Diagnostic utility of spirometry for children with induced laryngeal obstruction or chronic non-specific cough.

PURPOSE: To determine the diagnostic utility of spirometry in distinguishing children with Induced Laryngeal Obstruction (ILO) or chronic non-specific cough (a.k.a. tic cough) from those with mild or moderate to severe asthma. METHODS: Retrospective cross sectional design. Children diagnosed with ILO (N = 70), chronic non-specific cough (N = 70), mild asthma (N = 60), or moderate to severe asthma (N = 60) were identified from the electronic medical record of a large children's hospital. Spirometry was completed before ILO, non-specific cough, or asthma diagnoses were made by pediatric laryngologists or pulmonologists. Spirometry was performed following American Thoracic Society guidelines and was interpreted by a pediatric pulmonologist. Forced Vital Capacity (FVC), Forced Expiratory Volume in 1 Second (FEV1), FEV1/FVC Ratio (FEV1/FVC), Forced Mid-Expiratory Flow 25--75 % (FEF25-75%), pulmonologist interpretation of flow volume loops, and overall exam findings were extracted from the medical record. RESULTS: Ninety seven percent of children with ILO or chronic non-specific cough presented with spirometry values within normative range. Patients with ILO, non-specific cough, and mild asthma presented with FVC, FEV1, FEV1/FVC, and FEF25-75% values in statistically similar range. Children with moderate to severe asthma presented with significantly reduced FVC (p < .001), FEV1 (p < .001), FEV1/FVC (p < .001), and FEF25-75% (p < .001) values when compared with patients in the other groups. Flow volume loops were predominantly normal for children with ILO and non-specific cough. CONCLUSIONS: Findings indicate that ILO and chronic non-specific cough can neither be diagnosed nor differentiated from mild asthma using spirometry alone. Spirometry should therefore be used judiciously with this population, bearing in mind the limitations of the procedure. Future research should determine the most effective and efficient ways of delineating ILO and non-specific cough from other respiratory conditions in children.

Examining therapy duration in adults with induced laryngeal obstruction (ILO).

OBJECTIVE: This study examined the number of therapy sessions required to sufficiently improve (exercise) induced laryngeal obstruction (EILO/ILO) symptoms for discharge. Factors predicting therapy duration were examined as was the likelihood of patients returning for additional therapy sessions following initial discharge. METHODS: Retrospective observational cohort design. Data for 350 patients were gathered from the University of Wisconsin-Madison Voice and Swallow Clinics Outcome Database. Patients (>18 years of age) diagnosed with EILO/ILO received therapy from a Speech-Language Pathologist (SLP) and were successfully discharged. EILO/ILO treatment details, symptoms, triggers, medical comorbidities, and patient demographics were collected from initial evaluations and subsequent course of therapy. RESULTS: Patients required an average of 3.59 (SD = 3.7) therapy sessions prior to discharge. A comorbid behavioral health diagnosis (p = .026), higher Vocal Handicap Index Score (p = .009) and reduced physical activity due to EILO/ILO symptoms (p = .032) were associated with increased therapy duration. Patients with ILO or EILO with secondary environmental triggers required significantly more sessions than those with exercise-induced symptoms (p < .01). Eight percent of patients returned for additional sessions following discharge. Patients returning for additional sessions all came from affluent neighborhoods as measured by the Area Deprivation Index (ADI). CONCLUSIONS: Patients with EILO/ILO required an average of 3.59 therapy sessions prior to discharge. As such, 4 sessions is a reasonable estimate for clinicians to provide patients. Six sessions may be a more conservative estimate for patients who present with a behavioral health diagnosis, a voice complaint, or reduced physical activity from EILO/ILO symptoms.

Piezo1-expressing vocal fold epithelia modulate remodeling via effects on self-renewal and cytokeratin differentiation.

