Effects of halofuginone on fibrosis formation secondary to experimentally induced subglottic trauma

OBJECTIVE: The aim of the study was to assess the antifibrotic effect of systemically applied halofuginone after subglottic injury. MATERIALS AND METHODS: After standardized trauma to subglottic area, rats were divided into two groups: a study group that received treatment and a control group that did not. The rats were treated with 0.1 mg/kg/day intraperitoneal halofuginone injection for 30 days. The larynx specimens were examined histopathologically under light microscope for epithelization, inflammation, necrosis, and fibrosis. RESULTS: The fibrosis indexes of the treated group were significantly less than those of the control group (P < .01). CONCLUSIONS: Systemically applied halofuginone hydrobromide decreases fibrosis/scar tissue formation secondary to experimentally induced acute subglottic trauma.     

Differential effects of endolaryngeal trauma upon the growth of the subglottis

In young rabbits (4-week-old) the growth of the subglottis was studied after an endolaryngeal soft tissue trauma. The injury induced the formation of a subglottic stenosis. In contrast to earlier observations on endolaryngeal trauma extending into the cricoid cartilage, the cricoid developed normally to the adult size and shape at 24 weeks. Similar to the earlier findings, the subepithelial layer showed considerable thickening as the result of formation of scar tissue, ectopic cartilage and fatty tissue with interruption of the elastic tunica (= conus elasticus). It was concluded that after endolaryngeal trauma in rabbits two types of subglottic stenosis can develop, determined by the depth of the injury.     

Characterization of submucosal lesions using optical coherence tomography in the rabbit subglottis

OBJECTIVE: To evaluate the efficacy of optical coherence tomography in differentiating between several simulated subglottic lesions, using an ex vivo rabbit laryngotracheal model. DESIGN: Laryngotracheal complexes were harvested from euthanized rabbits and divided into the following 4 groups: (1) control, (2) submucosal collagen injection (simulating scar formation), (3) dehydration and rehydration (simulating edema), and (4) repeated intubation trauma. The subglottic region was imaged using optical coherence tomography. Images were later correlated with conventional histologic findings. RESULTS: The epithelium, basement membrane, lamina propria, perichondrium, and cartilage (cricoid and tracheal) were clearly imaged. In group 2, an increase in the thickness of the lamina propria was observed, in addition to a characteristic optical pattern of the injected collagen. Dehydration (in group 3) produced a visible reduction in the thickness of the lamina propria, while rehydration of the same specimen with distilled water revealed a significant increase in submucosal swelling. Repeated intubation (in group 4) resulted in tissue edema that was seen as wavy heterogeneous thickening of the lamina propria. Edema produced by repeated intubation or distilled water immersion was easily differentiated from native and collagen-injected tissues. CONCLUSION: Optical coherence tomography successfully identifies the microstructure layers of the subglottis and can differentiate between edema and increased collagen deposition in the rabbit model.     

A comparison of the efficacy of 5-fluorouracil/triamcinolone, carnitine and dexamethasone therapy on wound healing in tracheal injury: potential for preventing tracheal stenosis?

The aim of the study was to investigate the efficacy of 5-flourouracil/triamcinolone acetonide (5-FU/TA), carnitine and dexamethasone on wound healing in an animal model of tracheal injury. Twenty-eight rats underwent surgical injury of the tracheal mucosa and perichondrium under general anesthesia, and were randomized into four groups. Group I (the control group) received nothing after tracheal trauma. Group II received intratracheal 5 mg 5–FU/TA combined with 0.2 ml sodium hyaluronic acid once. Group III received intraperitoneal carnitine 100 mg/kg/day for 20 days. Group IV received intramuscular dexamethasone 0.1 mg/day for 20 days. After 1 month, the levels of superoxide dismutase enzyme activity (SOD) and levels of malondialdehide (MDA) and nitric oxide (NO) in serum were measured and the tracheal specimens were harvested for histopathologic examination. SOD was significantly lower in the carnitine group compared to the control group. A statistically significant decrease in MDA levels were observed in the 5-FU/TA and carnitine groups compared to the control group. NO levels were significantly lower in the 5-FU/TA group, but significantly higher in the dexamethasone group, compared to the control group. The fibrotic tissue volumes of tracheal specimens were significantly lower in both the 5-FU/TA and carnitine groups than those of the control group, but trended toward higher volumes in the dexamethasone group when compared to controls. We concluded that 5-FU/TA and carnitine diminish the occurrence of tracheal stenosis (TS) secondary to experimentally induced tracheal trauma.     

