Vocal Fold Healing Research Articles

Surgical alternatives to classical phonomicrosurgery for vocal fold cysts: our experience.

Treating options for vocal fold cysts became more diversified with office-based laryngological surgery advances, offering new soft interventions, promising a good vocal outcome within a shorter healing period. We proposed two types of in-office surgeries, performed in the outpatient setting: a cyst punction-aspiration emptying the cyst, relieving its tension and improving the mucosal wave, with 4 cases showing no refilling within one year of follow-up, and a cyst marsupialization with Blue Laser, with 3 reported cases showing good vocal fold healing within three months. Both techniques showed promising results in improving vocal function and minimizing recovery time.

The role of systemic dehydration in vocal fold healing: Preliminary findings.

Systemic dehydration negatively alters the expression of vocal fold inflammatory and cell junction markers. These biological changes can have downstream effects on the healing processes of injured vocal folds. In the dermis, reduced hydration prolongs inflammation and delays healing. It is unknown whether this biological effect is observed in vocal fold tissue. To investigate the effects of systemic dehydration on vocal fold healing outcomes following acute, bilateral vocal fold injury in a rodent model. Eighteen systemic dehydrated and 18 euhydrated adult male Sprague Dawley rats experienced bilateral vocal fold injuries or no injury (N= 9/group). Vocal fold gene expression levels of inflammatory mediators and epithelial cell junction markers were measured 24 h post-injury. Pro-inflammatory gene markers (IL-1β; TNF-α) were differentially expressed in response to systemic dehydration with vocal fold injury compared to non-injury. Epithelial cell junction markers (Cadherin-3, Desmoglein-1) also exhibited divergent trends following systemic dehydration, but these data were not statistically significant. Systemic dehydration may affect cellular vocal fold healing processes within 24 h. These findings lay the groundwork for further investigation of how hydration status can affect vocal fold tissue recovery and influence clinical care.

Cryotherapy Modifies Extracellular Matrix Expression of Vocal Fold in Rat Models

Vocal fold (VF) scarring is the major cause of voice disorders. Cryotherapy is an effective anti-scarring therapy for skin lesions. The aim of this study was to explore the anti-scarring potential of cryotherapy in vocal folds. The extracellular matrix (ECM) mRNA expression of cryotherapy on normal VF tissue and the histologic results of cryotherapy on vocal fold healing were studied. Fifteen rats were introduced cryotherapy on the normal VF bilaterally and were harvested for real-time polymerase chain reaction (RT-PCR) analysis for collagen I, collagen III, TGFβ1, decorin, fibronectin and HAS1 at 1 day, 3 days and 7 days. Ten rats were unilaterally injured by stripping lamina propria and immediately treated with or without cryotherapy and were harvested at 2 months for histological and immunohistochemical analysis. Regenerative effect of cryotherapy was validated of ECM gene expression. Histological and immunohistochemical analysis showed significantly increased hyaluronan, decreased collagen, and increased decorin deposition in injury-cryotherapy cohort compared with injury control cohort and normal control cohort. Cryotherapy may provide an optimal environment for vocal fold tissue regeneration. The results of the present investigation suggest that cryotherapy has therapeutic potential in prevention and treatment of vocal fold scarring.

Effect of application of fibrin glue on vocal fold healing after surgical manipulation in rabbits

IntroductionA challenge in phonosurgery is achieving good vocal quality with minimal vocal fold fibrosis. Fibrin glue can be applied to minimize fibrosis; however, its use in the larynx is based primarily on clinical experience, particularly in extensive lesions. ObjectiveThe objective of this study was to evaluate the effect of application of fibrin glue on collagen concentration at the late phase of the healing process after detaching a pedicled flap of the vocal fold cover in rabbits. MethodsIn this prospective animal study, twelve adult male rabbits underwent laryngeal microsurgery, in which an incision was made along the entire length of both vocal folds, followed by extensive mucosal detachment and section of the incision ends. Fibrin glue was applied in the left vocal fold, and the mucosa was repositioned. In the right vocal fold, the mucosa was repositioned without treatment with fibrin glue. After 3-months, the rabbits were euthanized. Histological analyses were performed, and the data collected were subjected to statistical analysis. ResultsVocal folds treated with fibrin glue presented higher collagen concentration in Masson trichrome staining and significantly higher (p < 0.05) collagen concentration in picrosirius red staining compared to control vocal folds. ConclusionTreatment with fibrin glue led to greater vocal fold fibrogenesis in the present study. Nonetheless, further studies are necessary to determine the prophylactic effect of sealants in laryngeal surgeries that require extensive detachment of the vocal fold cover.

