Ryan C. Branski

Background: Vocal tremor profoundly impacts communication, social participation, and quality of life. Although expert auditory-perceptual ratings of vocal tremor severity align with acoustic voice outcomes (e.g., extent of frequency (fo) and intensity modulation), patient perception of their voice remains unexamined despite its clinical importance. This study aimed to characterize the relationship between patient-reported vocal tremor severity and acoustic voice outcomes at baseline and after botulinum toxin injections. Method: Patients diagnosed with vocal tremor affecting multiple structures (ETvt) or tremor only observed in the larynx (LDvt) were recruited. Participants completed the voice section of the Quality of Life in Essential Tremor questionnaire to assess patient perception and performed sustained /ɑ/ at a comfortable pitch and volume, from which acoustic voice outcomes (rate and extent of fundamental frequency [fo] and amplitude [dB] modulation) were derived. A subset of participants received botulinum toxin injections and were reassessed within the therapeutic window (within 12 weeks). Results: Thirty participants (29 females; mean age = 72 years, SD = 11.40) were analyzed. Participants who rated their vocal tremor as “severe” demonstrated higher rate fo (β = 1.20, 95% CI: –0.10, 2.60 Hz) and rate dB (β = 2.30, 95% CI: 0.50, 4.10 Hz) compared to participants who rated their tremor as “moderate”. Participants who rated their tremor as “marked” demonstrated higher rate fo (β = 1.50, 95% CI: 0.30, 2.60 Hz) compared to “moderate” ratings. Improvements in patient perception of vocal tremor and acoustic outcomes were highly heterogenous among seven participants who received botulinum toxin. Discussion: Participants reporting more severe vocal tremor demonstrated more aberrant acoustic voice outcomes. After botulinum toxin injection, substantial heterogeneity was observed in acoustic voice measures which varied based on patient perception of change. These preliminary, exploratory findings provide a foundation for future investigations to define meaningful change in this population.

Introduction: Glucocorticoids (GCs) are commonly prescribed for laryngeal indications due to their potent anti‐inflammatory properties. However, GCs effect on vocal fold (VF) epithelial morphology and barrier function following injury is overlooked and may be key to efficacy. In this study, the effects of GCs on epithelial morphology and barrier function were quantified in injured VFs. We seek to increase our understanding of biochemical processes underlying GC mechanisms to refine therapeutic strategies.Methods: Microflap injury was induced in 65 rabbits. Seven days after injury, animals received bilateral 20 μL intracordal injections of saline, dexamethasone, methylprednisolone, or triamcinolone (n = 15 per condition). Five rabbits in each condition were euthanized 1, 7, or 60 days following treatment. An additional five animals served as non‐injured/untreated controls. To quantify transepithelial electrical resistance (TEER), 1 mm epithelial biopsies were placed in an Ussing chamber. The contralateral VF was processed for transmission electron microscopy and epithelial depth analysis.Results: At 60 days, GC treatment maintained TEER levels similar to non‐injured/untreated controls. However, triamcinolone reduced TEER compared with saline‐treated conditions. Acutely, epithelial hyperplasia typically persisted in all injured VFs. At 60 days, only dexamethasone and triamcinolone increased epithelial depth in injured VFs; all GCs increased epithelial depth compared with non‐injured/untreated controls.Conclusion: Acutely, GCs did not alter TEER. Additionally, GCs did not alter epithelial depth compared with saline treatment, indicating alignment with natural healing responses. At 60 days, GCs exhibited varying degrees of TEER restoration and epithelial hyperplasia, possibly due to distinct pharmacodynamic profiles.Level of Evidence: NA Laryngoscope, 2024

Objective: The diversity of glucocorticoid (GC) properties may underlie variability of clinical efficacy for vocal fold (VF) disease. Optimized therapeutic approaches must account for tissue complexity as well as interactions between cell types. We previously reported that reduced GC concentrations inhibited inflammation without eliciting fibrosis in mono‐cultured VF fibroblasts and macrophages. These data suggested that a refined approach to GC concentration may improve outcomes. In the current study, co‐culture of VF fibroblasts and macrophages was employed to investigate the effects of different concentrations of methylprednisolone on fibrotic and inflammatory response genes in VF fibroblasts to optimize management paradigms.Study Design: In vitro.Methods: THP‐1 monocyte‐derived macrophages were stimulated with interferon‐γ (IFN‐γ), lipopolysaccharide (LPS), or transforming growth factor‐β (TGF‐β) to induce inflammatory (M(IFN/LPS)) and fibrotic (M(TGF)) phenotypes. Macrophages were then co‐cultured with a human VF fibroblast cell line using a 0.4 μm pore membrane with or without 0.1–3000 nM methylprednisolone. Inflammatory (CXCL10, TNF, and PTGS2) and fibrotic (ACTA2, CCN2, and COL1A1) gene expression was quantified in fibroblasts.Results: Incubating VF fibroblasts with M(IFN/LPS) macrophages increased expression of TNF and PTGS2, and this effect was inhibited by methylprednisolone. Incubation of VF fibroblasts with M(TGF) macrophages increased expression of ACTA2, CCN2, and COL1A1, and this effect was enhanced by methylprednisolone. The concentration of methylprednisolone required to downregulate inflammatory genes (TNF and PTGS2) was lower than that to upregulate fibrotic genes (ACTA2, CCN2, and COL1A1).Conclusion: Reduced concentration of methylprednisolone effectively suppressed inflammatory genes without enhancing fibrotic genes, suggesting that a refined approach to GC concentration may improve clinical outcomes.Level of Evidence: N/A Laryngoscope, 133:3116–3122, 2023