Role of vascular endothelial growth factor A in children with acquired airway stenosis

OBJECTIVES: Vascular endothelial growth factor A (VEGF-A) is important in the angiogenic response for wound healing. This study investigated whether VEGF-A may play a role in the pathogenesis of acquired airway stenosis. METHODS: Eight lesions from 5 pediatric patients with subglottic stenosis after airway reconstruction (N = 4) or prolonged intubation (N = 1) and normal laryngeal tissue from 5 autopsy patients were included. Formalin-fixed sections of subglottic tissue from each patient were examined by in situ hybridization for the presence of messenger RNA (mRNA) for VEGF-A, vascular endothelial growth factor receptor 1 (VEGFR-1), and vascular endothelial growth factor receptor 2 (VEGFR-2). RESULTS: Strong expression of VEGF-A mRNA was noted in hyperplastic squamous epithelium overlying granulation tissue. Strong expression of VEGFR-1 and VEGFR-2 was noted in the endothelial cells within granulation tissue. No strong labeling of VEGF-A mRNA or its receptors was noted in 2 specimens with mature scar tissue or in the control specimens. CONCLUSIONS: The angiogenic growth factor VEGF-A is strongly expressed in hyperplastic epithelium overlying granulation tissue in airway stenosis. Also, VEGFR-1 and VEGFR-2 mRNAs are strongly expressed in the endothelial cells of granulation tissue. This finding suggests an important role of VEGF-A in the pathogenesis of airway scar formation and stenosis.     

Can Mitomycin C Really Prevent Airway Stenosis?

BACKGROUND/OBJECTIVES: Mitomycin C (MMC) is used in the prevention of upper airway stenosis. However, the efficacy of MMC is still debatable, and the potential complications of MMC use have been scarcely reported. We evaluated the efficacy and early complications of MMC in a wound model of the subglottis. STUDY DESIGN: A randomized animal study. METHODS: Sixty rabbits underwent a full-thickness injury to the posterior subglottis by diode laser. The animals were randomly assigned to three groups according to the different topical treatments of MMC 0.4 or 10 mg/mL or saline alone for 5 minutes. Animals were killed at 4 weeks, and gross and histologic findings were compared among different groups and 10 age-matched, non-wound, normal controls. RESULTS: Thirty-two of the 60 (53%) animals died mainly of acute airway obstruction by necrotic debris, sloughs on, or cartilage collapse of the unhealed posterior subglottis during early weeks after wounding, higher in the MMC-treated animals (67%) than in the wound controls (25%) (P = .007). This resulted from the significant delay of wound healing in the MMC-treated groups compared with the wound controls (P = .012). The degree of subglottic stenosis was comparable among different groups (P > .8), although collagen deposition was significantly lower in the MMC-treated groups compared with the untreated controls (P = .036). CONCLUSIONS: There was a significant risk of acute airway obstruction from delayed wound healing in rabbits that received subglottic laser wounding and MMC treatment. This risk should be considered in the use of MMC in clinical practice.     

A histologic study of the growth of the subglottis after endolaryngeal trauma

In young rabbits the histologic features of the growth of the subglottis were studied after an endolaryngeal trauma. This lesion of the larynx leading to a subglottic stenosis, caused specific pathologic changes of the cricoid cartilage and the subepithelial tissues. Activation of chondrocytes resulted in proliferation and regeneration of destructed cartilage. The normal subepithelial structures were replaced by granulation tissue differentiating into a thick fibrous scar and into cartilage. This ectopic cartilage together with the repaired cricoid formed a composite thickened 'ring'-like cartilaginous structure in the adult stage.     