Laryngeal and Pharyngeal Movements During Inner Singing: A Cross-Sectional Study

Laryngeal and pharyngeal activity during inner singing is discussed in the context of vocal hygiene. Inner singing is defined as imagined singing, reading music silently, and listening to vocal music. When vocal rest is prescribed, doctors, speech therapists, and voice pedagogues recommend avoiding listening to music or reading music silently, since it is suggested that inner singing unconsciously influences the glottis, and thus moves the vocal folds involuntarily. The aim of this study was to compare the degree to which involuntary laryngeal and/or pharyngeal activity occur during inner singing, inner speech, and at rest, and to evaluate if current recommendations concerning vocal hygiene are still reasonable. Thirty vocally healthy participants were examined transnasally with a flexible videoendoscope. The sample consisted of 10 nonsingers, 10 lay singers, and 10 professional singers. Participants were examined during five tasks including rest, silent reading, imagining a melody, listening to music, and reading music. Two medical doctors specializing in phoniatrics analyzed the videos both qualitatively and quantitatively. During the endoscopic examination, the raters identified movements at the base of the tongue, the posterior and lateral pharynx wall, the arytenoid cartilage, and the vocal folds. The inner singing tasks showed significantly more laryngeal movements as well as significantly more glottal closures than the control tasks (at rest, silent reading). Pharyngeal structures did not show an increase in activity during inner singing. These findings were independent of the level of proficiency in singing. When total vocal rest is prescribed, patients should also be advised to avoid music imagination. Still, further research is needed to survey in detail the actual effects of these involuntary movements during inner singing on the regeneration process of vocal fold healing.

The effects of cryotherapy on vocal fold healing in a rabbit model.

Cryotherapy has been shown to be a scarless treatment modality for dermal lesions; however, there are limited data addressing the effect of cryotherapy on vocal fold tissue. The aim of this study was to clarify the effectiveness of cryotherapy for prevention of postsurgical vocal fold scarring. Prospective animal study in rabbits. The lamina propria of 20 rabbit vocal folds was bilaterally stripped, followed by randomized unilateral cryotherapy. Five larynges were harvested for real-time polymerase chain reaction (RT-PCR) analysis at 1 day, 3 days, and 7 days postinjury. The remaining five were harvested for histologic analysis at 3 months. Images of the healing phase were recorded by laryngoscopy. Analyses of RT-PCR for cyclooxygenase (COX)-2, interleukin (IL)-6, collagen I, collagen III, matrix metallopeptidase 1 (MMP1), transforming growth factor β (TGFβ1), α smooth muscle actin (α-SMA), and hyaluronan synthase 1 (HAS1) were completed. Histological samples were completed for collagen and hyaluronic acid analysis. RT-PCR results revealed that higher expressions of HAS1 and MMP1 and lower expressions of COX-2, IL-6, collagen I, collagen III, TGFβ1, and α-SMA were observed, and histological examination showed significantly increased hyaluronic acid, decreased deposition, and more organized configuration of collagen in injury with the cryotherapy cohort compared with the injury cohort. Cryotherapy can inhibit the inflammatory reaction and simulate a fetal healing environment in extracellular matrix synthesis to regenerate vocal fold tissue with less fibrosis. Histological results showed that cryotherapy achieves a mature healing result with less scar, which tends to return to normal. In summary, the findings of this study suggest that administration of cryotherapy at the time of injury has the potential to minimize vocal fold scarring. NA Laryngoscope, 129:E151-E157, 2019.

The effects of cytosporone-B, a novel antifibrotic agent, on vocal fold fibroblasts.

