Is Acupuncture Efficacious for Treating Phonotraumatic Vocal Pathologies? A Randomized Control Trial.

OBJECTIVES: To investigate the effectiveness of acupuncture in treating phonotraumatic vocal fold lesions. STUDY DESIGN/METHODS: A total of 123 dysphonic individuals with benign vocal pathologies were recruited. They were given either genuine acupuncture (n = 40), sham acupuncture (n = 44), or no treatment (n = 39) for 6 weeks (two 30-minute sessions/wk). The genuine acupuncture group received needles puncturing nine voice-related acupoints for 30 minutes, two times a week for 6 weeks, whereas the sham acupuncture group received blunted needles stimulating the skin surface of the nine acupoints for the same frequency and duration. The no-treatment group did not receive any intervention but attended just the assessment sessions. One-hundred seventeen subjects completed the study (genuine acupuncture = 40; sham acupuncture = 43; and no treatment = 34), but only 84 of them had a complete set of vocal functions and quality of life measures (genuine acupuncture = 29; sham acupuncture = 33; and no-treatment = 22) and 42 of them with a complete set of endoscopic data (genuine acupuncture = 16; sham acupuncture = 15; and no treatment = 11). RESULTS: Significant improvement in vocal function, as indicated by the maximum fundamental frequency produced, and also perceived quality of life, were found in both the genuine and sham acupuncture groups, but not in the no-treatment group. Structural (morphological) improvements were, however, only noticed in the genuine acupuncture group, which demonstrated a significant reduction in the size of the vocal fold lesions. CONCLUSIONS: The findings showed that acupuncture of voice-related acupoints could bring about improvement in vocal function and healing of vocal fold lesions.

Wound-healing effect of acupuncture for treating phonotraumatic vocal pathologies: A cytokine study.

OBJECTIVES/HYPOTHESIS: Acupuncture is a less-invasive procedure when compared with surgical treatment for benign vocal pathologies caused by vocal overuse. This study aimed to determine the wound-healing effect of acupuncture in treating phonotraumatic vocal fold lesions. STUDY DESIGN: Two-way, mixed-model, between- and within-subjects, prospective randomized, placebo-controlled, blinded group design. METHODS: Seventeen dysphonic individuals with vocal nodules were recruited from a university clinic in Hong Kong. Each participant was randomly assigned to receive one session of either genuine or sham acupuncture. The genuine acupuncture group (n = 9) received needles puncturing nine voice-related acupoints for 30 minutes, whereas the sham acupuncture group (n = 8) received blunted needles stimulating the skin surface of the nine acupoints for the same frequency and duration. Laryngeal secretions were suctioned from the surface of the vocal folds immediately before, immediately after, and 24 hours after the acupuncture. The protein concentration levels of wound-healing-related cytokines (interleukin [IL]-1ß and IL-10) in these secretion samples were measured. RESULTS: Following acupuncture, a significant increase in the anti-inflammatory cytokine IL-10 was found in the genuine acupuncture group (n = 9) but not in the sham acupuncture group (n = 8). CONCLUSIONS: The findings showed that acupuncture of voice-related acupoints facilitated an anti-inflammatory process in phonotraumatic vocal pathologies. This could be considered as supporting evidence to consider acupuncture as a less-invasive alternative option, when compared to surgery, for treating phonotraumatic vocal pathologies.

Microstructural and mechanical characterization of scarred vocal folds.

The goal of this study was to characterize the vocal folds microstructure and elasticity using nonlinear laser scanning microscopy and atomic force microscopy-based indentation, respectively. As a pilot study, the vocal folds of fourteen rats were unilaterally injured by full removal of lamina propria; the uninjured folds of the same animals served as controls. The area fraction of collagen fibrils was found to be greater in scarred tissues two months after injury than the uninjured controls. A novel mathematical model was also proposed to relate collagen concentration and tissue bulk modulus. This work presents a first step towards systematic investigation of microstructural and mechanical characteristics in scarred vocal fold tissue.

Study of extracellular matrix in vocal fold biomechanics using a two-phase model.

