Regenerative treatment for soft tissue defects of the external auditory meatus.

OBJECTIVE: To establish a regenerative treatment for soft tissue defects of the external auditory meatus (EAM) without conventional surgical therapy. STUDY DESIGN: Controlled clinical pilot study. SETTING: General hospitals. PATIENTS: Sixty-five patients with new or old EAM defects without active inflammation were selected. Ages ranged from 12 to 87 years (average age of 58 yr). INTERVENTION: Therapeutic nonsurgical treatment of EAM defects. Gelatin sponge, basic fibroblast growth factor (b-FGF), fibrin glue, and water proof transparent dressing were used in the repair procedure. Patients were divided into 2 groups: treatment with (n = 54) and without (n = 11) b-FGF. After mechanically disrupting the edge of the EAM defect, gelatin sponge immersed in b-FGF was placed over the defect and covered with fibrin glue. In cases of extensive EAM defects, the EAM was filled with gelatin sponge/b-FGF, and the auricle was wrapped in water proof dressing. Two or 3 weeks postprocedure, crust over the defect was removed. If complete defect closure was not achieved after 1 treatment course, the treatment was repeated. MAIN OUTCOME MEASURE: Evaluation of complete closure of EAM defects 3 months posttreatment. RESULTS: Complete closure of the EAM defect was achieved within 3 treatment courses in 92.6% (50/54) and 18.2% (2/11) of the patients with or without b-FGF, respectively. No inflammation/infection or severe sequelae were observed. CONCLUSION: This study demonstrated the effectiveness of combining gelatin sponge, b-FGF, and fibrin glue for EAM defect regeneration. This innovative regenerative therapy is an easy, simple, cost-effective and minimally invasive method for treating EAM defects.

Improvement of eustachian tube function by tissue-engineered regeneration of mastoid air cells.

OBJECTIVES/HYPOTHESIS: Most cases of chronic otitis media (OMC) are associated with poor development of the mastoid air cells (MACs) and poor Eustachian tube (ET) function. We have previously reported that MAC regeneration can effectively eliminate intractable OMC. In this study, we assessed the ability of regenerated MACs to restore normal gas exchange function and contribute to improved ET function. STUDY DESIGN: Clinical trial with control. SETTING: General hospitals. MATERIALS AND METHODS: Seventy-six patients with OMC, including cholesteatoma and adhesive otitis media, received tympanoplasty and MAC regeneration therapy. At the first-stage of tympanoplasty, artificial pneumatic bones and/or autologous bone fragments were implanted into the opened mastoid cavity. At the 2nd-stage operation, a nitrous oxide (N2O) gas study was performed in 10 patients to measure middle ear pressure (MEP). For the control group, MEP was measured in five patients with good MAC development during cochlear implantation or facial nerve decompression. ET function was measured twice in each patient, once before the 1st operation and 6 months after the second operation. RESULTS: At the 2nd-stage operation, in all cases with regenerated MACs and in the normal control group, MEP changed after administration of N2O. In contrast, no change in MEP was observed in cases with unregenerated MACs. In 70% (n = 37/53) of the regenerated MAC group, ET function was improved, whereas improvement of ET function was observed in only 13% (n = 3/23) of the unregenerated MAC group. CONCLUSIONS: Tissue-engineered regeneration of MACs improves ET function and gas exchange in the middle ear.

Regenerative treatment for tympanic membrane perforation.

OBJECTIVE: To establish a tissue engineering therapy for the treatment of large tympanic membrane perforation (TMP) without the need for conventional surgical therapy. STUDY DESIGN: Randomized control trial. SETTING: General hospital. PATIENTS AND METHODS: A total of 63 chronic TMPs were randomly selected from outpatients. INTERVENTION: Of the total 63 chronic TMPs, 53 were randomly assigned to the basic fibroblast growth factor (b-FGF) group and the remaining 10 were randomly assigned to the control group. Materials used for the TM repair were gelatin sponge and fibrin glue with/without b-FGF. After creating a mechanical disruption of the edge of the TMP, a gelatin sponge was immersed in b-FGF or saline (for the control group) and placed over the perforation. Fibrin glue was dripped over the sponge as a sealant. MAIN OUTCOME MEASURES: The effectiveness of this therapy was evaluated by closure rates, hearing level, and sequelae 3 weeks after treatment. The treatment was repeated up to 4 times for cases in which complete closure of the TMP was not achieved after 1 round of treatment. RESULTS: Complete closure of the TMP was achieved in more than 98.1% (52/53) of the patients in the b-FGF group and 10% (1/10) of the patients in the control group. The average hearing level of all patients with successful TM repair was improved. Serious sequelae were not observed in any patient. CONCLUSION: This study demonstrates that a combination of gelatin sponge, b-FGF, and fibrin glue enables the regeneration of the TM without conventional operative procedures. This innovative regenerative therapy is an easy, safe, cost-effective, and minimally invasive outpatient treatment.