Mechanoreceptors are implicated as functional afferents within mucosa of the airways and the recent discovery of mechanosensitive channels Piezo1 and Piezo2 has proved essential for cells of various mechanically sensitive tissues. However, the role for Piezo1/2 in vocal fold (VF) mucosal epithelia, a cell that withstands excessive biomechanical insult, remains unknown. The purpose of this study was to test the hypothesis that Piezo1 is required for VF mucosal repair pathways of epithelial cell injury. Utilizing a sonic hedgehog (shh) Cre line for epithelial-specific ablation of Piezo1/2 mechanoreceptors, we investigated 6wk adult VF mucosa following naphthalene exposure for repair strategies at 1, 3, 7 and 14 days post-injury (dpi). PIEZO1 localized to differentiated apical epithelia and was paramount for epithelial remodeling events. Injury to wildtype epithelium was most appreciated at 3 dpi. Shhcre/+; Piezo1loxP/loxP, Piezo2 loxP/+ mutant epithelium exhibited severe cell/nuclear defects compared to injured controls. Conditional ablation of Piezo1 and/or Piezo2 to uninjured VF epithelium did not result in abnormal phenotypes across P0, P15 and 6wk postnatal stages compared to heterozygote and control tissue. Results demonstrate a role for Piezo1-expressing VF epithelia in regulating self-renewal via effects on p63 transcription and YAP subcellular translocation-altering cytokeratin differentiation.

Risk Factors for Dysphagia in Patients Hospitalized with COVID-19.

Patients hospitalized with COVID-19 may be at risk for dysphagia and vulnerable to associated consequences. We investigated predictors for dysphagia and its severity in a cohort of patients hospitalized with COVID-19 at a single hospital center. A large level I trauma center database was queried for all patients hospitalized with COVID-19. Demographics, medical information associated with COVID-19, specific to dysphagia, and interventions were collected. 947 patients with confirmed COVID-19 met the criteria. 118 (12%) were seen for a swallow evaluation. Individuals referred for evaluation were significantly older, had a lower BMI, more severe COVID-19, and higher rates of intubation, pneumonia, mechanical ventilation, tracheostomy placements, prone positioning, and ARDS. Pneumonia (OR 3.57, p = 0.004), ARDS (OR 3.57, p = 0.029), prone positioning (OR 3.99, p = 0.036), ventilation (OR 4.01, p = 0.006), and intubation (OR 4.75, p = 0.007) were significant risk factors for dysphagia. Older patients were more likely to have more severe dysphagia such that for every 1-year increase in age, the odds of severe dysphagia were 1.04 times greater (OR 1.04, p = 0.028). Patients hospitalized with COVID-19 are at risk for dysphagia. We show predictive variables that should be considered when referring COVID-19 patients for dysphagia services to reduce time to intervention/evaluation.

Are Children with Cleft Palate at Increased Risk for Laryngeal Pathology?

To determine the prevalence of laryngeal pathology in children presenting with cleft palate with or without cleft lip (CP ± L) who underwent nasoendoscopy to assess palatal function. A secondary aim was to determine the relationship between patient demographics, resonance, articulation, and prevalence of laryngeal pathology in this population.Retrospective, observational cohort study.Outpatient pediatric cranio-facial anomalies clinic.Children ≤18 years of age presenting with CP ± L (N = 215) who underwent nasoendoscopy, speech language pathology, plastic surgery, and otolaryngological evaluations between 2009 and 2020.Laryngeal diagnosis by pediatric otolaryngologists.21.9% of children presented with laryngeal pathology. Diagnoses included benign vocal fold lesions and laryngeal edema sufficiently severe to alter vocal fold edge contour. Likelihood of laryngeal pathology increased by approximately 12% with every increase of 1 year in age (P = .001, OR = 1.12). Children with laryngeal pathology were 50% more likely to have undergone palatal repair (P < .001, OR = 1.50). In addition, children with severely hypernasal resonance were 78% less likely to present with laryngeal pathology (P =.046, OR = 0.22).This population is at increased risk for laryngeal pathologies as determined by nasoendoscopy. This finding underscores the importance of careful laryngeal imaging in assessing these children. Additional research is warranted to identify the mechanisms underlying the increased risk for morphological vocal fold changes.