The extracellular matrix (ECM) of the vocal fold tissue consists primarily of fibrous and interstitial proteins. The purpose of this study was to investigate the effects of selective enzymatic digestion of two ECM proteins, namely elastin and versican, on the elasticity of rabbit vocal fold tissue. Quasi-static, sinusoidal, uniaxial tensile tests were performed. The data were analyzed within the framework of a model of the ECM as a two-phase composite material consisting of collagen fibrils as the reinforcing fibers and noncollagenous ECM proteins as the matrix. To validate the two-phase model, the regression parameters for the fibers' volume fraction and shear modulus in a different animal model were compared with corresponding published data. The proposed model was then used to analyze rabbit vocal fold tissues. The mean value and the standard deviation of the fiber volume fraction were found to be 8.49 ± 3.75 % for the control samples (n = 4), 0.59 ± 1.13 % after elastin removal (n = 4), and 8.22 ± 1.06 % after versican removal (n = 4). The results suggest that elastin removal may lead to a reduction in tissue stiffness, through counteracting the reinforcement of collagen fibrils.

An in vivo study of composite microgels based on hyaluronic acid and gelatin for the reconstruction of surgically injured rat vocal folds.

PURPOSE The objective of this study was to investigate local injection with a hierarchically microstructured hyaluronic acid-gelatin (HA-Ge) hydrogel for the treatment of acute vocal fold injury using a rat model. METHOD Vocal fold stripping was performed unilaterally in 108 Sprague-Dawley rats. A volume of 25 µl saline (placebo controls), HA-bulk, or HA-Ge hydrogel was injected into the lamina propria (LP) 5 days after surgery. The vocal folds were harvested at 3, 14, and 28 days after injection and analyzed using hematoxylin and eosin staining and immunohistochemistry staining for macrophages, myofibroblasts, elastin, collagen type I, and collagen type III. RESULTS The macrophage count was statistically significantly lower in the HA-Ge group than in the saline group (p < .05) at Day 28. Results suggested that the HA-Ge injection did not induce inflammatory or rejection response. Myofibroblast counts and elastin were statistically insignificant across treatment groups at all time points. Increased elastin deposition was qualitatively observed in both HA groups from Day 3 to Day 28, and not in the saline group. Significantly more elastin was observed in the HA-bulk group than in the uninjured group at Day 28. Significantly more collagen type I was observed in the HA-bulk and HA-Ge groups than in the saline group (p < .05) at Day 28. The collagen type I concentration in the HA-Ge and saline groups was found to be comparable to that in the uninjured controls at Day 28. The concentration of collagen type III in all treatment groups was similar to that in uninjured controls at Day 28. CONCLUSION Local HA-Ge and HA-bulk injections for acute injured vocal folds were biocompatible and did not induce adverse response.

Role of steroids in acute phonotrauma: A basic science investigation.

OBJECTIVES/HYPOTHESIS: Steroids are used for the treatment of laryngitis in vocal performers and other individuals despite the absence of evidence demonstrating their impact on vocal fold inflammation. Our objective was to examine laryngeal secretion cytokine inflammatory profile changes associated with corticosteroid treatment in a human phonotrauma model. STUDY DESIGN: Prospective, individual, randomized, double-blinded, controlled trial. METHODS: Participants included 10 healthy females who were randomized to either treatment with oral hydrocortisone or placebo, each given in three doses over 20 hours after the experimental induction of acute phonotrauma. Cytokines associated with inflammation and healing (interleukin [IL]-1ß, IL-6, IL-10) were measured in laryngeal secretions before and after vocal loading and at 4 and 20 hours after treatment. RESULTS: Proinflammatory mediators IL-1ß and IL-6 were doubled in the controls versus the steroid treatment group at 21 hours following induction of acute vocal fold inflammation. Anti-inflammatory cytokine IL-10 showed a 6.3-fold increase in the steroid treatment group versus the controls, indicating anti-inflammatory modulation by steroid treatment. CONCLUSIONS: This study provides biologic evidence supporting the use of steroids for acute vocal fold inflammation associated with phonotrauma. LEVEL OF EVIDENCE: 1b.

Dose-dependent effect of mitomycin C on human vocal fold fibroblasts.