Laryngeal regeneration using tissue engineering techniques in a canine model.

OBJECTIVES: We previously reported that polypropylene mesh covered with collagen sponge is a useful material for the regeneration of the trachea and the cricoid cartilage. The aim of this study was to regenerate larynges after partial hemilaryngectomy with this new biomaterial. METHODS: A left partial hemilaryngectomy was performed on 12 adult beagles. The defect size was about 1.8 x 1.0 cm. Both sides of polypropylene mesh were coated with either 1% or 3% collagen sponge. This scaffold was wrapped in fascia lata harvested from the left thigh and then fixed in place over the defect. Endoscopic examinations were performed periodically. Six months after treatment, 3-dimensional computed tomographic scanning was performed. Vibratory examinations were also performed with excised larynges. RESULTS: In the 1% collagen group, exposure or dislocation of the mesh was found in 3 of 6 cases, but in the 3% group, no exposure of the mesh was seen. The morphological findings in the vocal fold were better in the 3% group than in the 1% group, but a difference in the vertical levels of the vocal folds was found in both groups. CONCLUSIONS: This study suggests that 3% collagen-coated polypropylene mesh wrapped with autologous fascia is a useful material for laryngeal regeneration.

A tissue-engineering approach for stenosis of the trachea and/or cricoid.

CONCLUSION: This new regenerative therapy shows great potential for the treatment of stenosis of the trachea and/or cricoids (STC). OBJECTIVES: To estimate the potential of tissue-engineered artificial trachea (AT) for treatment of STC in clinical applications. We previously reported that AT was a useful material for implantation into a tracheal defect after resection of cancer. There are many causes of stenosis of the respiratory tract and STC is particularly difficult to treat. METHODS: The AT was a spiral stent composed of Marlex mesh made of polypropylene and covered with collagen sponge made from porcine skin. Three patients with STC were treated by this tissue-engineering method. All of them suffered from STC caused by long endotracheal intubations. They underwent a two-stage operation. In the first operation, after resection of the stenotic regions, the edge of the tracheal cartilage was sutured to the edge of the skin. The tracheal lumen was exposed and a T-shaped cannula was inserted into the large tracheostoma. At 3 weeks to 2 months after the first operation, the trachea and skin were separated. The trimmed AT with venous blood and basic fibroblast growth factor (b-FGF) was then implanted into the cartilage defect. RESULTS: Postoperatively, all patients were able to breathe easily and had no discomfort in their daily activities. Six months after the second operation, we observed enough air space in the trachea and cricoid by computed tomography (CT) imaging and fiber endoscopy.

Glottal reconstruction with a tissue engineering technique using polypropylene mesh: a canine experiment.

OBJECTIVES: The larynx must be resected in some cases of cancer or stenosis, and various techniques are generally employed to fill the resulting defect. No ideal way, however, has been established to restore vocal function after this form of insult. The aim of this preliminary feasibility study in a canine model was to investigate the effectiveness of a polypropylene-based tissue engineering approach to repair a partial glottal defect. METHODS: Eight dogs were used in this study. A laryngeal defect involving resection of the left vocal fold was created through a thyroid cartilage window. A scaffold made of polypropylene and collagen was preclotted and wrapped with autologous fascia lata, inserted through the window, and sutured to the laryngeal defect in 5 dogs. The defect was reconstructed with an adjacent sternohyoid muscle flap in 3 control dogs. The surgical site was evaluated 3 months after operation by fiberscopic examination, computed tomographic imaging, histologic evaluation, and study of excised larynges. RESULTS: On fiberscopic examination, the experimental group implants were completely covered with regenerated mucosa in all cases, and a favorable vocal fold contour was found in 4 of the 5 cases. One case was characterized by a concave vocal fold shape and red granulation. In the control group, the muscle flap was replaced by scarred mucosa with a concave vocal fold contour in 2 cases, and there was soft white granulation at the anterior resected edge in the third case. The histologic data revealed the regeneration of lined epithelium, subepithelial tissue, and muscle structure in both groups. The excised larynx phonatory data revealed reduced vibratory amplitude in the experimental group compared with the control group; however, excised phonation was not achieved in 2 of the 3 cases in the control group. CONCLUSIONS: This polypropylene-based tissue engineering technique appears to be a viable tool for glottal reconstruction; however, additional refinement is required to maximize long-term phonatory function.