Our laboratory recently described NR4A1 as an endogenous inhibitor of TGF-β-induced vocal fold (VF) fibrosis. Our prior report described the temporal expression of NR4A1 during VF healing in vivo and the effects of NR4A1 knockdown on fibroplastic cell activities in vitro. Based on these findings, we hypothesized that cytosporone-B (Csn-B), an NR4A1 agonist, may hold significant therapeutic potential. In vitro. Human VF fibroblasts were exposed to TGF-β1+/-Csn-B. Expression of genes related to fibrosis were quantified. In addition, contraction was assayed as a surrogate for the fibrotic phenotype in our cell line. TGF-B1 stimulated COL1A1 and ACTA2, as expected. Csn-B significantly downregulated TGF-β1-mediated upregulation of these genes (P = .009, P = .03, respectively). Csn-B had no effect on genes related to TGF-β/Smad signaling. Csn-B also decreased the TGF-β1-mediated contractile phenotype in our cells (P = .004). NR4A1 is an endogenous inhibitor of fibrosis in the vocal folds and Csn-B, as an NR4A1 agonist, may evolve as an ideal, therapeutic candidate for this challenging condition. NA Laryngoscope, 128:E425-E428, 2018.

Cryotherapy has antifibrotic and regenerative effects on human vocal fold fibroblasts.

Vocal fold scarring remains a major treatment challenge, and scar prevention without residual lesions remains a dilemma. Cryotherapy has shown cosmetic outcomes on skin lesions with minimal scarring. The aim of this study was to clarify the beneficial effects of cryotherapy for the prevention and the treatment of vocal fold scarring. In vitro. Primary cultures of human vocal fold fibroblasts (VFFs) were used in this study. Myofibroblast differentiation was stimulated by transforming growth factor β1 (TGF-β1). We mimicked the cryotherapy effect on vocal fold healing in vivo by freezing VFFs ± TGF-β1 in vitro. The influence of freezing on cell viability, proliferation, migration, and contractile properties were analyzed. The expression of collagen I, collagen III, fibronectin, TGF-β1, matrix metallopeptidase 1 (MMP1), hyaluronan synthase 1 (HAS1) were investigated by real-time polymerase chain reaction (RT-PCR), and the expression of alpha smooth muscle actin (α-SMA) and decorin were investigated by RT-PCR and Western blot. Freezing was found to modify extracellular matrix (ECM) synthesis and differentiation of VFFs. Expression of collagen I, collagen III, fibronectin, α-SMA, and TGF-β1 was downregulated, and MMP1 was upregulated in VFFs + TGF-β1 (myofibroblast) by freezing. HAS1 and decorin were upregulated in both VFFs ± TGF-β1 by freezing. Freezing VFFs + TGF-β1 (myofibroblast) with fast thawing had a lower expression of α-SMA when compared with slow thawing. Freezing reduced the migration and collagen contraction of VFFs + TGF-β1 (myofibroblast). Cryotherapy induces antifibrotic and regenerative ECM alterations in VFFs. These data provide insight into the prevention and the treatment of vocal fold scarring with cryotherapy in phonomicrosurgery. NA Laryngoscope, 129:E143-E150, 2019.

Adipose stromal cells improve healing of vocal fold scar: Morphological and functional evidences.

Adipose derived stromal cells (ASCs) are abundant and easy to prepare. Such cells may be useful for treating severe vocal disturbance caused by acute vocal fold scars. Prospective animal experiments with controls. Twenty New-Zealand white rabbits were used in the present study. We evaluated vocal fold healing, with or without injection of autologous ASCs, after acute scarring. A defined lesion was created and the ASCs were immediately injected. Vocal fold regeneration was evaluated histomorphometrically and via viscoelastic analysis using an electrodynamic shaker. Six weeks after ASC injection, vocal folds exhibited significantly less inflammation than control folds (P < 0.005). In addition, hypertrophy of the lamina propria and fibrosis were significantly reduced upon ASC injection (P < 0.02). The decrease in viscoelastic parameters was less important in the ASC injected group compared to the noninjected group (P = 0.08). Injection of autologous ASCs improved vocal fold healing in our preclinical model. Further studies are needed, but this method may be useful in humans. NA. Laryngoscope, 126:E278-E285, 2016.