Relationship Between Auditory-Perceptual and Objective Measures of Resonance in Children with Cleft Palate: Effects of Intelligibility and Dysphonia.

To investigate the relationship between auditory-perceptual ratings of resonance and nasometry scores in children with cleft palate. Factors which may impact this relationship were examined including articulation, intelligibility, dysphonia, sex, and cleft-related diagnosis.Retrospective, observational cohort study.Outpatient pediatric cranio-facial anomalies clinic.Four hundred patients <18 years of age identified with CP ± L, seen for auditory-perceptual and nasometry evaluations of hypernasality as well as assessments of articulation and voice.Relationship between auditory-perceptual ratings of resonance and nasometry scores.Pearson's correlations indicated that auditory-perceptual resonance ratings and nasometry scores were significantly correlated across oral-sound stimuli on the picture-cued portion of the MacKay-Kummer SNAP-R Test (r values .69 to.72) and the zoo reading passage (r = .72). Linear regression indicated that intelligibility (p ≤ .001) and dysphonia (p = .009) significantly impacted the relationship between perceptual and objective assessments of resonance on the Zoo passage. Moderation analyses indicated that the relationship between auditory-perceptual and nasometry values weakened as severity of speech intelligibility increased (P < .001) and when children presented with moderate dysphonia (p ≤ .001). No significant impact of articulation testing or sex were observed.Speech intelligibility and dysphonia alter the relationship between auditory-perceptual and nasometry assessments of hypernasality in children with cleft palate. SLPs should be aware of potential sources of auditory-perceptual bias and shortcomings of the Nasometer when following patients with limited intelligibility or moderate dysphonia. Future study may identify the mechanisms by which intelligibility and dysphonia affect auditory-perceptual and nasometry evaluations.

Inactivation of Lats1 and Lats2 highlights the role of hippo pathway effector YAP in larynx and vocal fold epithelium morphogenesis.

Proliferation and differentiation of vocal fold epithelial cells during embryonic development is poorly understood. We examined the role of Hippo signaling, a vital pathway known for regulating organ size, in murine laryngeal development. Conditional inactivation of the Hippo kinase genes Lats1 and Lats2, specifically in vocal fold epithelial cells, resulted in severe morphogenetic defects. Deletion of Lats1 and Lats2 caused abnormalities in epithelial differentiation, epithelial lamina separation, cellular adhesion, basement membrane organization with secondary failed cartilage, and laryngeal muscle development. Further, Lats1 and Lats2 inactivation led to failure in differentiation of p63+ basal progenitors. Our results reveal novel roles of Hippo-Lats-YAP signaling in proper regulation of VF epithelial fate and larynx morphogenesis.

Novel immortalized human vocal fold epithelial cell line: In vitro tool for mucosal biology.

Study of vocal fold (VF) mucosal biology requires essential human vocal fold epithelial cell (hVFE) lines for use in appropriate model systems. We steadily transfected a retroviral construct containing human telomerase reverse transcriptase (hTERT) into primary normal hVFE to establish a continuously replicating hVFE cell line. Immortalized hVFE across passages have cobblestone morphology, express epithelial markers cytokeratin 4, 13 and 14, induced hTERT gene and protein expression, have similar RNAseq profiling, and can continuously grow for more than 8 months. DNA fingerprinting and karyotype analysis demonstrated that immortalized hVFE were consistent with the presence of a single cell line. Validation of the hVFE, in a three-dimensional in vitro VF mucosal construct revealed a multilayered epithelial structure with VF epithelial cell markers. Wound scratch assay revealed higher migration capability of the immortalized hVFE on the surface of collagen-fibronectin and collagen gel containing human vocal fold fibroblasts (hVFF). Collectively, our report demonstrates the first immortalized hVFE from true VFs providing a novel and invaluable tool for the study of epithelial cell-fibroblast interactions that dictate disease and health of this specialized tissue.

Correlation Between EAT-10 and Aspiration Risk Differs by Dysphagia Etiology.