Chronic vocal fold scar restoration with hepatocyte growth factor hydrogel.

OBJECTIVES/HYPOTHESIS: Therapeutic challenges exist in the management of vocal fold scarring. We have previously demonstrated the therapeutic potential of hepatocyte growth factor (HGF) in the management of acute phase vocal fold scarring using a novel hydrogel-based HGF drug delivery system (DDS). However, the effect of HGF on matured vocal fold scarring remains unclear. The current study aims to investigate the effect of HGF-DDS on chronic vocal fold scarring using a canine model. STUDY DESIGN: Animal model. METHODS: Vocal folds from eight beagles were unilaterally scarred by stripping the entire layer of the lamina propria; contralateral vocal folds were kept intact as normal controls. Six months after the procedures, hydrogels (0.5 mL) containing 1 microg of HGF were injected into the scarred vocal folds of four dogs (HGF-treated group). Hydrogels containing saline solution were injected into the other four dogs (sham group). Histological and vibratory examinations on excised larynges were completed for each group 9 months after the initial surgery. RESULTS: Experiments conducted on excised larynges demonstrated significantly better vibrations in the HGF-treated group in terms of mucosal wave amplitude. Although phonation threshold pressure was significantly lower in the HGF-treated group compared with the sham group, no significant differences were observed in the normalized glottal gap between HGF-treated and sham groups. Histological examinations of the HGF-treated vocal folds showed reduced collagen deposition and less tissue contraction with favorable restoration of hyaluronic acid. CONCLUSIONS: Results suggest that administration of HGF may have therapeutic potential in the treatment of chronic vocal fold scarring.

Atelocollagen sponge as a stem cell implantation scaffold for the treatment of scarred vocal folds.

OBJECTIVES: Treatment of vocal fold scarring remains a therapeutic challenge. Our group previously reported the efficacy of treating injured vocal folds by implantation of bone marrow-derived stromal cells containing mesenchymal stem cells. Appropriate scaffolding is necessary for the stem cell implant to achieve optimal results. Terudermis is an atelocollagen sponge derived from calf dermis. It has large pores that permit cellular entry and is degraded in vivo. These characteristics suggest that this material may be a good candidate for use as scaffolding for implantation of cells. The present in vitro study investigated the feasibility of using Terudermis as such a scaffold. METHODS: Bone marrow-derived stromal cells were obtained from GFP (green fluorescent protein) mouse femurs. The cells were seeded into Terudermis and incubated for 5 days. Their survival, proliferation, and expression of extracellular matrix were examined. RESULTS: Bone marrow-derived stromal cells adhered to Terudermis and underwent significant proliferation. Immunohistochemical examination demonstrated that adherent cells were positive for expression of vimentin, desmin, fibronectin, and fspl and negative for beta III tubulin. These findings indicate that these cells were mesodermal cells and attached to the atelocollagen fibers biologically. CONCLUSIONS: The data suggest that Terudermis may have potential as stem cell implantation scaffolding for the treatment of scarred vocal folds.

In situ tissue engineering of canine skull with guided bone regeneration.

