA), and medial thyroarytenoid (MTA), respectively. After high-throughput RNA-Sequencing, 88 samples were organized into 12 datasets according to their age (P15/adult), sex (male/female), and muscle type (PCA/LTA/MTA). A comprehensive bioinformatics analysis was conducted to compare the left-right ILMs across different conditions. RESULTS: A total of 774 differentially expressed genes were identified across the 12 experimental groups, revealing age, sex, and muscle-specific differences between the left versus right ILMs. Enrichment analysis of Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathways implicated several genes with a left-right laryngeal muscle asymmetry. These genes are associated with neuronal and muscular physiology, immune/inflammatory response, and hormone control. CONCLUSION: Bioinformatics analysis confirmed divergent transcriptome profiles between the left-right ILMs. This preliminary study identifies putative gene targets that will characterize ILM laterality. LEVEL OF EVIDENCE: N/A Laryngoscope, 134:3741-3753, 2024.

Neurotrophin expression and laryngeal muscle pathophysiology following recurrent laryngeal nerve transection.

Laryngeal palsy often occurs as a result of recurrent laryngeal or vagal nerve injury during oncological surgery of the head and neck, affecting quality of life and increasing economic burden. Reinnervation following recurrent laryngeal nerve (RLN) injury is difficult despite development of techniques, such as neural anastomosis, nerve grafting and creation of a laryngeal muscle pedicle. In the present study, due to the limited availability of human nerve tissue for research, a rat model was used to investigate neurotrophin expression and laryngeal muscle pathophysiology in RLN injury. Twenty-five male Sprague-Dawley rats underwent right RLN transection with the excision of a 5-mm segment. Vocal fold movements, vocalization, histology and immunostaining were evaluated at different time-points (3, 6, 10 and 16 weeks). Although vocalization was restored, movement of the vocal fold failed to return to normal levels following RLN injury. The expression of brain­derived neurotrophic factor and glial cell line-derived neurotrophic factor differed in the thyroarytenoid (TA) and posterior cricoarytenoid muscles. The number of axons did not increase to baseline levels over time. Furthermore, normal muscle function was unlikely with spontaneous reinnervation. During regeneration following RLN injury, differences in the expression levels of neurotrophic factors may have resulted in preferential reinnervation of the TA muscles. Data from the present study indicated that neurotrophic factors may be applied for restoring the function of the laryngeal nerve following recurrent injury.

Differences in laryngeal neurotrophic factor gene expression after recurrent laryngeal nerve and vagus nerve injuries.

OBJECTIVES: Recurrent laryngeal nerve (RLN) and vagus nerve (VN) injuries characteristically are followed by differing degrees of spontaneous reinnervation, yet laryngeal muscle neurotrophic factor (NF) expression profiles after RLN and VN injuries have not been well elucidated. This study's objective was to determine the relative changes in gene expression of 5 well-characterized NFs from laryngeal muscle after RLN or VN injuries in a time-dependent fashion, and demonstrate how these changes correspond with electromyography-assessed innervation status. METHODS: Thirty-six male rats underwent left RLN transection (12 rats), left VN transection (12 rats), or a sham procedure (12 rats). The primary outcomes included electromyographic assessment and laryngeal muscle NF expression quantification with reverse transcription polymerase chain reaction at 3 days and at 1 month. RESULTS: Electromyography at 3 days demonstrated electrical silence in the VN injury group, normal activity in the sham group, and nascent units with decreased recruitment in the RLN injury group. Reverse transcription polymerase chain reaction demonstrated that changes in NF gene expression from laryngeal muscles varied depending on the type of nerve injury (RLN or VN) and the specific laryngeal muscle (posterior cricoarytenoid or adductor) assessed. CONCLUSIONS: Laryngeal muscle NF expression profiles after cranial nerve X injury depend both upon the level of nerve injury and upon the muscles involved.