Faculty Bibliography

OBJECTIVE/UNASSIGNED:Oversimplified clinical dogma suggests that laryngeal diseases fall into two broad, mutually exclusive diagnostic categories-mucosal injury or neuromuscular/functional disorders. Extensive investigation in the lower airway as well as other organ systems suggest complex interactions between tissue types underlying both tissue health and pathological states. To date, no such relationship has been described in the vocal folds, likely the most bioactive organ in the body. We hypothesize interactions between the vocal fold muscle and mucosa likely contribute to aberrant phonatory physiology and warrant further investigation to ultimately develop novel therapeutic strategies. METHODS/UNASSIGNED:Primary culture of myoblasts from rat thyroarytenoid muscle and fibroblasts from the vocal fold mucosa were established. Co-culture and conditioned media experiments were performed to established bidirectional interactions between cell types. Transforming Growth Factor (TGF)-β was employed to stimulate a fibrotic phenotype in culture. In addition to quantitative PCR, standard migration and proliferation assays were performed as well as immunocytochemistry. RESULTS/UNASSIGNED:Bidirectional cell-cell interactions were observed. Without TGF-β stimulation, myoblast conditioned media inhibited fibroblast migration, but enhanced proliferation. Conversely, fibroblast conditioned media increased both myoblast proliferation and migration. Myoblast conditioned media decreased TGF-β-mediated gene expression and of particular interest, ACTA2 mRNA expression. In both co-culture and in response to fibroblast conditioned media, myosin heavy chain (Myh2) mRNA expression decreased in myoblasts. CONCLUSIONS/UNASSIGNED:These data are the first to describe interactions between cell types within the vocal fold. The implications for these interactions in vivo warrant further investigation to develop and refine optimal treatment strategies.

The application of exercise science training knowledge has been of growing interest to voice professionals. This tutorial, derived from the authors' invited presentations from the "Exercise and the Voice" Special Session at the 2018 Voice Foundation Symposium, proposes a foundational theoretical structure based in exercise science, clarifies the wide range of variables that may influence voice training, and summarizes our present understanding of voice physiology from the perspective of muscle training. The body of literature on voice exercise was then analyzed from the perspective of this framework, identifying what we currently know and what we still have yet to learn.

OBJECTIVES/HYPOTHESIS/OBJECTIVE:Menopause adversely affecs power and endurance of the limb muscles. However, despite clinical observations that menopause corresponds to negative changes of the voice, the direct effects of estrogen deprivation on the thyroarytenoid muscles are unknown. The purpose of this study was to investigate the effects of estrogen deprivation via ovariectomy on three neuromuscular parameters of the thyroarytenoid muscles using a rat model. STUDY DESIGN/METHODS:Animal model. METHODS:Cryosections of vocal folds of 20 (10 control and 10 ovariectomized) female rats were stained to label neuromuscular junctions, fiber size, or parvalbumin levels using immunohistochemical techniques and compared between experimental groups. RESULTS:The neuromuscular junctions, thyroarytenoid fiber sizes, and parvalbumin levels of the vocal folds were similar between experimental groups. CONCLUSIONS:The loss of estrogen did not change neuromuscular parameters of the vocal folds of adult female rats; therefore, vocal changes within the outer vibratory layers of the vocal folds may primarily be responsible for clinically observed menopausal vocal changes. LEVEL OF EVIDENCE/METHODS:NA Laryngoscope, 2020.

