Microarray Analysis Gene Expression Profiles in Laryngeal Muscle After Recurrent Laryngeal Nerve Injury.

OBJECTIVES: The pathophysiology of recurrent laryngeal nerve (RLN) transection injury is rare in that it is characteristically followed by a high degree of spontaneous reinnervation, with reinnervation of the laryngeal adductor complex (AC) preceding that of the abducting posterior cricoarytenoid (PCA) muscle. Here, we aim to elucidate the differentially expressed myogenic factors following RLN injury that may be at least partially responsible for the spontaneous reinnervation. METHODS: F344 male rats underwent RLN injury (n = 12) or sham surgery (n = 12). One week after RLN injury, larynges were harvested following euthanasia. The mRNA was extracted from PCA and AC muscles bilaterally, and microarray analysis was performed using a full rat genome array. RESULTS: Microarray analysis of denervated AC and PCA muscles demonstrated dramatic differences in gene expression profiles, with 205 individual probes that were differentially expressed between the denervated AC and PCA muscles and only 14 genes with similar expression patterns. CONCLUSIONS: The differential expression patterns of the AC and PCA suggest different mechanisms of reinnervation. The PCA showed the gene patterns of Wallerian degeneration, while the AC expressed the gene patterns of reinnervation by adjacent axonal sprouting. This finding may reveal important therapeutic targets applicable to RLN and other peripheral nerve injuries.

Stem cell-derived tissue-engineered constructs for hemilaryngeal reconstruction.

OBJECTIVES: As an initial step toward our goal of developing a completely tissue-engineered larynx, the aim of this study was to describe and compare three strategies of creating tissue-engineered muscle-polymer constructs for hemilaryngeal reconstruction. METHODS: Cartilage-mimicking polymer was developed from electrospun poly(D,L-lactide-co-ε-caprolactone) (PCL). Primary muscle progenitor cell cultures were derived from syngeneic F344 rat skeletal muscle biopsies. Twenty F344 rats underwent resection of the outer hemilaryngeal cartilage with the underlying laryngeal adductor muscle. The defects were repaired with muscle stem cell-derived muscle-PCL constructs (5 animals), myotube-derived muscle-PCL constructs (5 animals), motor end plate-expressing muscle-PCL constructs (5 animals), or PCL alone (controls; 5 animals). The outcome measures at 1 month included animal survival, muscle thickness, and innervation status as determined by electromyography and immunohistochemistry. RESULTS: All of the animals survived the 1-month implant period and had appropriate weight gain. The group that received motor end plate-expressing muscle-PCL constructs demonstrated the greatest muscle thickness and the strongest innervation, according to electromyographic activity and the percentage of motor end plates that had nerve contact. CONCLUSIONS: Although all of the tissue-engineered constructs provided effective reconstruction, those that expressed motor end plates before implantation yielded muscle that was more strongly innervated and viable. This finding suggests that this novel approach may be useful in the development of a tissue-engineered laryngeal replacement.

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.

Safety practices for in-office laryngology procedures during clinical reintroduction amidst COVID-19.

Objective: Describe safety practices for performing in-office laryngology procedures during clinical re-introduction amidst the coronavirus disease 2019 (COVID-19) pandemic. Methods: An anonymous survey in Qualtrics was created to evaluate demographics, preprocedure testing, practice settings, anesthesia, and personal protective equipment (PPE) use for five procedure categories (non-mucosal-traversing injections, mucosal-traversing injections, endoscopy without suction, endoscopy with suction/mucosal intervention via working channel, and laser via working channel). The survey was emailed to the Fall Voice Community on Doc Matter and to members of the American Broncho-Esophagological Association (ABEA) from May to June 2020. Results: Eighty-two respondents were analyzed (response rate: 10%). Respondents represented diverse locations, including international. Most reported academic (71%) or private practices (16%), laryngology fellowship training (76%), and a significant practice devotion to laryngology and broncho-esophagology. During the early re-introduction, most continued to perform all procedure categories. The office was preferred to the OR setting for most, though 36% preferred the OR for laser procedures. There was a preference for preprocedural SARS-Cov2 testing for procedures involving a working channel (>67%), and these procedures had the highest proportion of respondents discontinuing the procedure due to COVID-19. Various types of topical anesthesia were reported, including nebulizer treatments. The most common forms of personal protective equipment utilized were gloves (>95%) and N95 masks (>67%). Powered-air purifying respirators and general surgical masks were used infrequently. Conclusions: During the early re-introduction, respondents reported generally continuing to perform office laryngology procedures, while greater mucosal manipulation affected decisions to stop procedures due to COVID-19, perform preprocedural SARS-Cov2 testing, and alter topical anesthesia. Gloves and N95 masks were the predominate PPE. Level of Evidence: N/A.