CONCLUSION: Calcium alginate (CA) membrane prevents excessive fibrous tissue intrusion and/or dislocation of a bone scaffold. However, CA membrane did not always accelerate cranial bone regeneration. OBJECTIVE: We previously reported skull regeneration using a bone substitute material (BSM), which consisted of collagen-coated beta-tricalcium phosphate and autologous bone fragments, and bone marrow-derived stromal cells (BSCs). However, excessive fibrous tissue intrusion or dislocation of the BSM occasionally interrupted bone regeneration. To avoid such problems, we examined CA membrane, which is useful for guided bone regeneration (GBR), to investigate whether this material maintains the bone regenerative space. MATERIALS AND METHODS: Bone defects (2x2 cm) were created in the skulls of 12 adult beagle dogs using the same clinical procedure. Four experimental models were tested with or without BSM plus BSCs or CA membrane. In group I, the original free bone flap was replaced at the defect. In group II, after replacing the bone flap, the defect was covered with CA membrane. In group III, BSM plus BSCs were used as a gap filler. In group IV, BSM plus BSCs and CA membrane were applied. Histological examinations were performed 3 and 6 months after the operation. RESULTS: In groups I and II, bone regeneration was not observed but fibrous tissue intrusion was prevented in group II. Bone neogenesis was more observed in group III than in group IV at 3 months (p<0.05). At 6 months, the regenerated areas were larger than those observed at 3 months, but the differences between groups III and IV were not statistically significant.

Expression of extracellular matrix proteins in the vocal folds and bone marrow derived stromal cells of rats.

Vocal fold scarring remains a therapeutic challenge. Our research group has indicated that bone marrow-derived stromal cells (BSCs) may have therapeutic potential in restoration of injured vocal folds. However, it is still unclear how BSCs restore the viscoelasticity of vocal fold mucosa. Since a feature of vocal fold scarring is the disorganization of the extracellular matrix (ECM), it is important to understand how BSCs produce ECM. The present study aimed to clarify ECM gene expression in BSCs, and also examined the effects of hepatocyte growth factor (HGF) on this expression. BSCs obtained from the femurs of four Sprague-Dawley rats were cultured with or without HGF. The mRNA expression of ECM components (type I procollagen, decorin, Has2, CD44, MMP-1, and GAPDH) were examined in cultured BSCs and the vocal fold mucosa by the reverse transcription-polymerase chain reaction (RT-PCR). The mRNA expression of Has2 and MMP-1 was significantly stronger in BSCs than in the vocal folds (P < 0.05). Expression of Has2 in BSCs was significantly increased by the administration of HGF (P < 0.05). There was no significant difference in the gene expression of other ECM molecules between BSCs and vocal fold mucosa. Increased expression of Has2 and MMP-1 genes from BSCs may have a positive potential in the treatment of vocal fold scarring.

Drug delivery system of hepatocyte growth factor for the treatment of vocal fold scarring in a canine model.

OBJECTIVES: Vocal fold scarring remains a therapeutic challenge. Previous studies have indicated that hepatocyte growth factor (HGF), a strong antifibrotic element, has therapeutic potential for restoring scarred vocal folds. To enhance the effect of HGF in vivo, we developed a novel drug delivery system (DDS) in which HGF is embedded in gelatin hydrogel and continuously released over a period of 2 weeks. In the present study we investigated the therapeutic efficacy of the HGF DDS on vocal fold scarring by using a canine model. METHODS: The vocal folds of 8 beagles were unilaterally scarred by stripping the entire layer of the lamina propria. The contralateral vocal folds were kept intact as normal controls. One month after the procedure, hydrogels (0.5 mL) containing 1 microg of HGF were injected into the scarred vocal folds of 4 dogs (HGF-treated group), whereas hydrogels containing saline solution were injected in the other 4 dogs (sham group). Histologic and vibratory examinations were completed for each group 6 months after the initial surgery. RESULTS: The excised larynx experiments showed significantly better vibration in terms of mucosal wave amplitude and glottal closure in the HGF-treated group compared to the sham group. Histologic evaluation of the vocal folds indicated remarkable reduction in collagen deposition and tissue contraction, with favorable restoration of hyaluronic acid and elastin in the HGF-treated group. CONCLUSIONS: The present findings suggest that the novel HGF DDS may provide favorable effects in restoring the vibratory properties of scarred vocal folds.

Bone regeneration of canine skull using bone marrow-derived stromal cells and beta-tricalcium phosphate.

