Vocal Fold Epithelium Research Articles

Utility of cell viability assays for use with ex vivo vocal fold epithelial tissue.

Ex vivo models are routinely used to investigate the barrier function of the vocal fold epithelium. However, there are limited reports on assays that can be used to investigate the effect of clinically relevant challenges on vocal fold epithelial tissue viability. Our objective was to determine the utility of two assays routinely used in cell culture-a cellular metabolic activity assay and a cell membrane integrity assay-to investigate the viability of ex vivo porcine vocal fold epithelium. Prospective, ex vivo animal study. Porcine vocal folds were exposed to acrolein, hydrochloric acid, or hydrogen peroxide challenge. An untreated, sham challenge was included as a control. Assays including metabolic activity, cell membrane integrity, and histology were used to determine whether challenges reduced epithelial viability as compared to sham. Cell membrane integrity and metabolic activity assays detected reductions in viability following hydrochloric acid and hydrogen peroxide challenges but not acrolein challenge as compared to sham. No challenge produced significant changes in epithelial appearance as evidenced by light microscopy. Metabolic activity and cell membrane integrity assays are valuable tools that can be used to evaluate the viability of ex vivo vocal fold epithelial tissue following clinically relevant challenges. As viability is reduced, the ability of epithelial tissue to maintain its barrier function is compromised. Accurate assessment of viability may provide us clues into understanding mechanisms underlying vocal fold epithelial injury and disease. NA Laryngoscope, 125:E180-E185, 2015.

Dysphonia and Cystic Fibrosis

Objectives: Because cystic fibrosis transmembrane conductance regulator (CFTR) chloride channels are found in human vocal fold epithelium, we aimed to (1) assess the prevalence and severity of dysphonia in patients with cystic fibrosis (CF) and (2) compare these with control patient populations. Methods: Voice samples and the Voice Handicap Index-10 (VHI-10) were collected from patients with cystic fibrosis and control patients with and without chronic sinusitis. Voice samples underwent consensus auditory-perceptual evaluation of voice (CAPE-V) assessment by blinded speech-language pathologists. Sinus-related symptoms were assessed using the SinoNasal Outcome Test (SNOT-20). Statistical analysis was performed using unpaired t-tests and one-way analysis of variance (ANOVA). Results: Thirty-seven patients participated, including 17 with CF, 10 with chronic sinusitis, and 10 healthy controls. Prevalence of dysphonia by VHI-10 criteria was 41% in CF, 20% in chronic sinusitis, and 0% in healthy controls. CF patients had worse VHI-10 scores than healthy controls ( P = .001). There was no significant difference in VHI-10 or SNOT-20 between CF and chronic sinusitis patients, but on CAPE-V assessment, patients with CF had greater overall severity of dysphonia and strain compared with control patients with and without chronic sinusitis ( P < .001) and ( P = .022). Conclusions: Auditory-perceptual evaluation demonstrated greater severity of dysphonia in patients with cystic fibrosis than in controls. This may identify an area of clinical need as well as an opportunity for improved understanding of the role of CFTR in human voice.

The clinical utility of vocal dosimetry for assessing voice rest.

Voice rest is frequently recommended following surgical disruption of vocal fold epithelium, but patients report variable adherence to voice rest recommendations. The objective of this study was to assess the clinical utility of an ambulatory vocal dosimeter for measuring adherence to voice rest recommendations. Outcomes research. Part 1: To determine the utility of the dosimeter in nonclinical use, the relationship between self-reported voice use and dosimeter measurements was examined in normal subjects (n = 11) who prospectively logged voice use while wearing the dosimeter. Part 2: To determine clinical utility of the dosimeter, patients undergoing vocal fold surgery for which postoperative voice rest was recommended (n = 11) wore a dosimeter for 2 days prior to and 2 days after surgery. Phonation percent and sound level were compared at baseline and during voice rest. The dosimeter performed as hypothesized with both normal subjects and patients. A moderate correlation (r = 0.62) was noted between self-reported voice use and dosimeter measurements in normal subjects. In patients on voice rest, a statistically and clinically significant decrease was observed in measured voice use, both in phonation time (P = .002) and intensity of phonation (P = .004). Ambulatory vocal dosimetry may have clinical utility for assessing adherence to voice rest recommendations. This information will be useful for the design of future studies on voice rest.

Vocal fold ion transport and mucin expression following acrolein exposure.