Influence of anxiety and depression on the predictive value of the Reflux Symptom Index.

OBJECTIVES: Although the Reflux Symptom Index (RSI) is a validated laryngopharyngeal reflux (LPR) outcomes tool, its predictive value for LPR is controversial. Because psychiatric problems may lead to exaggerated patient-perceived symptoms and RSI values,the aim of this study was to determine whether the positive predictive value of the RSI for pH probe-documented LPR is influenced by anxiety and depression. METHODS: We reviewed the charts of all patients who underwent pH probe testing for LPR between January 2006 and July 2008 at our institution. The RSI, Reflux Finding Score (RFS), medical history, and pH probe findings were recorded. Patients with anxiety or depression were included in the psychiatric disorder (+PSY) group, and those without anxiety or depression comprised the non-psychiatric disorder (-PSY) group. Predictive values of the RSI for pH probe-documented LPR were determined for each group. RESULTS: We included 51 patients: 30 patients (59%) in the -PSY group and 21 patients (41%) in the +PSY group. The mean RSI of the +PSY group was higher than that of the -PSY group (p < 0.05), but the +PSY patients actually had a lower incidence of abnormal probe studies (p < 0.02). The positive predictive value of an elevated RSI for an abnormal pH probe study was poor in the +PSY patients (p = 0.495), but strong in the -PSY group (p = 0.004). CONCLUSIONS: The presence of anxiety and depression impairs the predictive value of the RSI for LPR. This finding potentially explains some of the controversy over the diagnostic utility of the RSI.

Local neurotoxins for prevention of laryngeal synkinesis after recurrent laryngeal nerve injury.

OBJECTIVES: Persistent vocal fold motion impairment after recurrent laryngeal nerve (RLN) injury is not characteristically due to absent reinnervation, but often results from spontaneous aberrant reinnervation (synkinesis). We administered local neurotoxins to selected laryngeal muscles after RLN injury to determine whether aberrant reinnervation could be selectively inhibited. METHODS: Unilateral RLN transection was performed in 24 male rats. Three weeks later, the denervated laryngeal adductor complex was injected with phenol, high- or low-dose vincristine sulfate (VNC), or saline solution. One month later, rat larynges were evaluated via videolaryngoscopy and laryngeal electromyography (LEMG). Larynges from euthanized animals were analyzed via immunofluorescent staining for the presence of reinnervation. RESULTS: One animal that received phenol and 3 animals that received high-dose VNC died of toxicity-related complications. In the surviving neurotoxin-treated animals, videolaryngoscopy showed increased lateralization of the immobile vocal fold. Only 1 phenol-injected rat had adductor complex motor recruitment (score of 3+) with LEMG. The other neurotoxin-treated animals demonstrated an absence of adductor complex reinnervation, with only insertional activity and fibrillations (no motor units/recruitment). Spontaneous ipsilateral abductor reinnervation was not affected by the adductor injections. CONCLUSIONS: Low-dose VNC injections appear to be relatively safe and effective in selectively inhibiting spontaneous aberrant reinnervation after RLN injury in an animal model.

Current Indications for Transnasal Esophagoscopy: An American Broncho-Esophagological Association Survey.

INTRODUCTION:: The aim of this study was to evaluate the current indications for and clinical factors influencing the use of transnasal esophagoscopy (TNE). METHODS:: An online survey was sent to American Broncho-Esophagological Association members, including questions on demographics, indications, and factors influencing the use of TNE. RESULTS:: Sixty of 251 members (24%) completed the survey. Ninety-three percent of respondents reported academic practices, while 98% practice in medium to large urban settings. Thirty-five (58%) completed laryngology fellowships. Mean monthly TNE procedure count was 7.15 (range, <1-35). The most common indications were dysphagia (82%), biopsy (50%), and laryngopharyngeal reflux (47%). Chronic cough, head and neck cancer screening, gastroesophageal reflux (GER), and tracheoesophageal puncture were also commonly reported indications (44% each). For laryngopharyngeal reflux and GER, most respondents perform TNE for recalcitrant disease following a medical trial of at least 3 months. Long-standing GER symptoms, documentation of GER on pH and impedance testing, and abnormal findings on previous esophagoscopy lead to greater TNE use. Specific dysphagia indications included abnormal esophagographic findings (70%), history or examination localizing to the esophagus (60%), solid dysphagia only (53%), and solid and/or liquid dysphagia (40%). The primary sites most likely to prompt TNE use for head and neck cancer surveillance were the esophagus (92.3%) and hypopharynx (84.6%). Balloon dilation was the most common indication for which respondents do not currently perform TNE but would like to (n = 8). CONCLUSIONS:: TNE indications have not been well established. According to respondents from the American Broncho-Esophagological Association, TNE is most commonly used for dysphagia and laryngopharyngeal reflux and slightly less so for GER and head and neck cancer screening and surveillance. Several clinical indicators were identified that influence the decision to perform TNE.