Homeostasis of Hyaluronic Acid in Normal and Scarred Vocal Folds

Vocal fold scarring is one of the most challenging laryngeal disorders to treat. Hyaluronic acid (HA) is the main component of lamina propria, and it plays an important role in proper vocal fold vibration and is also thought to be important in fetal wound healing without scarring. Although several animal models of vocal fold scarring have been reported, little is known about the way in which HA is maintained in vocal folds. The purpose of this study was to clarify the homeostasis of HA by examining the expression of hyaluronan synthase (Has) and hyaluronidase (Hyal), which produce and digest HA, respectively. Experimental prospective animal study. Vocal fold stripping was performed on 38 Sprague-Dawley rats. Vocal fold tissue was collected at five time points (3days-2months). Expression of HA was examined by immunohistochemistry, and messenger RNA (mRNA) expression of Has and Hyal was examined by real-time polymerase chain reaction and in-situ hybridization. In scarred vocal folds, expression of Has1 and Has2 increased at day 3 together with expression of HA and returned to normal at 2weeks. At 2months, Has3 and Hyal3 mRNA showed higher expressions than normal. Expression patterns of Has and Hyal genes differed between normal, acute-scarred, and chronic-scarred vocal folds, indicating the distinct roles of each enzyme in maintaining HA. Continuous upregulation of Has genes in the acute phase may be necessary to achieve scarless healing of vocal folds.

A reporter assay for the next generation of biomaterials: Porous‐wall hollow glass microspheres

The primary objective was to design a reporter assay to measure molecular release kinetics from a new porous-wall hollow glass microsphere biomaterial with great potential in regenerative medicine and drug delivery. Second, future avenues for research will be discussed specifically in regard to potential clinical uses in laryngology. Basic science data report. We developed an assay using fluorescent nanocrystals or quantum dots (Qdot 605) as a reporter. A Nuance FX multispectral imaging system was used to detect fluorescence in aqueous phase. Spectral output of known concentrations of aqueous Qdot 605 was measured by the Nuance system to create a standard curve. These data were plotted and fit to a curve. Qdot 605 emission demonstrates excellent correlation with concentration in a log-log relationship [R(2) = 0.99649, median error = 9.9%], indicating that the Qdot 605 assay is reliable and should be explored regarding its ability to evaluate the drug-eluting properties of this material. We have derived a method to measure Qdot concentration using fluorescent microscopy, which will facilitate future research on this exciting new biomaterial. This material has great potential for use in head and neck surgery. Specific avenues within laryngology to be investigated include laryngeal and tracheal reconstruction, vocal fold healing, and nerve regeneration. Furthermore, we believe this is the first documented use of the Nuance system to determine aqueous molecular concentrations. NA.

Characterization of extracellular matrix proteins during wound healing in the lamina propria of vocal fold in a canine model: A long-term and consecutive study

The characterization of vocal fold wound healing can be reflected by the changes of extracellular matrix (ECM) proteins in the lamina propria. Although the expression of ECM proteins after vocal fold injury has been widely studied, such observations have lacked time continuity and integrity of marker proteins. In this study, we observed the morphology of injured vocal folds in a canine model. We used immunofluorescence staining to evaluate the expression and distribution of ECM proteins, such as collagen, elastin, hyaluronic acid, decorin and fibronectin, from 15 days to 6 months after injury. The results showed that large amounts of ECM proteins were secreted 15–40 days after injury. Collagen and fibronectin secretion increased significantly, and were disorderly deposited. The secretion of decorin and elastin increased slightly, while hyaluronic acid decreased. The 15–40 day post-injury period may be the critical intervention stage in wound healing of vocal folds. From 3 to 6 months after injury, the secretion of ECM proteins declined. However, collagen and fibronectin secretion were still significantly higher than normal with irregular arrangement, while the secretion of elastin, hyaluronic acid and decorin decreased significantly at 6 months. This led to vocal fold inelasticity and stiffness, which required effective long-term interventions to treat scar formation.