Agreement between self-reported dysphagic symptoms and actual swallowing physiology can vary widely across individuals. The Eating Assessment Tool-10 (EAT-10) is a self-report questionnaire commonly used to identify individuals with oropharyngeal dysphagia, but its interpretation for highly prevalent populations is poorly defined. Our primary objective was to determine if correlation strength between EAT-10 and Penetration-Aspiration Scale (PAS) scores differed by dysphagia etiology. Our secondary objective was to identify clinical factors that were associated with a mismatch between EAT-10 scores and videofluoroscopic findings. Outpatients with Parkinson disease (PD), stroke, and/or head and neck cancer (HNC) who completed EAT-10 and underwent videofluoroscopy were included (n = 203). EAT-10/PAS correlations were calculated by dysphagia etiology. We found that across the sample, higher EAT-10 scores were significantly correlated to higher PAS scores (rs = 0.31, p < 0.001). EAT-10 and PAS were moderately correlated in the HNC group (rs = 0.41, p < 0.001, n = 87), but correlations were modest in the PD (rs = 0.18, n = 41) and stroke groups (rs = 0.12, n = 59). Clinical characteristics of individuals with a "matched" profile (normal EAT-10 score and normal swallow physiology) and a "mismatched" profile (normal EAT-10 score and abnormal swallow physiology) were also compared. Individuals with a "mismatched" EAT-10/PAS profile appeared to be significantly older and had a worse Charlson Comorbidity Index than individuals with a "matched" profile. Within the HNC subgroup, EAT-10/PAS correlations for specific tumor sites, treatment types, and time since treatment are reported. Clinicians may consider these aspiration risk profiles when making recommendations for instrumented swallowing assessment.

Drainage of amniotic fluid delays vocal fold separation and induces load-related vocal fold mucosa remodeling.

In the present study, we investigated the role of mechanical load as generated by amniotic fluid in the vocal fold embryogenesis. In utero, amniotic fluid flows through the laryngeal inlet down into the lungs during fetal breathing and swallowing. In a mouse model, the onset of fetal breathing coincides with epithelial lamina recanalization. The epithelial lamina is a temporal structure that is formed during early stages of the larynx development and is gradually resorbed whereby joining the upper and lower airways. Here, we show that a temporary decrease in mechanical load secondary to drainage of amniotic fluid and subsequent flow restoration, impaired timing of epithelial lamina disintegration. Moreover, re-accumulation of fluid in the laryngeal region led to VF tissue deformation triggering remodeling of the epithelium and pressure generated changes in the elastic properties of the lamina propria, as measured by atomic force microscopy. We further show that load-related structural changes were likely mediated by Piezo 1 -Yap signaling pathway in the vocal fold epithelium. Understanding the relationship between the mechanical and biological parameters in the larynx is key to gaining insights into pathogenesis of congenital laryngeal disorders as well as mechanisms of vocal fold tissue remodeling in response to mechanotransduction.

ß-Catenin signaling is essential for mammalian larynx recanalization and the establishment of vocal fold progenitor cells.

Congenital laryngeal webs result from failure of vocal fold separation during development in utero Infants present with life-threatening respiratory problems at birth, and extensive lifelong difficulties in breathing and voicing. The molecular mechanisms that instruct vocal fold formation are rarely studied. Here, we show, for the first time, that conditional inactivation of the gene encoding ß-catenin in the primitive laryngopharyngeal epithelium leads to failure in separation of the vocal folds, which approximates the gross phenotype of laryngeal webbing. These defects can be traced to a series of morphogenesis defects, including delayed fusion of the epithelial lamina and formation of the laryngeal cecum, failed separation of the larynx and esophagus with reduced and disorganized cartilages and muscles. Parallel to these morphogenesis defects, inactivation of ß-catenin disrupts stratification of epithelial cells and establishment of p63+ basal progenitors. These findings provide the first line of evidence that links ß-catenin function to the cell proliferation and progenitor establishment during larynx and vocal fold development.