Histologic study of the growth of the subglottis after interruption of the circular structure of the cricoid

In a previous study in growing rabbits it was demonstrated that after resection of a ventral part of the cricoid the lumen of the subglottic airway developed normal size during further growth up to the adult stage. Histologic investigations of these cases suggest that a subepithelial network of elastic fibres, ventrally suspended to the thyroid and the tracheal rings, ensures the airway lumen and prevents the development of a subglottic stenosis. Resection of a ventral part of the cricoid including the adjacent soft tissue lining of the airway causes an irreversible loss of the supporting elastic layer and the formation of scar tissue resulting in a stenosis. It was concluded that the larynx and trachea can be considered as an inner, elastic tube (= conus elasticus) lined with epithelium, suspended to an outer, segmented cartilaginous tube. At the level of the subglottis an intact conus elasticus is of more importance for the normal development of the airway lumen in young rabbits than an intact ventral half of the cricoid ring.     

Prevention of posterior glottic stenosis by mitomycin C

OBJECTIVES: Posterior glottic stenosis (PGS) is a disabling disease commonly induced by endolaryngeal injury from intubation or surgery. Few experiments on PGS prevention, however, have been performed. The purpose of this study was to investigate the preventive effect of mitomycin C (MMC) on PGS in a randomized, controlled animal study. METHODS: Twenty-six rabbits underwent laser injury of 6 W, continuous-focus mode on the posterior glottis. There were 6 uninjured controls. Thirteen animals were treated by a single topical use of 0.4 mg/mL MMC solution on the injured posterior glottis for 5 minutes, and 13 were sham-treated controls. The posterior glottis of all groups was blindly evaluated by telescopic examination under direct laryngoscopy 2 months after injury and was harvested for histologic analyses. RESULTS: Scar, granulation tissue, and synechia formation of the posterior glottis were clearly induced by laser injury. Microscopic examination showed increased collagen content and fibroblast proliferation in the region. The use of MMC significantly decreased the incidence of such gross and microscopic changes of the posterior glottis (p < .05). CONCLUSIONS: This study suggests that topical MMC can be helpful in preventing the progression of PGS from mucosal injury of the posterior glottis.     

丝裂霉素C对LASEK术后Haze形成及角膜基质细胞的影响

目的 探讨在准分子激光角膜上皮瓣下磨镶术(LASEK)中,使用不同浓度丝裂霉素C(MMC)、作用不同时间,对Haze的形成及角膜基质细胞变化的影响.方法 40只兔(80眼),取随机一眼为实验眼,行LASEK术+MMC,对侧眼为手术对照组,仅行LASEK手术.实验眼又随机分为4组:T1组为0.02％的MMC作用10 s,T2组为0.02％的MMC作用30 s,T3组为0.08％的MMC作用10 s,T4组为0.08％的MMC作用30s.另设5兔(10眼)为正常对照组,不做任何手术及药物处理.术后观察Haze形成情况及并发症,并对角膜基质细胞凋亡进行实验研究.结果 术后1周内T1、T2、T3组角膜基本愈合,T4组仍有部分未愈合.术后1周,所有实验组的双眼角膜均有不同程度的Haze形成,其中T2、T3、T4组与手术对照组相比,差异有统计学意义(P＜0.05),而此实验三组之间差异无统计学意义(P＞0.05).T1组与手术对照组相比差异无统计学意义(P＞0.05).各组之间的角膜基质TUNEL阳性细胞数量差异均具有统计学意义(P＜0.05),PT4组＞PT3组＞PT2组＞PT1组＞P手术对照组.结论 MMC可以减少Haze形成,增加作用时间及浓度都会明显抑制Haze的形成,增强角膜基质细胞凋亡.浓度为0.08％,作用时间为30 s的MMC作用最明显,但其对于角膜的刺激作用较大,上皮修复时间较长.     