Objective: Variable outcomes of glucocorticoid (GC) therapy for laryngeal disease are putatively due to diverse interactions of the GC receptor (GR) with cell signaling pathways, limited consideration regarding concentration‐dependent effects, and inconsistent selection of GCs. In the current study, we evaluated the concentration‐dependent effects of three frequently administered GCs on transcription factors with an emphasis on the phosphorylation of GR at Ser203 and Ser211 regulating the nuclear translocation of GR. This study provides foundational data regarding the diverse functions of GCs to optimize therapeutic approaches.Study design: In vitro.Methods: Human vocal fold fibroblasts and THP1‐derived macrophages were treated with different concentrations of dexamethasone, methylprednisolone, and triamcinolone in combination with IFN‐γ, TNF‐α, or IL4. Phosphorylated STAT1, NF‐κB family molecules, and phosphorylated STAT6 were analyzed by Western blotting. Ser211‐phosphorylated GR (S211‐pGR) levels relative to GAPDH and Ser203‐phosphorylated GR (S203‐pGR) were also analyzed.Results: GCs differentially altered phosphorylated STAT1 and NF‐κB family molecules in different cell types under IFN‐γ and TNF‐α stimuli. GCs did not alter phosphorylated STAT6 in IL4‐treated macrophages. The three GCs were nearly equivalent. A lower concentration of dexamethasone increased S211‐pGR/GAPDH ratios relative to increased S211‐pGR/S203‐pGR ratios regardless of cell type and treatment.Conclusion: The three GCs employed in two cell lines had nearly equivalent effects on transcription factor regulation. Relatively high levels of Ser203‐phosphorylation at low GC concentrations may be related to concentration‐dependent differential effects of GCs in the two cell lines.Level of Evidence: NA Laryngoscope, 133:2704–2711, 2023

Objective: Glucocorticoids (GCs) modulate multiple cellular activities including inflammatory and fibrotic responses. Outcomes of GC treatment for laryngeal disease vary, affording opportunity to optimize treatment. In the current study, three clinically employed GCs were evaluated to identify optimal in vitro concentrations at which GCs mediate favorable anti‐inflammatory and fibrotic effects in multiple cell types. We hypothesize a therapeutic window will emerge as a foundation for optimized therapeutic strategies for patients with laryngeal disease.Study Design: In vitro.Methods: Human vocal fold fibroblasts and human macrophages derived from THP‐1 monocytes were treated with 0.03–1000 nM dexamethasone, 0.3–10,000 nM methylprednisolone, and 0.3–10,000 nM triamcinolone in combination with interferon‐γ, tumor necrosis factor‐α, or interleukin‐4. Real‐time polymerase chain reaction was performed to analyze inflammatory (CXCL10, CXCl11, PTGS2, TNF, IL1B) and fibrotic (CCN2, LOX, TGM2) genes, and TSC22D3, a target gene of GC signaling. EC₅₀ and IC₅₀ to alter inflammatory and fibrotic gene expression was calculated.Results: Interferon‐γ and tumor necrosis factor‐α increased inflammatory gene expression in both cell types; this response was reduced by GCs. Interleukin‐4 increased LOX and TGM2 expression in macrophages; this response was also reduced by GCs. GCs induced TSC22D3 and CCN2 expression independent of cytokine treatment. EC₅₀ for each GC to upregulate CCN2 was higher than the IC₅₀ to downregulate other genes.Conclusion: Lower concentrations of GCs repressed inflammatory gene expression and only moderately induced genes involved in fibrosis. These data warrant consideration as a foundation for optimized clinical care paradigms to reduce inflammation and mitigate fibrosis.Level of Evidence: NA Laryngoscope, 133:1169–1175, 2023

Objectives: Functional outcomes following microflap surgery for vocal fold pathology are favorable. Although the stratified squamous epithelium appears to heal rapidly, persistent physiologic tissue alterations are likely. We sought to elucidate key biochemical processes including recruitment of immune cells, regulation of cellular junction proteins, and long‐term alterations to epithelial tissue permeability following microflap with an eye toward enhanced clinical outcomes.Methods: Forty New Zealand rabbits were assigned to eight groups (n = 5/group): no‐injury control or bilateral microflap with survival for 0 h, 12 h, 1 day, 3 days, 7 days, 30 days, and 60 days post‐microflap. The epithelium was dissected from one vocal fold and transepithelial resistance was quantified. The contralateral fold was subjected to transmission electron microscopy. Images were evaluated by a blinded rater and paracellular space dilation was quantified using ImageJ. Immune cell infiltration was evaluated and recorded qualitatively.Results: Increased innate immune response was observed 12 h as well as 7 and 30 days after microflap. At 60 days following injury, decreased epithelial resistance was observed. Paracellular spaces were dilated at all time‐points following injury.Conclusions: The vocal fold epithelium was significantly altered at 60 days following microflap. The implications for this tissue phenotype are unclear. However, compromised epithelial barrier function is implicated in various diseases and may increase the risk of subsequent injury.Level of Evidence: NA Laryngoscope, 133:350–356, 2023