BACKGROUND: The purpose of this study was to evaluate in vitro cytotoxicity and antifibrotic effects of mitomycin C on normal and scarred human vocal fold fibroblasts. METHODS: Fibroblasts were subjected to mitomycin C treatment at 0.2, 0.5, or 1 mg/mL, or serum control. Cytotoxicity, immunocytochemistry, and Western blot for collagen I/III were performed at days 0, 1, 3, and 5. RESULTS: Significant decreases in live cells were measured for mitomycin C-treated cells on days 3 and 5 for all doses. Extracellular staining of collagen I/III was observed in mitomycin C-treated cells across all doses and times. Extracellular staining suggests apoptosis with necrosis, compromising the integrity of cell membranes and release of cytosolic proteins into the extracellular environment. Western blot indicates inhibition of collagen at all doses except 0.2 mg/mL at day 1. CONCLUSION: A total of 0.2 mg/mL mitomycin C may provide initial and transient stimulation of collagen for necessary repair to damaged tissue without the long-term risk of fibrosis.

Assessment of fine needle aspiration feasibility and specimen adequacy for molecular diagnostics of benign vocal fold lesions.

OBJECTIVES/ HYPOTHESIS: The use of molecular testing is becoming more significant for the diagnosis and classification of disease. The application of fine-needle aspiration (FNA) biopsy as the means of sampling lesions in union with molecular testing could be a powerful combination in laryngology. The objectives of this study were to investigate 1) if FNA was feasible to sample benign vocal fold lesions; 2) if FNA samples provided sufficient RNA quality for molecular analysis; and 3) if gene expression of FNA samples matched paired surgical excised specimens. STUDY DESIGN: Prospective cross-sectional. METHODS: Fifteen vocal fold specimens were obtained from adult patients undergoing routine surgical removal for benign vocal fold lesions using FNA and surgical excision. Comparisons were made between FNA and excision biopsies for RNA quality. Correlative analysis was completed for RNA expression of nine genes, including decorin (DCN), connective tissue growth factor (CTGF), vascular endothelial growth factor (VEGF), collagen type VI alpha 3 (COL6A3), superoxide dismutase 1 (SOD1), glutathione S-transferase (GST2), collagen type I alpha 2 (COL1A2), ATP binding cassette (ABC), and procollagen I alpha 1 (COL1A1). RESULTS: FNA and excision samples demonstrated similar RNA quality (P > 0.05). Per gene expression, four out of nine genes were moderately correlated between the paired samples (P < 0.05). CONCLUSIONS: FNA of the vocal fold lamina propria is technically feasible to perform. Further improvement in the FNA technology is desirable to optimize RNA quality for reliable gene expression analysis.

Current Understanding and Future Directions for Vocal Fold Mechanobiology.

The vocal folds, which are located in the larynx, are the main organ of voice production for human communication. The vocal folds are under continuous biomechanical stress similar to other mechanically active organs, such as the heart, lungs, tendons and muscles. During speech and singing, the vocal folds oscillate at frequencies ranging from 20 Hz to 3 kHz with amplitudes of a few millimeters. The biomechanical stress associated with accumulated phonation is believed to alter vocal fold cell activity and tissue structure in many ways. Excessive phonatory stress can damage tissue structure and induce a cell-mediated inflammatory response, resulting in a pathological vocal fold lesion. On the other hand, phonatory stress is one major factor in the maturation of the vocal folds into a specialized tri-layer structure. One specific form of vocal fold oscillation, which involves low impact and large amplitude excursion, is prescribed therapeutically for patients with mild vocal fold injuries. Although biomechanical forces affect vocal fold physiology and pathology, there is little understanding of how mechanical forces regulate these processes at the cellular and molecular level. Research into vocal fold mechanobiology has burgeoned over the past several years. Vocal fold bioreactors are being developed in several laboratories to provide a biomimic environment that allows the systematic manipulation of physical and biological factors on the cells of interest in vitro. Computer models have been used to simulate the integrated response of cells and proteins as a function of phonation stress. The purpose of this paper is to review current research on the mechanobiology of the vocal folds as it relates to growth, pathogenesis and treatment as well as to propose specific research directions that will advance our understanding of this subject.