Mechanisms of larynx and vocal fold development and pathogenesis.

The larynx and vocal folds sit at the crossroad between digestive and respiratory tracts and fulfill multiple functions related to breathing, protection and phonation. They develop at the head and trunk interface through a sequence of morphogenetic events that require precise temporo-spatial coordination. We are beginning to understand some of the molecular and cellular mechanisms that underlie critical processes such as specification of the laryngeal field, epithelial lamina formation and recanalization as well as the development and differentiation of mesenchymal cell populations. Nevertheless, many gaps remain in our knowledge, the filling of which is essential for understanding congenital laryngeal disorders and the evaluation and treatment approaches in human patients. This review highlights recent advances in our understanding of the laryngeal embryogenesis. Proposed genes and signaling pathways that are critical for the laryngeal development have a potential to be harnessed in the field of regenerative medicine.

Comparing Quality of Life and Perceptual Speech Ratings in Children With Cleft Palate.

OBJECTIVE: To delineate the relationship between patient and parent-reported quality of life (QOL) ratings and perceptual characteristics of speech assigned by a speech-language pathologist (SLP) in children with repaired cleft palate. DESIGN: Prospective. SETTING: Academic Children's Hospital. PARTICIPANTS: This population-based sample included children, aged 3 to 18 with a history of repaired cleft palate, and their parents. INTERVENTION: Participants completed the Velopharyngeal Insufficiency Effects on Life Outcomes Questionnaire (VELO). Children's speech was judged perceptually by an expert SLP using the Pittsburgh Weighted Speech Scale (PWSS). MAIN OUTCOME MEASURE(S): Velopharyngeal Insufficiency Effects on Life Outcomes questionnaire assessed participant and parent perceptions of impact of velopharyngeal function on QOL. Pittsburgh Weighted Speech Scale assessed nasal emissions, facial grimacing, nasality, quality of phonation, and articulation. RESULTS: Enrollment included 48 participant parent dyads. Overall, participants reported high QOL scores within the 95% CI with children reporting slightly better yet not significantly different QOL (86.27 ± 8.96) compared to their parents (81.81 ± 15.2). Children received an average score of 1.38 ± 1.96 on the PWSS corresponding to borderline velopharyngeal competence. A significant moderate negative correlation was found between PWSS total score and parent VELO total score (r = -0.51103, P = .0002). Mild-moderate significant negative correlations were measured between PWSS total and the 5 subscales of the VELO. No significant correlations were measured between PWSS and child VELO total responses or between total scores and subscales. CONCLUSIONS: Results suggest that as perceptual analysis of speech improves, overall QOL improves moderately.

Correction to: Assessing Hyolaryngeal Excursion: Comparing Quantitative Methods to Palpation at the Bedside and Visualization During Videofluoroscopy.

This letter notifies the readers of the Dysphagia journal of an error in the original published version of this manuscript, for which a previously available open source spreadsheet tool had been used to calculate the position of the hyoid bone or larynx on lateral view videofluoroscopic images. An error in the mathematical formula built into the spreadsheet resulted in a reversal of the results for the X and Y planes of measurement. This erratum provides corrections to the results and interpretations of the original manuscript.

Tissue specific human fibroblast differential expression based on RNAsequencing analysis.