OBJECTIVES/UNASSIGNED:(1) Determine the feasibility of obtaining a global, unbiased metabolomic profile on laryngeal muscle in a rat model; (2) evaluate the impact of biological aging on the laryngeal metabolome; and (3) characterize biochemical expression differences between aged and non-aged laryngeal and hindlimb muscle. METHODS/UNASSIGNED:Thyroarytenoid laryngeal muscle and plantaris hindlimb muscle were harvested from 5 young adult (9 months old) and 5 older adult (32 months old) F344BN rats. Tissue was processed and analyzed using LC-MS methods. Detected metabolites were compared to widely used metabolite databases and KEGG pathway enrichment was performed on significant metabolites. RESULTS/UNASSIGNED:The greatest differences in metabolite expression were between laryngeal and limb muscle with 126 different metabolites found between laryngeal and limb within the young group and 149 different metabolites within the old group. Significant hits between muscle groups highlighted amino acid differences between these tissues. There were more robust differences with age in limb muscle compared to laryngeal muscle. CONCLUSIONS/UNASSIGNED:Amino acid metabolism is a key difference between muscles of the limbs and larynx. Due to the number of differentially expressed metabolites between the 2 muscle groups, caution should be exercised when applying skeletal limb muscle physiology and biology concepts to the vocal muscles in both aged and non-aged musculoskeletal systems. Mechanisms underlying less robust effects of age on laryngeal muscle compared to limb muscle require elucidation.

The application of exercise science training knowledge has been of growing interest to voice professionals. This tutorial, derived from the authors' invited presentations from the "Exercise and the Voice" Special Session at the 2018 Voice Foundation Symposium, proposes a foundational theoretical structure based in exercise science, clarifies the wide range of variables that may influence voice training, and summarizes our present understanding of voice physiology from the perspective of muscle training. The body of literature on voice exercise was then analyzed from the perspective of this framework, identifying what we currently know and what we still have yet to learn.

The purpose of this investigation was to determine the effects of vocal training on neuromuscular junction (NMJ) morphology and muscle fiber size and composition in the thyroarytenoid muscle, the primary muscle in the vocal fold, in younger (9-month) and older (24-month) Fischer 344 × Brown Norway male rats. Over 4 or 8 weeks of vocal training, rats of both ages progressively increased their daily number of ultrasonic vocalizations (USVs) through operant conditioning and were then compared to an untrained control group. Neuromuscular junction morphology and myofiber size and composition were measured from the thyroarytenoid muscle. Acoustic analysis of USVs before and after training quantified the functional effect of training. Both 4- and 8-week training resulted in less NMJ motor endplate dispersion in the lateral portion of the thyroarytenoid muscle in rats of both ages. Vocal training and age had no significant effects on laryngeal myofiber size or type. Vocal training resulted in a greater number of USVs with longer duration and increased intensity. This study demonstrated that vocal training induces laryngeal NMJ morphology and acoustic changes. The lack of significant effects of vocal training on muscle fiber type and size suggests vocal training significantly improves neuromuscular efficiency but does not significantly influence muscle strength changes.

The goal of this study was to evaluate the effects of ovarian hormones on female rat ultrasonic vocalizations (USVs). Twenty (10 control and 10 ovariectomized) 3-month-old female rats were recorded in 3 recording conditions (elicitation, dyad, and isolation) over a full estrous cycle or time-matched duration. There were differences in USV acoustics (frequency and complexity parameters) across recording conditions but no differences in USV acoustics between control and ovariectomized groups. USVs produced in isolation had lower frequency and complexity parameters than elicited USVs for both control and ovariectomized rats. Additionally, for control rats, USV parameters of frequency, complexity, duration, and intensity changed depending on the estrous state. Therefore, although fluctuating hormone levels may influence USV acoustics, this variation can be controlled for by ovariectomizing female rats.

Laryngeal videoendoscopy is one of the main tools in clinical examinations for voice disorders and voice research. Using high-speed videoendoscopy, it is possible to fully capture the vocal fold oscillations, however, processing the recordings typically involves a time-consuming segmentation of the glottal area by trained experts. Even though automatic methods have been proposed and the task is particularly suited for deep learning methods, there are no public datasets and benchmarks available to compare methods and to allow training of generalizing deep learning models. In an international collaboration of researchers from seven institutions from the EU and USA, we have created BAGLS, a large, multihospital dataset of 59,250 high-speed videoendoscopy frames with individually annotated segmentation masks. The frames are based on 640 recordings of healthy and disordered subjects that were recorded with varying technical equipment by numerous clinicians. The BAGLS dataset will allow an objective comparison of glottis segmentation methods and will enable interested researchers to train their own models and compare their methods.