OBJECTIVE: The aim of this study was to regenerate high-quality cranial bone using tissue engineering techniques, with subsequent extension to clinical application. Our previous study with a 3-month observation period indicated that a composite scaffold composed primarily of beta-tricalcium phosphate (TCP) had the potential for cranial bone regeneration. In this study, we investigated whether bone marrow derived stromal cells (BSCs) could promote the regeneration of cranial bone as determined after 3 and 6 months. STUDY DESIGN: The pilot study was conducted with 14 adult beagle dogs. MATERIALS AND METHODS: Craniotomy was performed in the same manner used clinically. The bone defect (2 x 2 cm) was created at each canine temporoparietal region. The test animals were divided into three groups. In group I, the bone defect was closed by replacing the original free bone flap without filling the residual gaps. In group II, the gap was filled with a composite scaffold consisting of collagen coated beta-TCP and autologous bone fragments with fibrin glue. In group III, autologous cultured BSCs and the composite scaffold were used to fill the gap. The sites of craniotomy were analyzed with three-dimensional computed tomography and histologic examination 3 and 6 months after the operation. RESULTS: Bone regeneration was observed in groups II and III, with more extensive formation in group III than in group II. In group I, bone regeneration was not observed. CONCLUSION: This study showed that BSCs have the potential to promote cranial bone regeneration and confirmed the efficacy of a composite scaffold made of beta-TCP and autologous bone fragments with fibrin glue.

Experimental regeneration of canine larynx: a trial with tissue engineering techniques.

CONCLUSION: Since this tissue engineering technique is cost-effective and is less invasive to patients, it may replace conventional approaches in laryngeal reconstructive surgeries. OBJECTIVE: Laryngeal cancer is one of the most prevalent cancers in the head and neck region, and frequently requires surgical resection. Although there are many ways to reconstruct the larynx after resection, donor tissue is usually required. Recently, tissue engineering techniques have become widely accepted in clinical medicine and have already been applied to some organs. This animal experiment was designed to elucidate the efficacy of laryngeal regeneration using tissue engineering technique. MATERIALS AND METHODS: A bioartificial scaffold was designed from a replica of a canine larynx. A dental cast was used to replicate the intricate inside shape of the larynx. After copying its shape on a polypropylene mesh sheet, this sheet was coated with spongy collagen from porcine skin. A hemilaryngectomy was performed on beagle dogs under general anesthesia. Then the scaffold, preclotted with a mixture of peripheral blood and bone marrow-derived stromal cells, was implanted and fixed. The postoperative status was examined fiberscopically. RESULTS: On the eighth day after the operation, the surface of the implant was covered with soft tissue. Finally, the implant was completely covered with regenerated mucosa.

A case of floppy epiglottis in adult: a simple surgical remedy.

A new operative technique for the treatment of floppy epiglottis is introduced in this report. Though its diagnosis is not so difficult by close observations using a flexible fiberscope, it is likely to be treated as an asthma attack or a paradoxical vocal cord movement. In previous reports of upper airway obstruction caused by adult floppy epiglottis: epiglottis prolapse during inspiration, the cause of this condition has been divided broadly into two categories: idiopathic cases and secondary cases. Partial or total epiglottectomy has been usually reported for the treatment of the relief of airway obstruction caused by a floppy epiglottis. It is very difficult to decide adequately how much volume of epiglottis to cut without postoperative sequelae. A V-shaped partial epiglottidectomy is described for its treatment. This technique is very easy, simple, and effective.

[Neurinoma of tympanic plexus: a case report].

We report a unique case of neurinoma originating from the tympanic plexus. A 23-year-old man reparting hearing impairment was found in otoscopic examination to have a tumor contour identified through the ear drum. Computed tomography showed that the tumor extended from the hypotympanum to the mesotympanum, eroding the promontory. A small specimen obtained after myringotomy indicated neurinoma. The man did not report facial weakness or taste disturbance. When we operated to remove the tumor, we found the long crus of Incus had disappeared and the stapes superstructure was dislocated upward due to the tumor expanse. Pathologically, the extirpated tumor was neurinoma of Antoni A. A postoperative salivary gland function test using Tc showed hypofunction at the lesion side of the parotid gland. We surmise that neurinoma originated from the tympanic plexus because of its location and its lack of a relationship to the chorda tympanic nerve or facial nerve.