The vocal fold epithelium is exposed to inhaled particulates including pollutants during breathing in everyday environments. Yet, our understanding of the effects of pollutants on vocal fold epithelial function is extremely limited. The objective of this study was to investigate the effect of the pollutant acrolein on two vocal fold epithelial mechanisms: ion transport and mucin (MUC) synthesis. These mechanisms were chosen as each plays a critical role in vocal defense and in maintaining surface hydration which is necessary for optimal voice production. Healthy, native porcine vocal folds (N = 85) were excised and exposed to an acrolein or sham challenge. A 60-min acrolein, but not sham challenge significantly reduced ion transport and inhibited cyclic adenosine monophosphate-dependent, increases in ion transport. Decreases in ion transport were associated with reduced sodium absorption. Within the same timeline, no significant acrolein-induced changes in MUC gene or protein expression were observed. These results improve our understanding of the effects of acrolein on key vocal fold epithelial functions and inform the development of future investigations that seek to elucidate the impact of a wide range of pollutant exposures on vocal fold health.

Vocal fold epithelial barrier in health and injury: a research review.

Vocal fold epithelium is composed of layers of individual epithelial cells joined by junctional complexes constituting a unique interface with the external environment. This barrier provides structural stability to the vocal folds and protects underlying connective tissue from injury while being nearly continuously exposed to potentially hazardous insults, including environmental or systemic-based irritants such as pollutants and reflux, surgical procedures, and vibratory trauma. Small disruptions in the epithelial barrier may have a large impact on susceptibility to injury and overall vocal health. The purpose of this article is to provide a broad-based review of current knowledge of the vocal fold epithelial barrier. A comprehensive review of the literature was conducted. Details of the structure of the vocal fold epithelial barrier are presented and evaluated in the context of function in injury and pathology. The importance of the epithelial-associated vocal fold mucus barrier is also introduced. Information presented in this review is valuable for clinicians and researchers as it highlights the importance of this understudied portion of the vocal folds to overall vocal health and disease. Prevention and treatment of injury to the epithelial barrier is a significant area awaiting further investigation.

Bicarbonate Availability for Vocal Fold Epithelial Defense to Acidic Challenge

Bicarbonate is critical for acid-base tissue homeostasis. In this study we investigated the role of bicarbonate ion transport in vocal fold epithelial defense to acid challenges. Acidic insults to the larynx are common in gastric reflux, carcinogenesis and metastasis, and acute inflammation. Ion transport was measured in viable porcine vocal fold epithelium. First, 18 vocal folds were exposed to either the carbonic anhydrase antagonist acetazolamide or to vehicle. Second, 32 vocal folds were exposed to either a control buffer or a bicarbonate-free buffer on their luminal or basolateral surface or both. Third, 32 vocal folds were challenged with acid in the presence of bicarbonate-free or control buffer. The vocal fold transepithelial resistance was greater than 300 Ω*cm(2), suggesting robust barrier integrity. Ion transport did not change after exposure to acetazolamide (p > 0.05). Exposure to bicarbonate-free buffer did not compromise vocal fold ion transport (p > 0.05). Ion transport increased after acid challenge. This increase approached statistical significance and was the greatest for the control buffer and for the bicarbonate-free buffer applied to the basolateral surface. Bicarbonate secretion may contribute to vocal fold defense against acid challenge. Our data offer a potential novel role for bicarbonate as a therapeutic agent to reduce pH abnormalities in the larynx and prevent associated pathological changes.

Methicillin‐resistant and methicillin‐sensitive Staphylococcus aureus laryngitis

Despite the fact that a wide variety of head and neck methicillin-resistant Staphylococcus aureus (MRSA) infections have been described, only four cases of MRSA laryngitis are reported in the literature. Our clinical experience suggests that this diagnosis is more common and can be more subtle that previously reported. The objective of this study was to identify and describe the clinical presentation, diagnosis, treatment, and outcomes of MRSA and methicillin-sensitive S aureus (MSSA) laryngitis, highlighting the in-office workup of these patients. Retrospective case series. All patients with a culture-proven diagnosis of S aureus laryngitis treated within the Emory Voice Center, Department of Otolaryngology-Head and Neck Surgery at Emory University between 2007 and 2011. Demographic, diagnostic, and treatment data were retrospectively collected from the patients' hospital records. Three patients with culture-proven MRSA laryngitis were identified. Three further cases of MSSA were also identified. Patients ranged in age from 34 to 74 years. All three patients with MRSA were diabetics. All six patients in the study were current or past users of cigarettes. The most common presenting symptoms were vocal roughness, vocal fatigue, and decreased vocal endurance. There were no symptoms of airway or swallowing compromise. The duration of symptoms at the time of initial assessment ranged from 3 months to 5 years, and most patients had undergone numerous previous treatments. Common signs on laryngeal examination included thickened vocal fold epithelium, whitish debris or the appearance of leukoplakia, edema, and crusting. Signs and symptoms were similar in MRSA and MSSA patients. The diagnosis was made in all patients via awake in-office culture of the larynx. All patients were treated with a prolonged course of trimethoprim-sulfamethoxazole (2-4 weeks). Although repeated courses of treatment were required, most patients had an excellent response to treatment. This is the largest, single, case series of patients with MRSA laryngitis. Our study findings suggest that the diagnosis may be more common than previously recognized, and that the presenting signs and symptoms may be subtle and similar to MSSA. Diagnosis can be made via in-office laryngeal culture. Clinicians must have a high index of suspicion for this diagnosis.