Vocal exercise may attenuate acute vocal fold inflammation.

OBJECTIVES/HYPOTHESES: The objective was to assess the utility of selected "resonant voice" (RV) exercises for the reduction of acute vocal fold inflammation. The hypothesis was that relatively large-amplitude, low-impact vocal fold exercises associated with RV would reduce inflammation more than spontaneous speech (SS) and possibly more than voice rest. STUDY DESIGN: The study design was prospective, randomized, and double blind. METHODS: Nine vocally healthy adults underwent a 1-hour vocal loading procedure, followed by randomization to a SS condition, vocal rest condition, or RV exercise condition. Treatments were monitored in clinic for 4 hours and continued extraclinically until the next morning. At baseline (BL), immediately after loading, after the 4-hour in-clinic treatment, and 24 hours post-BL, secretions were suctioned from the vocal folds bilaterally and submitted to enzyme-linked immunosorbent assay to estimate concentrations of key markers of tissue injury and inflammation: interleukin (IL)-1ß, IL-6, IL-8, tumor necrosis factor α, matrix metalloproteinase (MMP)-8, and IL-10. RESULTS: Complete data sets were obtained for three markers--IL-1ß, IL-6, and MMP-8--for one subject in each treatment condition. For these markers, results were poorest at 24-hour follow-up in the SS condition, sharply improved in the voice rest condition, and was the best in the RV condition. Average results for all markers and responsive subjects with normal BL mediator concentrations revealed an almost identical pattern. CONCLUSIONS: Some forms of tissue mobilization may be useful to attenuate acute vocal fold inflammation.

Preliminary data on prevention and treatment of voice problems in student teachers.

OBJECTIVES/HYPOTHESES: To assess the utility of a targeted voice hygiene (VH) program compared to VH plus voice training intervention (VH+VT) for the prevention and treatment of voice problems in student teachers. STUDY DESIGN: Prospective, randomized. METHODS: Thirty-one student teachers with low (good) and high (poor) voice handicap index (VHI) scores in Pittsburgh and Hong Kong were randomly assigned to (1) a targeted, individually tailored VH program, (2) the VH program plus resonant VT (VH+VT), or (c) a control group. Participants assigned to intervention groups were monitored for their adherence to their programs for their first 4 weeks of student teaching. VHI data were collected again 4 weeks postintervention (both sites) and 8 weeks postintervention, following a no-contact washout period (Pittsburgh). RESULTS: Descriptive data analysis indicated that across both sites, for initially healthy participants, the VH program was sufficient to prevent worsening of VHI scores that occurred in all control participants over the first 4-8 weeks of student teaching. The addition of VT did not consistently enhance protective benefits over VH alone. In contrast, for participants with initially poor VHI scores, the VH program failed to produce VHI benefits over the control condition. The addition of VT was required to optimize results for that cohort. CONCLUSIONS: Preliminary data suggest that a minimalist, individually tailored VH program may be sufficient to prevent voice problems from teaching in healthy student teachers. However, for student teachers with existing voice problems, VT may be required to optimize results of intervention.

Temporal and spatial expression of high-mobility group box 1 in surgically injured rat vocal folds.

OBJECTIVES/HYPOTHESIS: High-mobility group box 1 (HMGB1) protein has been identified as a principal instigator of injury-induced inflammation in many organ systems. Physiologically, HMGB1 binds to chromatin in cell nucleus. Upon injury, cells release HMGB1 to extracellular milieu, triggering a destructive inflammatory response. Neutralizing or removing HMGB1 has been shown to control inflammation. Unfortunately, the role of HMGB1 in laryngeal inflammation and healing has yet to be defined. The purpose of this study was to determine spatial and temporal patterns of HMGB1 expression in surgically injured rat vocal folds up to 2 weeks after injury. STUDY DESIGN: Prospective animal study. METHODS: Bilateral vocal fold injury was performed on 70 Sprague-Dawley rats. An additional 14 rats served as uninjured controls. Animals were sacrificed at 1 day, 3 days, 5 days, 1 week, and 2 weeks following surgery. Immunohistochemistry staining and enzyme-linked immunosorbent assay (ELISA) were performed to determine the spatial distribution and temporal expression, respectively, of HMGB1 in vocal fold tissue. Hematoxylin-and-eosin staining for cell counting was performed to evaluate cell infiltration. RESULTS: Cell number peaked significantly 5 days after injury. HMGB1 was positively stained in the nuclear, cytoplasmic, and extracellular compartments from days 1 to 7 after injury, whereas a strict nuclear staining was observed in uninjured controls and week 2 animals. Staining results were corroborated by ELISA. CONCLUSIONS: Spatial and temporal changes of HMGB1 expression were shown in injured vocal fold tissue, indicating this protein may be one of the principal drivers of inflammation and healing response to surgical injury in the larynx.