Age effects on extracellular matrix production of vocal fold scar fibroblasts in rats

Vocal fold (VF) fibroblasts are the central subject of interest in fibrogenesis and wound healing after VF injury. Scar fibroblasts (SF) exhibit an aberrant production of several extracellular matrix (ECM) components which lead either to VF fibrosis or scarless wound healing. This study aimed to investigate the role of age at the time of injury on ECM production of SF. This is designed as an animal study. VF injury was established unilaterally in eight male Sprague-Dawley rats [3 months of age (n = 4), 11 months of age (n = 4)], while the other side was left intact. Three months after injury the larynges were excised and fibroblasts were extracted from VF [normal fibroblasts (NF)–scar fibroblasts (SF)] and cultured in vitro. After first passage, VF fibroblasts were plated in 24-well plates and levels of hyaluronic acid (HA) and collagen type I were determined enzymatically from supernatant after 24 and 72 h. Cultured SF from younger animals produced significantly higher levels of HA compared to NF fibroblasts from the same animals. HA concentrations of the older animals did not differ significantly between the NF and SF cultures, but the range in SF cultures was large. In contrast to previous studies, we found that even 3 months after VF injury cultured SF from young animals expressed higher levels of HA in comparison to SF from older animals. No difference in collagen levels were observed between the younger and older animals. Age of animals is an essential factor during VF healing and has to be considered for study design.

Histologic comparison of vocal fold microflap healing with sutures and glue

Improved voice quality is the expected outcome of microphonosurgery. To this end, the complex vibratory movement of the vocal folds must be preserved. Scarring of the vocal folds may compromise vocal outcome and is one of the most difficult to treat conditions. To minimize scar formation, a consensus exists on the need for maximum preservation of the epithelium and superficial lamina propria, and minimal exposure of vocal ligament. However, the need to cover the microflap defect is controversial. The aim of this study was to compare healing characteristics of microflap technique when the microflap is left to heal by second intention, when the defect is closed with sutures, and when it is covered with glue. Experimental animal study. An experimental study comparing the three techniques was carried out on 37 New Zealand rabbits. Vocal fold healing was evaluated 7, 30, and 90 days after surgical intervention. Collagen concentration, inflammatory reaction, epithelium, and lamina propria thickness were evaluated. Collagen concentration significantly increased in all groups. After 7 days, epithelium thickness significantly increased and lamina propria thickness was not significantly altered. At 90 days, only the suture group showed no significant alteration in epithelium thickness. After 90 days, lamina propria thickness decreased, except in the fibrin glue group. No difference was seen in the number of inflammatory cells among the techniques. The use of microsutures or fibrin glue to close microflap defects did not consistently improve vocal fold healing nor produce better scarring results when compared to healing by second intention.

Comparison of Subjective and Objective Tools in Transoral Laser Cordectomy for Early Glottic Cancer: Importance of Voice Handicap Index

Several studies in recent years have already reported good oncologic results with laser microsurgery in the treatment of early glottic carcinoma. We conducted a longitudinal voice evaluation, in patients with early glottic cancer who underwent transoral laser cordectomy, in order to assess the voice quality outcome and its relationship with objective and subjective (voice handicap index questionnaire and GIRBAS scale) means. Twenty-four previously untreated patients underwent transoral laser cordectomy for early glottic cancer. All patients underwent subjective and objective voice assessment according to the multidimensional voice protocol recommended by the European Laryngological Society including acoustic, perceptual and stroboscopic analysis combined to patient self assessment of voice (voice handicap index-VHI). These evaluations were performed before treatment and 6 months after the treatment. Vocal fold healing was complete in all cases by 6 months following surgery. Main voice parameters (subjective and objective) resulted improved at the 6-month control after surgery. In particular, voice handicap index (VHI) and GIRBAS scale resulted reliable for voice assessment and for the postoperative follow-up. In conclusion, VHI, GIRBAS, multidimensional voice program (MDVP) and spectroacoustic parameters showed a close trend in the present study, and this confirms the validity of the voice analysis performed by each tool.

Expression of Reactive Oxygen Species during Wound Healing of Vocal Folds in a Rat Model

Previous studies have indicated that although normal wound healing requires low levels of reactive oxygen species (ROS), excessive amounts of ROS impair wound healing. In injured vocal folds, this excess may result in dysphonia due to scarring that is difficult to treat. However, the expression of ROS during vocal fold wound healing has yet to be investigated. In this study, we assessed the expression and localization of ROS in injured vocal folds by immunohistochemical analysis. Vocal folds of Sprague-Dawley rats were unilaterally injured by stripping the mucosa under transoral endoscopy. The larynges were harvested at specific time points after injury and were immunohistochemically examined for 4-hydroxy-2-nonenal (4-HNE), an ROS marker, and for the presence of inflammatory cells. We found that 4-HNE-immunopositive cells were significantly increased in the lamina propria of the injured vocal folds as compared to the normal vocal folds on postinjury days 1 and 3. More than half of the 4-HNE-immunopositive cells were also immunopositive for a macrophage- and granulocyte-specific antibody. This study suggests that a large amount of ROS is produced during early-phase wound healing, until postinjury day 3, and that this period may be crucial for regulating ROS levels. The results also suggest that inflammatory cells may contribute to ROS generation.