Bacterial Biofilm Associated With Chronic Laryngitis

To determine the incidence of biofilm on the vocal fold epithelium of patients with chronic laryngitis. Bacteria can grow in biofilm protected by a glycoprotein mass. Recent studies have shown the importance of biofilm in chronic otolaryngologic infections. Because chronic laryngitis is often recurrent and occasionally purulent, we hypothesized that it is biofilm related. Prospective, controlled, blinded study. Epithelial biopsy specimens from true vocal folds from patients with chronic laryngitis undergoing diagnostic laryngomicroscopy were prepared for confocal scanning laser microscopy (CSLM) and polymerase chain reaction (PCR) microarray: CSLM images were evaluated for bacterial biofilm morphologic characteristics; PCR with microarray-based diagnostic assay was used to identify the bacterial species involved. Patients with vocal fold polyp served as control cases. Tertiary care university hospital. Eighteen consecutive patients were enrolled in the study. Thirteen of them had chronic laryngitis, and 5 had vocal fold polyps. In 9 cases, the CSLM revealed bacterial growth in biofilm form, and most of these specimens (8 out of 9) were from patients with chronic laryngitis. The PCR results were positive in 13 cases, including all 9 biofilm-positive cases. Direct detection of biofilm in laryngeal biopsy specimens from patients with chronic laryngitis supports the hypothesis that chronic laryngitis may be biofilm related. Biofilm was found in 62% of the cases of chronic laryngitis. To our knowledge, this is the first report of bacterial biofilm associated with chronic laryngitis; however, further investigation is warranted before a clear conclusion can be drawn.

Tight Junction-Related Barrier Contributes to the Electrophysiological Asymmetry across Vocal Fold Epithelium

Electrophysiological homeostasis is indispensable to vocal fold hydration. We investigate tight junction (TJ)-associated components, occludin and ZO-1, and permeability with or without the challenge of a permeability-augmenting agent, histamine. Freshly excised ovine larynges are obtained from a local abattoir. TJ markers are explored via reverse transcriptase polymerase chain reaction (RT-PCR). Paracellular permeabilities are measured in an Ussing system. The gene expression of both TJ markers is detected in native ovine vocal fold epithelium. Luminal histamine treatment significantly decreases transepithelial resistance (TER) (N = 72, p<0.01) and increases penetration of protein tracer (N = 35, p<0.001), respectively, in a time-, and dose-dependent fashion. The present study demonstrates that histamine compromises TJ-related paracellular barrier across vocal fold epithelium. The detection of TJ markers indicates the existence of typical TJ components in non-keratinized, stratified vocal fold epithelium. The responsiveness of paracellular permeabilities to histamine would highlight the functional significance of this TJ-equivalent system to the electrophysiological homeostasis, which, in turn, regulates the vocal fold superficial hydration.

Acute Stress to Excised Vocal Fold Epithelium From Reactive Oxygen Species

Acute Stress to Excised Vocal Fold Epithelium From Reactive Oxygen Species

Objective and Subjective Evaluation of Larynx in Smokers and Nonsmokers: A Comparative Study