Injection of autologous muscle stem cells (myoblasts) for the treatment of vocal fold paralysis: a pilot study.

OBJECTIVE: Autologous muscle stem cell (myoblast) therapy may be an ideal treatment for vocal fold paralysis because of its technical ease (administered by injection), its potential to restore muscular defects and dynamic function, and its autologous origin. The goal of this project was to determine whether autologous myoblast injection into the thyroarytenoid (TA) muscle after recurrent laryngeal nerve (RLN) injury could attenuate TA muscle atrophy and enhance spontaneous reinnervation. STUDY DESIGN: This was an animal experiment. METHODS: Unilateral RLN transection and sternocleidomastoid muscle (approximately 1 g) biopsies were performed in 16 male Wistar rats. Biopsies were used to create myoblast cultures for each animal. One month later, 10(6) autologous myoblasts labeled with fluorescent cell membrane marker (PKH26) were injected into the denervated TA of each study animal, with saline injected into controls. Animals were euthanized at 2 weeks and 2 months after myoblast injection. Outcomes included myoblast survival, TA fiber diameter and volume, and reinnervation status (motor endplate to nerve contact staining). RESULTS: All denervated TA study specimens demonstrated viable myoblasts under fluorescent microscopy, with the myoblasts demonstrating fusion with the TA myofibers at 2 months. The myoblast-treated group had greater mean TA fiber diameter than denervated TA controls at 2 months (25.1 vs. 21.1 microm; P = .04) but not at 2 weeks (25.7 microm vs. 23.5 microm; P = .06). Mean TA volumes were greater in the myoblast-treated groups at both time points. Two of the animals in the myoblast-treated group demonstrated adductor motion at 2 months, whereas none of the 2 week study animals or controls recovered adduction. Reinnervation was not significantly different between the myoblast-treated groups and the denervated controls. CONCLUSIONS: Autologous myoblast therapy may be a future treatment for vocal fold paralysis, with current findings demonstrating myoblast survival with attenuation of TA muscle atrophy.

Cricopharyngeal electromyography: patterns of injury based on etiology.

OBJECTIVE: Although cricopharyngeal electromyography (CP-EMG) is recognized as a diagnostic tool for dysphagia assessment, few reports in the literature characterize CP-EMG abnormalities in relation to clinical presentation. The aim of this study was to review a large series of CP-EMG studies, and compare the CP-EMG results with the patients' diagnoses. METHODS: A retrospective review of all CP-EMG performed at our institution over a 10-year period was executed. CP-EMG findings were then compared with the patients' clinical history, focusing on potential etiologies of neurogenic injury. RESULTS: Seventy CP-EMGs were reviewed, with 47 (67%) demonstrating neural injury. Of those cases with neural injury on EMG, 29 (60%) had known vagal injuries, 13 (28%) had idiopathic nerve palsies, and 5 (11%) had central etiologies, such as stroke. Each of these three neurogenic subgroups revealed a distinct pattern of EMG abnormalities. CONCLUSION: This study suggests there is an association between patterns of CP-EMG abnormalities and underlying etiology based on clinical history.

Differentiation of adductor-type spasmodic dysphonia from muscle tension dysphonia by spectral analysis.

OBJECTIVES: To determine the utility of spectral analysis in the differentiation of adductor-type spasmodic dysphonia (AdSD) from muscle tension dysphonia (MTD). STUDY DESIGN: Prospective blinded study. METHODS: Forty-seven samples of AdSD-connected speech spectrograms from 27 subjects and 17 samples of MTD-connected speech spectrograms from 15 subjects were selected from clinical charts and de-identified. These spectrograms were reviewed independently and blindly by two speech language pathologists experienced in spectrography. The speech language pathologists designated the spectrogram as consistent with AdSD and MTD, and these designations were compared with actual clinical diagnoses. RESULTS: The ability to differentiate AdSD from MTD with spectral analysis was 94% for rater #1 and 98% for rater #2. No MTD subjects were incorrectly diagnosed as having SD (100% specificity). CONCLUSIONS: This study suggests that experienced speech language pathologists can distinguish AdSD from MTD with a high degree of sensitivity and specificity based on spectral analysis. Spectral analysis is especially useful in cases where perceptual analysis and clinical evaluation alone are insufficient.