5-FU乙醇脂质体制备及局部植入治疗家兔喉气管狭窄的效果分析

目的:评价5-氟尿嘧啶(5-FU)脂质体对家兔瘢痕性喉气管狭窄的治疗效果。方法:利用薄膜分散法制备5-FU乙醇脂质体,并检测脂质体形态、包封率等特性。采用刮擦法制备喉气管狭窄模型,环形刮除气管黏膜约0.5cm宽度,诱导气管内瘢痕形成,将诱导瘢痕性喉气管狭窄成功的模型动物随机分为3组:5-FU乙醇脂质体组(A组)、5-FU水溶液组(B组)和生理盐水组(C组),各组分别在内镜监视下经皮穿刺将药物注射入瘢痕内部,并记录此时狭窄度,记为0d,其后,在用药后7、14、21d纤维喉镜下观察记录气管狭窄情况。结果:治疗初期A组和B组并无差别,21d时A组狭窄度明显小于B组,并在观察期间没有出现再狭窄。结论:5-FU脂质体对家兔瘢痕型喉气管狭窄的治疗具有较好的效果,可以作为喉气管狭窄的治疗方法进行进一步研究。     

Therapy of iatrogenic subglottic stenosis: a steroid/antibiotic regimen.

The authors have evaluated the efficacy of a medical regimen, consisting of systemic antibiotics and steroids, in the management of acute iatrogenic subglottic stenosis. The study consisted of the infliction of a standardized subglottic injury to three groups of five dogs: Group I animals were placed on prednisone 1 mg/kg/24 hours and potassium phenoxymethyl penicillin 50 mg/kg/24 hours from the day of the trauma. Group II were placed on a similar regimen from the eighth day after subglottic trauma. Group III received no medical therapy at all. Therapy was continued in treated Groups I and II for five weeks. At the end of the study the dogs were sacrificed and the final degree of subglottic stenosis evaluated, at which time the laryngotracheal complexes were submitted for pathological evaluation. A significant difference was found between the degree of stenosis attained in the three groups. The study suggests that the introduction of a steroid/antibiotic regimen has a beneficial effect in developing subglottic stenosis and that the timing of such therapy is of importance.     

Experience with laryngotracheal reconstruction in subglottic stenosis in a 30 years time period

BACKGROUND: Severe subglottic stenosis is a difficult condition to manage. It can be treated by laryngotracheal reconstruction or cricotracheal resection. PATIENTS AND METHODS: In this retrospective study the experiences for treatment of isolated subglottic stenosis in 37 patients (age: 3-78 years; stenosis grading: 20 x grade II, 13 x grade III, and 4 x grade IV) by laryngotracheal reconstruction in a 30-years experience are presented. RESULTS: In 33 out of 37 patients (89.2 %) a sufficient subglottic patency (postoperative endoscopic finding: stenosis less than 30 %) was achieved by laryngotracheal reconstruction. However, 5 patients of this series had required revision of laryngotracheal recontruction and in 22 patients endoscopic removal of granulation tissue had been performed. Sufficient widening of the subglottic space had been possible in all grade II stenosis (20/20), in 11 out of 13 patients with grade III stenosis, and in 3 out of 4 patients with grade IV stenosis. In one child an accidental decannulation occurred and due to asphyxia an apallic syndrome developed. CONCLUSIONS: Even through laryngotracheal reconstruction is a demanding surgical technique requiring great experience it is an effective option for treatment of subglottic stenosis less than 90 %. For severe subglottic stenosis (> 90 %) treatment by laryngotracheal reconstruction is possible and should be considered if mobilisation of the trachea by scar tissue is suited to be worse or to extended cricotracheal stenosis is present, both being not good candidates for cricotracheal resection.     