The deleterious effects of smoking on laryngeal mucosa are indisputable. Smoking not only causes histologic (microscopic) alterations in the vocal fold epithelium but also affect the acoustic property of voice. The goal of this study was to determine the effects of smoking on the laryngeal structures and phonatory physiology via videostroboscopy, acoustic voice analysis and electroglottography. Cross-sectional and observational study was carried out in Department of ENT, Pt. J. N. M. Medical College, Raipur, in 100 male volunteers (50 smokers and 50 nonsmokers) between the ages of 18 and 60 years with no voice complaints. All subjects were analyzed for detailed history and underwent videostroboscopic, acoustic voice analysis and electroglottographic examination. Videostroboscopic examination results showed significant relationships between smoking and abnormal vocal fold edge, abnormal texture, edema, erythema, abnormal mucosal cover, abnormal phase symmetry and abnormal pliability/stiffness. Voice analysis showed that smoking has a clear effect on some acoustic voice and electroglottographic parameters. The parameters most commonly affected by smoking are fundamental frequency (F0), jitter, shimmer, F0 tremor, normalized noise energy, signal noise ratio, maximum phonation time (MPT), s/z ratio, opening rate and closing rate. The results indicate that smoking has a significant effect on laryngeal structures and phonatory physiology.

Role for Ion Transport in Porcine Vocal Fold Epithelial Defense to Acid Challenge

The vocal fold epithelium is routinely exposed to gastric contents, including acid and pepsin, during laryngopharyngeal reflux events. The epithelium may possess intrinsic defenses to reflux. The first objective of the current study was to examine whether vocal fold epithelial ion transport is one potential mechanism of defense to gastric contents. The second objective was to determine whether ion transport in response to gastric contents is associated with the secretion of bicarbonate. Prospective design in excised porcine larynges. Laboratory. Porcine vocal folds (N = 56) were exposed on the luminal surface to acid, pepsin, or sham challenges. Ion transport at baseline and following challenge exposure was measured using electrophysiological techniques. To examine specific ion transport mechanisms, vocal folds were pretreated with either a sodium channel blocker or bicarbonate channel blocker. Within 60 seconds of acid but not pepsin exposure, there was a significant increase in ion transport. This rapid increase in ion transport was transient and related to bicarbonate secretion. The current data suggest that porcine vocal folds immediately increase bicarbonate secretion following exposure to acid. Bicarbonate secretion may act to neutralize acid. These findings contribute to the identification of the mechanisms underlying vocal fold defense to reflux and offer implications for the development of treatments for reflux-induced vocal fold injury.

Acute stress to excised vocal fold epithelium from reactive oxygen species.

Vocal fold epithelium is exposed to reactive oxygen species from the inhaled environment and from tissue inflammation. The objective of this study was to explore the functional and structural consequences of reactive oxygen species exposure on vocal fold epithelium. In vitro, prospective study design. Hydrogen peroxide (H(2)O(2)), a common reactive oxygen species, was utilized in this study. Freshly excised, viable porcine vocal fold epithelia (N = 32) were exposed to H(2) O(2) or sham challenge for 2 hours. Electrophysiology, western blotting, and light microscopy were used to quantify the functional and structural effects of reactive oxygen species on vocal fold epithelia. Exposure to reactive oxygen species did not significantly alter transepithelial resistance. There was a small, nonsignificant trend for decreased concentration of epithelial junctional complex protein with reactive oxygen species challenge. Minimal changes to the gross structural appearance of vocal fold epithelia were also noted. The stratified squamous epithelia of the vocal folds effectively defend against an acute reactive oxygen species challenge. The current study lays the groundwork for future investigations on the effects of reactive oxygen species on vocal fold epithelia that are compromised from phonotrauma.

Variations of the Vocal Fold Epithelium in a Menopause Induced Model

Durante la menopausia ocurren cambios en las estructuras laringeas que tienen implicancias en la voz. El objetivo de este estudio es determinar el efecto de la menopausia inducida sobre parametros morfologicos de la mucosa de los pliegues vocales en ratas de laboratorio. Se analizaron 10 ratas Sprague-Dowley hembras adultas, divididas en dos grupos, 5 fueron ooforectomizadas quirurgicamente (OVX) y 5 fueron sometidas a un procedimiento SHAM (SHAM). Luego de 30 dias postcirugia, se observaron las caracteristicas del epitelio del pliegue vocal, en terminos de disposicion celular y organizacion del epitelio. Adicionalmente con la ayuda del software Morphometrics XS se determino la altura del epitelio y la densidad de numero de estratos celulares. Nuestros resultados indican que existen alteraciones en numero de estratos celulares, asi como tambien en caracteristicas como cohesion celular, acompanado de un aumento de la matriz extracelular. El numero de estratos celulares fue significativamente mayor (p<0,01) en el grupo SHAM con 6,66 (DS 1,07), mientras que en el OVX fue de 3,2 (DS 0,83). El grosor medio del epitelio fue de 534,71 µm (DS 119,89), mientras que en el grupo SHAM fue de 486,84 µm (DS 82,95), estas diferencias no fueron estadisticamente significativas (p=0,112). Se sugiere que alteraciones en las caracteristicas del epitelio que reviste los pliegues vocales pueden relacionarse con alteraciones clinicas como la disminucion de la calidad vocal y la presbifonia caracteristicas de la mujer postmenopausica.