Developing a Porcine Model for Study of Vocal Fold Scar

The porcine larynx is very similar in size and structure to that of humans, and wound healing in pigs is very similar to that of humans. However, the pig is not often used in vocal fold scar research because it is difficult to view the vocal folds endoscopically. To further assess the pig as a model for studying vocal scar, we compared the plane of surgical dissection in the mucosa of four porcine vocal folds with that in eight human cadaver larynges. The plane of dissection was quite similar in porcine and human larynges, occurring within the loose layer of the superficial lamina propria. We also compared healing of porcine vocal folds after elevation and replacement of an epithelial flap versus excision of epithelium, leaving an open wound. After 6 weeks, larynges were harvested for histologic examination. There was no significant difference between the mucosa of the normal vocal fold and that of the healed microflap. However, after healing of epithelial excision, there was a depressed scar, with average lamina propria thickness of 302 μm versus 864 μm for the normal fold (P<0.05). Finally, to document that the mucosal wave can be evaluated in the porcine larynx, we developed a preparation that removes the false vocal folds, to allow ex vivo phonation. Experimentally created scar in the porcine larynx is a favorable model for the study of vocal fold healing and for assessment of treatments for vocal fold scar.

Influence of selant fibrin on the wound healing of the pigs vocal folds

Fibrin sealants or fibrin glue are products made from human plasma proteins, which mimic the final pathway of the coagulation cascade. Its application to stimulate the healing process has been a topic of debate in the literature. The use of fibrin sealants in phonosurgery has been empirical; there have been no studies that investigate the action of fibrin sealant in Reinke's space. AimTo evaluate the effect of fibrin glue in healing of the vocal folds of pigs after surgical manipulation. Materials and MethodsThis was a prospective and experimental study. Six animals had both vocal folds incised. Sealant was applied in one of them; the other served as a control. After three months, the animals were sacrificed and a collagen count was carried out. ResultsThe side on which glue was applied had an average of 27.8% against 20.4% of the side without glue. ConclusionThe collagen concentration in the samples where the fibrin sealant was applied was significantly higher compared to samples without glue. Thus, the presence of a fibrin sealant stimulates fibrogenesis in this tissue.

Biosimulation of acute phonotrauma: An extended model

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. Computer simulation. 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)-1β, 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. 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. 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.

Injection of human mesenchymal stem cells improves healing of vocal folds after scar excision—A xenograft analysis

Using a xenograft model the aim was to analyze if injection of human mesenchymal stem cells (hMSC) into the rabbit vocal fold (VF), after excision of an established scar, can improve the functional healing of the VF. Prospective design with an experimental xenograft model. The VFs of 12 New Zealand rabbits were injured by a bilateral localized resection. After 9 weeks the scar after the resection was excised and hMSC were injected into the VFs. After another 10 weeks 10 VFs were dissected and stained for histology. Lamina propria thickness and relative content of collagen type I were measured. Viscoelasticity of 14 VFs at phonatory frequencies was quantified by a simple-shear rheometer. The hMSC survival was determined using a human DNA specific reference probe, that is, FISH analysis. The viscoelastic measurements, that is, dynamic viscosity and elastic shear modulus for the hMSC-treated VFs, were found to be similar to those of normal controls and were significantly lower than those of untreated controls (P < .05). A significant reduction in lamina propria thickness was also shown for the hMSC treated VFs compared with the untreated VFs (P < .05). This histologic finding corresponded with the viscoelastic results. No hMSC survived 10 weeks after the injection. Human mesenchymal stem cells injected into the rabbit VF following the excision of a chronic scar, were found to enhance the functional healing of the VF with reduced lamina propria thickness and restored viscoelastic shear properties.