Graded carbon dioxide laser-induced subglottic injury in the rabbit model

OBJECTIVE: To conduct an endoscopic and histologic analysis of the subglottic effects of various carbon dioxide laser-induced injuries in the rabbit model. DESIGN: Animals were assigned to either a control (cricothyroidotomy only) group or 4 (cricothyroidotomy and posterior subglottic laser) groups that were injured using varying systematically controlled carbon dioxide laser power exposures (5 W, 8 W, and 12 W), with durations of 2 or 4 seconds, and surface area exposures (25% or 40%). SUBJECTS: Twenty-seven New Zealand white rabbits. INTERVENTIONS: The subglottis was approached via cricothyroidotomy. Control airways were immediately closed, while injured airways were subjected to graded carbon dioxide laser exposures prior to closure. Airways were endoscopically monitored preoperatively, immediately postoperatively, and on postoperative days 1, 7, 14, and 21, after which the animals were humanely killed and subglottic tissue harvested for histological evaluation. RESULTS: Clinical observation revealed no significantly obstructive (acute) stenosis during the duration of the study. Endoscopic visualization revealed the formation of posterior subglottic scarring. Histological analysis of the mucosa revealed that use of carbon dioxide laser resulted in a statistically significant (unpaired, 2-tailed t test, P<.05) proportional thickening of the lamina propria layer, without significant changes in the epithelial and cartilaginous layers. In addition, mucosal blood vessel size increased proportional to the power of the laser delivered to the area (P<.05). CONCLUSIONS: Carbon dioxide laser-induced injury to the subglottis caused localized scarring, lamina propria thickening, and increased vascularity, which resolved with time and was not associated with significant airway obstruction. This model describes a systematic, controlled, and reproducible method of investigating subglottic injury.     

Fibroblast transplantation in the airway: implications for subglottic stenosis

OBJECTIVE: Because subglottic stenosis (SGS) represents one of the most challenging pathologies confronting the pediatric otolaryngologist, our laboratory is investigating the role fibroblasts play in mucosal scar formation in the course of SGS development. Our objective is to establish cell transplantation into the subglottic mucosal wound bed as a viable tool for examining the cellular processes that underlie the development of SGS. DESIGN: A series of 2 animal experiments, with animals assigned to a control, vehicle-only, or cell-treated group. SETTING: John G. Rangos Sr Research Center, Children's Hospital of Pittsburgh, Pittsburgh, Pa. SUBJECTS: Twenty-six New Zealand white rabbits. This animal model has been well established in the study of SGS formation. INTERVENTIONS: Fluorescently labeled exogenous fibroblasts were transplanted into the injured subglottis of the rabbits. RESULTS: Exogenous fibroblasts derived from fetal and adult dermis and subglottic mucosa were successfully transplanted into the injured subglottic mucosa of adult rabbits. Transplanted fibroblasts survived into the latter stages of wound healing (at 14 and 21 days) and appeared to be associated with a mild inflammatory cell influx and active remodeling of the mucosal wound bed. CONCLUSION: Cell transplantation is a viable tool for the study of fibroblast activity in the mucosal wound bed.     

Effects of topical application of mitomycin-C and 5-fluorouracil on myringotomy in rats.

OBJECTIVE: The effects of topical application of Mitomycin-C (MMC) and 5-fluorouracil (5-FU) for maintaining myringotomy patency were investigated in this experimental study. STUDY DESIGN: We performed simple myringotomy with a knife on 140 tympanic membranes of 70 rats. Rats were divided in two study groups and a control group. Each study group had 60 tympanic membranes, and the control group had 20. We applied Mitomycin-C (0.4 mg/ml) in Group A, 5-fluorouracil (50 mg/ml) in Group B topically, and sterile saline in the control group for 10 minutes. Examination was made with otoendoscope on days 1, 3, 5, 7, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, and 70, and patency rates were recorded. RESULTS: MMC and 5-FU Groups remained open for a mean of 46.17 days and 14.62 days, respectively. The control ears healed within 10.4 days. Fibrosis of the MMC-treated group was the same as that of 5-FU-treated groups. Fibrosis of both study groups was significantly lower than that of the control group. CONCLUSIONS: MMC is more effective than 5-FU, which is more effective than the simple myringotomy procedure in extending the patency of myringotomies in rat tympanic membranes (p < 0.05). Both medications are useful as an adjunct in preventing myringotomy closure.     