Localization of Label-Retaining Cells in Murine Vocal Fold Epithelium

Epithelial homeostasis is critical for vocal fold health, yet little is known about the cells that support epithelial self-renewal. As a known characteristic of stem cells is that they are slow-cycling in vivo, the purpose of this prospective controlled study was to identify and quantify slow-cycling cells or putative stem cells in murine vocal fold epithelium. Twelve mice were administered daily intraperitoneal injections of a nucleotide dye, bromodeoxyuridine (BrdU), over 7 consecutive days. Under this pulse-chase paradigm, slow-cycling cells retain the dye (label-retaining cells; LRCs) while more rapidly cycling cells lose dye to dilution during multiple cell divisions. The percentage of label-retaining cells (%LRCs) was calculated following a chase period of 2, 4, and 8 weeks postinjections. The %LRCs decreased significantly from 9.4% at 2 weeks to 3.1% at 8 weeks following injections (p < .05). No statistically significant differences in the quantity of BrdU-positive cells were measured between the anterior, mid-membranous, or cartilaginous regions of the vocal fold (p > .05). These findings are consistent with the presence and first report of a small population of putative stem cells along the length of murine vocal fold epithelium.

Raised intensity phonation compromises vocal fold epithelial barrier integrity

We investigated the hypothesis that 30 minutes of raised intensity phonation alters transcript levels of vocal fold intercellular tight junction proteins and disrupts the vocal fold epithelial barrier. Prospective animal study. Eighteen New Zealand white breeder rabbits were randomly assigned to receive 30 minutes of raised intensity phonation or approximation of the vocal folds without phonation. Quantitative polymerase chain reaction (qPCR) was used to investigate transcript levels of the epithelial intercellular tight junction proteins, occludin and zonula occludin-1 (ZO-1), and the adherens junction proteins β-catenin and E-cadherin. Structural alterations to the vocal fold epithelium were further examined by scanning electron microscopy (SEM) and transmission electron microscopy (TEM). Mann-Whitney U revealed significantly decreased occludin (P = .016) and β-catenin (P = .016) gene expression from rabbits undergoing raised intensity phonation compared with control. There were no significant differences in Z0-1 and E-cadherin gene expression between groups (P > .025). SEM revealed significant obliteration, desquamation, and evidence of microhole formation in rabbit vocal folds exposed to raised intensity phonation compared with control, whereas TEM revealed dilated intercellular morphology between groups. Results provide support for the hypothesis that a transient episode of raised intensity phonation alters transcript levels of vocal fold intercellular tight junction proteins and disrupts integrity of the epithelial barrier. The loss of barrier integrity may have significant consequences on epithelial defenses and compromise protection of the underlying mucosa from damage secondary to prolonged vibration exposure.

E-Cadherin and Transglutaminase-1 Epithelial Barrier Restoration Precedes Type IV Collagen Basement Membrane Reconstruction following Vocal Fold Mucosal Injury

The vocal fold epithelium is critical to upper airway immunologic defense and water/ion transport; therefore, any form of physical trauma or insult increases the vulnerability of this structure to functional impairment and pathogen invasion/infection. In this study, we examined the reestablishment of epithelial and basement membrane barrier structures in a well-established rat model of vocal fold mucosal injury. We observed active cell recruitment culminating in peak hyperplasia at 3 days postinjury, the establishment of robust E-cadherin+ and transglutaminase-1+ biochemical barrier signals along the epithelial surface by 3 days postinjury, and the persistent absence of a type IV collagen+ basement membrane at 7 days postinjury. The distinct spatial and temporal immunoactivity of these molecules is consistent with a programmed repair process driving the restoration of vocal fold mucosal integrity and permeability. These data may inform future efforts to optimize functional mucosal recovery postinjury and avoid undesirable events such as barrier compromise or epithelial metaplasia.