Biosimulation of acute phonotrauma: an extended model.

OBJECTIVES/HYPOTHESIS: Personalized, preemptive, and predictive medicine is a central goal of contemporary medical care. The central aim of the present study was to investigate the utility of mechanistic computational modeling of inflammation and healing to address personalized therapy for patients with acute phonotrauma. STUDY DESIGN: Computer simulation. METHODS: Previously reported agent-based models (ABMs) of acute phonotrauma were extended with additional inflammatory mediators as well as extracellular matrix components. The models were calibrated with empirical data for a panel of biomarkers--interleukin (IL)-1ß, IL-6, IL-8, IL-10, tumor necrosis factor-α and matrix metalloproteinase-8--from individual subjects following experimentally induced phonotrauma and a randomly assigned voice treatment namely voice rest, resonant voice exercise, and spontaneous speech. The models' prediction accuracy for biomarker levels was tested for a 24-hour follow-up time point. RESULTS: The extended ABMs reproduced and predicted trajectories of biomarkers seen in experimental data. The simulation results also agreed qualitatively with various known aspects of inflammation and healing. Model prediction accuracy was generally better following individual-based calibration as compared to population-based calibration. Simulation results also suggested that the special form of vocal fold oscillation in resonant voice may accelerate acute vocal fold healing. CONCLUSIONS: The calibration of inflammation/healing ABMs with subject-specific data appears to optimize the models' prediction accuracy for individual subjects. This translational application of biosimulation might be used to predict individual healing trajectories, the potential effects of different treatment options, and most importantly, provide new understanding of health and healing in the larynx and possibly in other organs and tissues as well.

Possible cross-cultural differences in the perception of impact of voice disorders.

UNLABELLED: Perception of the impact of voice disorders may differ across different cultural backgrounds. This study investigated the difference in the perception of the impact of voice disorders between the American (Pittsburgh) and Chinese (Hong Kong) cultures. STUDY DESIGN AND SETTING: Sixty dysphonic subjects from Hong Kong, China, and 60 dysphonic subjects from Pittsburgh, USA, were recruited to complete the Chinese and English versions of the Voice Activity and Participation Profile (VAPP), respectively. Data analyses using independent t tests were conducted on (1) the total profile scores; (b) total activity limitation score (ALS) and total participation restriction score (PRS); (c) section scores of job, daily communication, and social communication; and (d) section ALS and section PRS. RESULTS: Hong Kong subjects showed significantly higher scores than the Pittsburgh subjects in total profile and other subsections except in the job PRS. CONCLUSION: RESULTS support the possible influence of individualist and collectivist cultures on the perceived impact of voice disorders on the activity and participation.

Biosimulation of inflammation and healing in surgically injured vocal folds.

OBJECTIVES: The pathogenesis of vocal fold scarring is complex and remains to be deciphered. The current study is part of research endeavors aimed at applying systems biology approaches to address the complex biological processes involved in the pathogenesis of vocal fold scarring and other lesions affecting the larynx. METHODS: We developed a computational agent-based model (ABM) to quantitatively characterize multiple cellular and molecular interactions involved in inflammation and healing in vocal fold mucosa after surgical trauma. The ABM was calibrated with empirical data on inflammatory mediators (eg, tumor necrosis factor) and extracellular matrix components (eg, hyaluronan) from published studies on surgical vocal fold injury in the rat population. RESULTS: The simulation results reproduced and predicted trajectories seen in the empirical data from the animals. Moreover, the ABM studies suggested that hyaluronan fragments might be the clinical surrogate of tissue damage, a key variable that in these simulations both is enhanced by and further induces inflammation. CONCLUSIONS: A relatively simple ABM such as the one reported in this study can provide new understanding of laryngeal wound healing and generate working hypotheses for further wet-lab studies.