Endoscopic treatment of subglottic and tracheal stenosis by radial laser incision and dilation

Five patients with severe fibrous subglottic and tracheal stenosis were treated by endoscopic radial laser incision and dilation using both carbon dioxide and neodymium:yttrium aluminum garnet lasers. Good results were noted without complications in all patients in a follow-up period of at least 1 year. Careful selection of patients, excluding those with tracheal collapse or tracheomalacia, and preservation of tracheal epithelium with minimal heat and mechanical trauma are considered essential for good results.     

Ectopic cartilage in subglottic stenosis: hamartoma or reaction to trauma?

In an experimental study in growing rabbits an endolaryngeal injury to the subglottis resulted in the development of a stenosis due to the formation of scar tissue containing ectopic cartilage. For comparison, biopsies taken from the subglottic stenosis in 8 children were studied histologically. In 6 cases ectopic cartilage was observed; all patients had a history of endotracheal intubation. In 3 children the diagnosis hamartoma was made. In the remaining 3 cases the formation of ectopic cartilage might have been a direct reaction to the endolaryngeal intubation. The observations suggest that the formation of ectopic cartilage in acquired subglottic stenosis is not always due to a developmental aberration such as a hamartoma.     

Subglottic wound healing in a new rabbit model of acquired subglottic stenosis

OBJECTIVES: It is difficult to develop models of subglottic stenosis in small animals that ensure reliable stenosis. We therefore sought to establish a new animal model of subglottic stenosis in rabbits and observe subglottic wound healing. METHODS: Using a diode laser under endoscopic visualization after anterior tracheal incision, we induced full-thickness injury to the subglottis in 60 rabbits: 20 on the anterior 120 degrees, 20 on the posterior 120 degrees, and 20 circumferentially. The animals were painlessly sacrificed 2 or 4 weeks later, and their gross and histologic findings were compared with those of each other and with those of 16 age-matched nonwounded controls. RESULTS: Of the 60 animals, 22 died, primarily of acute airway obstruction and mostly in the circumferential group. All of the injured rabbits showed subglottic stenosis compared with controls, ranging from 12% to 56% reduction in cross-sectional area in the 120 degrees injured groups and from 32% to 82% reduction in the 360 degrees injured group. The significant narrowing of the subglottic lumina resulted in substantial early mortality in the circumferential group. Histologic examination showed mucosal ulceration, inflammation, and formation of granulation tissues during the acute phase after wounding, and collapse of the injured cartilage and submucosal thickening and fibrosis at later times. CONCLUSIONS: Subglottic healing and stenosis depend on the extent of cartilaginous injury. This model may be useful for developing methods to treat subglottic stenosis.     

Treatment of severe subglottic laryngotracheal stenosis using hyoid graft with sternohyoid muscle flap]

OBJECTIVE: To evaluate the possibility and reliability of the hyoid-sternohyoid graft transfer in the correction of server subglottic laryngotracheal stenosis, and delineate the operation skills and clinical results. METHODS: Seven patients with severe subglottic stenosis underwent laryngotracheal reconstruction using the hyoid grafts with sternohyoid muscle flaps (HG-SHMF). Five of these patients had traumatic subglottic stenosis, one with scar tissue of unknown etiology arising in the subglottic region, another with tracheal narrowing caused by inhalation of hydrochloric acid. RESULTS: All seven patients were successfully decannulated with moderate good voice. The average time from reconstruction to decannulation was 15.4 months. The stent was endoscopically removed with a range of 3 to 22 months; the mean time required for stenting was 9.6 months. Two patients who received additional salvage reconstruction procedures because of graft or stent displacement were extubated with improved voices and satisfactory airway. CONCLUSIONS: The HG-SHMF transfer was a single-stage reconstruction, relatively simple procedure that can restore an adequate airway and a good voice. Patients undergoing laryngotracheal reconstruction with HG-SHMF must have regular, long-term follow-up since graft displacement and recurrent granulation tissue or scar reformation can cause restenosis after an initially successful surgery. This procedure should be used in a large number of patients to further test its reliability.