BACKGROUND: Physical forces, such as mechanical stress, are essential for tissue homeostasis and influence gene expression of cells. In particular, the fibroblast has demonstrated sensitivity to extracellular matrices with assumed adaptation upon various mechanical loads. The purpose of this study was to compare the vocal fold fibroblast genotype, known for its unique mechanically stressful tissue environment, with cellular counterparts at various other anatomic locales to identify differences in functional gene expression profiles. RESULTS: By using RNA-seq technology, we identified differentially expressed gene programs (DEseq2) among seven normal human fibroblast primary cell lines from healthy cadavers, which included: vocal fold, trachea, lung, abdomen, scalp, upper gingiva, and soft palate. Unsupervised gene expression analysis yielded 6216 genes differentially expressed across all anatomic sites. Hierarchical cluster analysis revealed grouping based on anatomic site origin rather than donor, suggesting global fibroblast phenotype heterogeneity. Sex and age-related effects were negligible. Functional enrichment analyses based on separate post-hoc 2-group comparisons revealed several functional themes within the vocal fold fibroblast related to transcription factors for signaling pathways regulating pluripotency of stem cells and extracellular matrix components such as cell signaling, migration, proliferation, and differentiation potential. CONCLUSIONS: Human fibroblasts display a phenomenon of global topographic differentiation, which is maintained in isolation via in vitro assays. Epigenetic mechanical influences on vocal fold tissue may play a role in uniquely modelling and maintaining the local environmental cellular niche during homeostasis with vocal fold fibroblasts distinctly specialized related to their anatomic positional and developmental origins established during embryogenesis.

Postextubation Dysphagia in Pediatric Populations: Incidence, Risk Factors, and Outcomes.

OBJECTIVE: To assess incidence, risk factors for, and impact on outcomes of postextubation dysphagia. We hypothesized that the incidence of postextubation dysphagia in pediatric patients would approximate or exceed that in adults, that age and duration of intubation would increase odds for postextubation dysphagia, and that the presence of postextubation dysphagia would negatively impact patient outcomes. STUDY DESIGN: We performed a retrospective, observational cohort study of patients aged 0-16 years admitted between 2011 and 2017. Patients were included if they were extubated in the intensive care unit and fed orally within 72 hours. Records were reviewed to determine dysphagia status and assess the impact of patient factors on odds of postextubation dysphagia. The impact of postextubation dysphagia on patient outcomes was then assessed. RESULTS: Following application of inclusion and exclusion criteria, the sample size was 372 patients. Postextubation dysphagia was observed in 29% of patients. For every hour of intubation, odds of postextubation dysphagia increased by 1.7% (P < .0001). Age of <25 months increased odds of postextubation dysphagia more than 2-fold (P < .05). When we controlled for age, diagnosis, number of complex chronic conditions, and dysphagia status, patients with dysphagia had an increase in total length of stay of 10.95 days (P < .0001). Postextubation dysphagia increased odds of gastrostomy or nasogastric tube at time of discharge (aOR 22.22, P < .0001). CONCLUSIONS: This study found that postextubation dysphagia is associated with increased time between extubation and discharge and with odds of gastrostomy or nasogastric tube at time of discharge.

Assessing Hyolaryngeal Excursion: Comparing Quantitative Methods to Palpation at the Bedside and Visualization During Videofluoroscopy.

PURPOSE: Hyolaryngeal excursion (HE) is typically assessed via palpation during clinical swallowing exams (CSE) or visually during videofluoroscopy (VFSS). Minimal evidence exists to support the use of these perceptual methods for judging HE. We investigated whether binary judgment of HE differentiates quantitative measures of hyoid movement, using frame-by-frame VFSS analysis to measure anatomically scaled peak hyoid positions. METHODS: Medical records of patients who received a CSE and VFSS within a 24-h period were reviewed. Clinician ratings of HE ('reduced' or 'normal') were collected from CSE and VFSS reports, along with rater experience. Five ml puree swallows were extracted from each VFSS for randomized, blinded analysis. Peak hyoid position from C4 was captured in anterior, superior, and hypotenuse positions and expressed relative to C2-C4 length. T-test comparisons of hyoid positions between patients judged to have reduced versus normal HE on palpation and VFSS were conducted. RESULTS: Eighty-seven patients (56 male, mean age 61) met criteria. Peak anterior hyoid position was significantly different between patients judged to have reduced (mean = 89.2% C2-C4) and normal (mean = 110.6% C2-C4) HE on palpation (p = 0.001). Further analysis revealed no effect of clinician experience on differentiation of objective measures based on palpation. No differences were found across any objective measures when compared to clinician VFSS ratings. CONCLUSIONS: Clinicians appeared to be able to differentiate peak anterior hyoid movement but not superior or hypotenuse movement on palpation. On VFSS visualization, no significant differences were found between swallows judged to have reduced versus normal HE in any directional dimension. While perceptual methods may contribute to clinical decision-making, clinicians should remain cautious when making judgments about HE using these methods.