Simulated reflux decreases vocal fold epithelial barrier resistance

The vocal fold epithelium provides a barrier to the entry of inhaled and systemic challenges. However, the location of the epithelium makes it vulnerable to damage. Past research suggests, but does not directly demonstrate, that exposure to gastric reflux adversely affects the function of the epithelial barrier. Understanding the nature of reflux-induced epithelial barrier dysfunction is necessary to better recognize the mechanisms for vocal fold susceptibility to this disease. Therefore, we examined the effects of physiologically relevant reflux challenges on vocal fold transepithelial resistance and gross epithelial and subepithelial appearance. Ex vivo, mixed design with between-group and repeated-measures analyses. Healthy, native porcine vocal folds (N = 52) were exposed to physiologically relevant acidic pepsin, acid-only, or pepsin-only challenges and examined with electrophysiology and light microscopy. For all challenges, vocal folds exposed to a neutral pH served as control. Acidic pepsin and acid-only challenges, but not pepsin-only or control challenges significantly reduced transepithelial resistance within 30 minutes. Reductions in transepithelial resistance were irreversible. Challenge exposure produced minimal gross changes in vocal fold epithelial or subepithelial appearance as evidenced by light microscopy. These findings demonstrate that acidic environments characteristic of gastric reflux compromise epithelial barrier function without gross structural changes. In healthy, native vocal folds, reductions in transepithelial resistance could reflect reflux-related epithelial disruption. These results might guide the development of pharmacologic and therapeutic recommendations for patients with reflux, such as continued acid-suppression therapy and patient antireflux behavioral education.

Cervids with different vocal behavior demonstrate different viscoelastic properties of their vocal folds

The authors test the hypothesis that vocal fold morphology and biomechanical properties covary with species-specific vocal function. They investigate mule deer (Odocoileus hemionus) vocal folds, building on, and extending data on a related cervid, the Rocky Mountain elk (Cervus elaphus nelsoni). The mule deer, in contrast to the elk, is a species with relatively little vocal activity in adult animals. Mule deer and elk vocal folds show the typical three components of the mammalian vocal fold (epithelium, lamina propria and thyroarytenoid muscle). The vocal fold epithelium and the lamina propria were investigated in two sets of tensile tests. First, creep rupture tests demonstrated that ultimate stress in mule deer lamina propria is of the same magnitude as in elk. Second, cyclic loading tests revealed similar elastic moduli for the vocal fold epithelium in mule deer and elk. The elastic modulus of the lamina propria is also similar between the two species in the low-strain region, but differs at strains larger than 0.3. Sex differences in the stress-strain response, which have been reported for elk and human vocal folds, were not found for mule deer vocal folds. The laminae propriae in mule deer and elk vocal folds are comparatively large. In general, a thick and uniformly stiff lamina propria does not self-oscillate well, even when high subglottic pressure is applied. If the less stiff vocal fold seen in elk is associated with a differentiated lamina propria it would allow the vocal fold to vibrate at high tension and high subglottic pressure. The results of this study support the hypothesis that viscoelastic properties of vocal folds varies with function and vocal behavior.

Hypertonic challenge to porcine vocal folds: Effects on epithelial barrier function

Objective Dehydration challenges can increase the chemical composition of surface fluid overlying vocal fold epithelia (hypertonic surface fluid). The vocal fold epithelium is posited to act as a barrier, shielding the lamina propria from perturbations in the airway lumen. However, the effects of hypertonic surface fluid on the barrier functions of vocal fold epithelia have not been quantified. We, therefore, sought to investigate whether hypertonic surface fluid compromises epithelial barrier function. We examined the effects of hypertonic surface fluid on vocal fold epithelial resistance, paracellular pathway morphology, and tight junction protein integrity. Study Design Ex vivo, between group design. Setting Laboratory. Methods Porcine vocal folds (n = 24) were exposed to hypertonic or isotonic challenge and examined by electrophysiology, transmission electron microscopy, and Western blot analyses. Results Hypertonic, but not isotonic, challenge significantly reduced transepithelial resistance. This decrease in resistance was observed immediately after the challenge and was consistent with the appearance of dilated paracellular pathway morphology. However, hypertonic challenge did not alter protein levels of occludin, zona occludens-1, E-cadherin, or β-catenin. Conclusion Hypertonic surface fluid alters epithelial barrier function in the vocal folds. Specifically, exposure to hypertonic challenges increases epithelial permeability. Given the important role of the vocal fold epithelium in shielding the underlying mucosa from inhaled pathogens and pollutants, our data provide the impetus for future studies on pharmacological treatments aimed at restoring the hydration level and chemical composition of vocal fold surface fluid.