Translational systems biology and voice pathophysiology.

OBJECTIVES/HYPOTHESIS: Personalized medicine has been called upon to tailor healthcare to an individual's needs. Evidence-based medicine (EBM) has advocated using randomized clinical trials with large populations to evaluate treatment effects. However, due to large variations across patients, the results are likely not to apply to an individual patient. We suggest that a complementary, systems biology approach using computational modeling may help tackle biological complexity in order to improve ultimate patient care. The purpose of the article is: 1) to review the pros and cons of EBM, and 2) to discuss the alternative systems biology method and present its utility in clinical voice research. STUDY DESIGN: Tutorial. METHODS: Literature review and discussion. RESULTS: We propose that translational systems biology can address many of the limitations of EBM pertinent to voice and other health care domains, and thus complement current health research models. In particular, recent work using mathematical modeling suggests that systems biology has the ability to quantify the highly complex biologic processes underlying voice pathophysiology. Recent data support the premise that this approach can be applied specifically in the case of phonotrauma and surgically induced vocal fold trauma, and may have particular power to address personalized medicine. CONCLUSIONS: We propose that evidence around vocal health and disease be expanded beyond a population-based method to consider more fully issues of complexity and systems interactions, especially in implementing personalized medicine in voice care and beyond.

A patient-specific in silico model of inflammation and healing tested in acute vocal fold injury.

The development of personalized medicine is a primary objective of the medical community and increasingly also of funding and registration agencies. Modeling is generally perceived as a key enabling tool to target this goal. Agent-Based Models (ABMs) have previously been used to simulate inflammation at various scales up to the whole-organism level. We extended this approach to the case of a novel, patient-specific ABM that we generated for vocal fold inflammation, with the ultimate goal of identifying individually optimized treatments. ABM simulations reproduced trajectories of inflammatory mediators in laryngeal secretions of individuals subjected to experimental phonotrauma up to 4 hrs post-injury, and predicted the levels of inflammatory mediators 24 hrs post-injury. Subject-specific simulations also predicted different outcomes from behavioral treatment regimens to which subjects had not been exposed. We propose that this translational application of computational modeling could be used to design patient-specific therapies for the larynx, and will serve as a paradigm for future extension to other clinical domains.

Acoustic and perceptual analysis of modal and falsetto registers in females with dysphonia.

Modal and falsetto registers are the basic vocal qualities used in female speaking voices. The purpose of this study was to identify the frequency at which modal register changed to falsetto register and the frequency range of each modal and falsetto register produced under three loudness levels: soft, comfortable and loud phonations in normal and dysphonic speakers. These data provide information on the relationship between vocal registers. It is hypothesized that vocal pathologies affect the frequency at which register change from one mode to another mode. Fifteen dysphonic and 15 non-dysphonic females were assessed on their ability to produce tone series from the lowest to the highest frequency at soft, comfortable and loud phonations. Fourteen listeners served as judges to identify the first production of falsetto voice in each tone series. The first falsetto tone perceived was considered to represent the frequency at which the modal-falsetto register change occurred perceptually. The change from modal to falsetto register in the tone series occurred between G4 (360.74 Hz) and B4 (485.23 Hz) in the dysphonic group and between A4 (421.00 Hz) and B4 (476.13 Hz) in the non-dysphonic group among the three loudness levels. The results showed that the presence of dysphonia affected the production of vocal registers The dysphonic group demonstrated the modal-falsetto register change at a significantly lower frequency and reduced frequency range in the modal register than those of the non-dysphonic group in the soft and comfortable phonations. In producing falsetto register, the dysphonic group demonstrated reduced ability in producing falsetto register in soft and loud phonations when compared to the non-dysphonic group. Physiological explanations are used to account for the observations.