Cell density, dimethylsulfoxide concentration and needle gauge affect hydrogel-induced bone marrow mesenchymal stromal cell viability.

Mesenchymal stromal cells (MSCs) have shown potential therapeutic benefits for a range of medical disorders and continue to be a focus of intense scientific investigation. Transplantation of MSCs into injured tissue can improve wound healing, tissue regeneration and functional recovery. However, implanted cells rapidly lose their viability or fail to integrate into host tissue. Hydrogel-seeded bone marrow (BM)-MSCs offer improved viability in response to mechanical forces caused by syringe needles, cell density and dimethylsulfoxide (DMSO) concentration, which in turn, will help to clarify which factors are important for enhancing biomaterial-induced cell transplantation efficiency and provide much needed guidance for clinical trials. In this study, under the control of cell density (<2 × 107 cells/mL) and final DMSO concentration (<0.5%), hydrogel-induced BM-MSC viability remained >82% following syringe needle passage by 25- or 27-gauge needles, providing improved cell therapeutic approaches for regenerative medicine.

Ontogeny of the mouse vocal fold epithelium.

This investigation provides the first systematic determination of the cellular and molecular progression of vocal fold (VF) epithelium development in a murine model. We define five principal developmental events that constitute the progression from VF initiation in the embryonic anterior foregut tube to fully differentiated and functional adult tissue. These developmental events include (1) the initiation of the larynx and vocal folds with apposition of the lateral walls of the primitive laryngopharynx (embryonic (E) day 10.5); (2) the establishment of the epithelial lamina with fusion of the lateral walls of the primitive laryngopharynx (E11.5); (3) the epithelial lamina recanalization and separation of VFs (E13.5-18.5); (4) the stratification of the vocal folds (E13.5-18.5); and (5) the maturation of vocal fold epithelium (postnatal stages). The illustration of these morphogenetic events is substantiated by dynamic changes in cell proliferation and apoptosis, as well as the expression pattern of key transcription factors, FOXA2, SOX2 and NKX2-1 that specify and pattern the foregut endoderm. Furthermore, we documented the gradual conversion of VF epithelial cells from simple precursors expressing cytokeratins 8 and 18 in the embryo into mature stratified epithelial cells also expressing cytokeratins 5 and 14 in the adult. Interestingly, in the adult, cytokeratins 5 and 14 appear to be expressed in all cell layers in the VF, in contrast to their preferential localization to the basal cell layer in surrounding epithelium. To begin investigating the role of signaling molecules in vocal fold development, we characterized the expression pattern of SHH pathway genes, and how loss of Shh affects vocal fold development in the mutant. This study defines the cellular and molecular context and serves as the necessary foundation for future functional investigations of VF formation.

Evaluation and management of the pediatric thyroid nodule.

Thyroid nodules are commonly diagnosed in adults. Although rare in children, the risk for thyroid cancer is much higher in the pediatric population compared with adults. Presenting as either a solitary nodule or a multinodular goiter, thyroid nodular disease in children requires a thorough workup that includes a detailed clinical examination comprised of prior history of thyroid disease in the patient or in their family, history of radiation exposure, careful palpation of the thyroid and lymph nodes, blood tests, ultrasonography, and cytological assessment. Thyroid surgery is the gold-standard treatment for pediatric thyroid nodules; nonetheless, the extent of surgery remains controversial. Because surgery is not without risk, the decision matrix necessitates focus on the benefits of surgery for the child contingent upon all the preoperative exams. New diagnostic technology such as molecular testing with fine needle aspiration biopsy may help distinguish between benign and malignant lesions while potentially decreasing surgery for benign disease. The objective of this review is to summarize new concepts in clinical disease management of nodular thyroid disease in the pediatric population, including patient history, medical examination, and diagnosis workup.