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A rat model of wound repair after laryngotracheal injury
Otolaryngology Head and Neck Surgery
Scientific Sessions- - Tuesday
Volume 115 Number 2
muscle fiber types present in human laryngeal muscles. Previous studies have relied on biochemical markers which can distinguish only between fast and slow fibers and cannot differentiate between fast myosin heavy chain (MHC) isoforms or hybrid fibers which express multiple isoforms. We examined the thyroarytenoid (TA), lateral cricoarytenoid (LCA), posterior cricoarytenoid (PCA), interarytenoid (IA), and cricothyroid (CT) muscles obtained from two organ donors and five cadavers with normal larynges using immunohistochemistry with MHC-specific antibodies and in situ hybridization using specific riboprobes to human slow, IIA, and IIX MHC. Muscle fiber type distribution and fiber size were determined for six of each muscle using MHC antibodies. A total of 300 to 600 fibers per muscle were analyzed. The specific fiber type distributions for each muscle were as follows: TA, 23% slow, 50% IIA, 27% IIX; LCA, 25% slow, 62% IIA, 12% IIX; PCA, 52% slow, 37% IIA, 11% IIX; IA, 32% slow, 62% IIA, 6% IIX; and CT, 40% slow, 57% IIA, 1.5% IIX. Average fiber size ranged from 750 to 1650 Hm2, which is one half to one third the size of limb skeletal muscle fibers. In all muscles slow fibers were significantly smaller than type IIA or IIX fibers. In situ hybridization revealed that fiber typing obtained via antibodies could not distinguish hybrid fibers, and depending on the muscle, 20% to 90% of those fibers typed as IIA using antibodies expressed both IIA and IIX MHC messenger RNA. No fibers expressed both slow and fast isoforms. All fibers hybridized with at least one of the three isoforms. In situ hybridization revealed that those fibers previously typed as liB actually expressed the IIX isoform. This study provides previously unavailable basic information about the MHC composition of normal laryngeal muscles, which is intimately linked with their function. This work was supported by a grant from the American Laryngologic Association. 8:45 AM
A Rat Model of Wound Repair after Laryngotracheal Injury DAVID G. DILLARD, MD (presenter), IAN N. JACOBS, MD, and ANTHONY A. GAL, MD, Atlanta, Ga,
Acquired subglottic stenosis occurs in a small percentage of premature infants who require chronic ventilation. Endotracheal tube injury to the mucosal surface of the airway may result in cartilage exposure and subsequent chondritis. This leads to cartilage remodeling and collagen deposition. The new scar tissue may narrow the subglottic airway, resuiting in airway compromise. Wound repair after injury is a complex process that involves epidermal cells, monocytes, fibroblasts, cytokines, and extracellular matrix proteins. To better understand airway wound healing we investigated sequential changes in wound repair and scar tissue formation after posterior cricoid injury in rats. Twenty-eight male adult rats underwent a midline laryngofissure to expose the posterior wall of the cricoid. A rectangular area (1 x 3 mm) of mucosa, submucosa, and
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perichondrium was removed under the microscope. The underlying cricoid cartilage was exposed. The laryngofissure site was then closed. Four rats were sacrificed at each time period (0, 1, 2, 4, 7, 14, and 21 days). Their larynges were harvested, and 5-p.m thick sections were stained with hematoxylin and eosin. Additional sections underwent immunehistochemical staining for fibronectin, cellular fibronectin, procollagen, and tenascin. Optical density measurements of staining intensity were performed on computer-aided microscopy equipment. The wound sites reepithelialized in 4 days by migration and proliferation of basal epithelial cells. Epithelial maturation was complete by 7 days. Fibroblasts infiltrated into the submucosa by 2 days. Fibronectin staining intensity peaked between two and four and returned to normal by 21 days after injury. Procollagen appears later and peaks at 7 days. Tenascin appears between 2 and 4 days at the periphery of the injury site. The expression of these extracellular matrix proteins and cellular elements results in the scar tissue in the airway. This immunohistochemical model of wound repair will serve as a basis for our future studies on the effect of growth factors in the etiology and prevention of subgiottic stenosis. 9:00 AM
Tracheal Reconstruction: Long-term Growth Potential of Auricular Perichondrium and Utility of a Tubed Myoperiosteal Free Flap PAUL T. HOFF, MD (presenter), a n d RAMON M. ESCLAMADO, MD, Ann Arbor, Mich., and Cleveland, Ohio
Reconstruction of extensive laryngotracheal stenoses continues to pose a significant challenge. Previous work from our laboratory has shown the utility of vascularized perichondrium for airway reconstruction. Preliminary work by other investigators suggests that vascularized periosteum may also provide suitable grafting material. We propose in this study to address two different questions using separate animal models. In the first study we propose to examine long-term cartilage growth, epithelial growth, and luminal patency in our previously described rabbit model. Second, we propose to use a canine model to evaluate the efficacy of vascularized periosteum as a donor material to reconstruct a segmental tracheal defect so that this may soon be applied in the clinical setting. 9:15 AM
The Effect of Platelet Derived Growth Factor in Laryngotracheal Surgery Utilizing a Rabbit Model DAVID L. WALNER, MD (presenter), ROBIN T. COTTON, MD, DEAN M. TORIUMI, MD, a n d J. PAUL WILLGING, MD, Cincinnati, Ohio, a n d Chicago, III.
Growth factors are proteins that help regulate the inflammatory response and wound healing in tissues. Multiple growth factors including platelet-derived growth factor (PDGF) have been identified as important cell mediators
Scientific Sessions- - Tuesday
Volume 115 Number 2
muscle fiber types present in human laryngeal muscles. Previous studies have relied on biochemical markers which can distinguish only between fast and slow fibers and cannot differentiate between fast myosin heavy chain (MHC) isoforms or hybrid fibers which express multiple isoforms. We examined the thyroarytenoid (TA), lateral cricoarytenoid (LCA), posterior cricoarytenoid (PCA), interarytenoid (IA), and cricothyroid (CT) muscles obtained from two organ donors and five cadavers with normal larynges using immunohistochemistry with MHC-specific antibodies and in situ hybridization using specific riboprobes to human slow, IIA, and IIX MHC. Muscle fiber type distribution and fiber size were determined for six of each muscle using MHC antibodies. A total of 300 to 600 fibers per muscle were analyzed. The specific fiber type distributions for each muscle were as follows: TA, 23% slow, 50% IIA, 27% IIX; LCA, 25% slow, 62% IIA, 12% IIX; PCA, 52% slow, 37% IIA, 11% IIX; IA, 32% slow, 62% IIA, 6% IIX; and CT, 40% slow, 57% IIA, 1.5% IIX. Average fiber size ranged from 750 to 1650 Hm2, which is one half to one third the size of limb skeletal muscle fibers. In all muscles slow fibers were significantly smaller than type IIA or IIX fibers. In situ hybridization revealed that fiber typing obtained via antibodies could not distinguish hybrid fibers, and depending on the muscle, 20% to 90% of those fibers typed as IIA using antibodies expressed both IIA and IIX MHC messenger RNA. No fibers expressed both slow and fast isoforms. All fibers hybridized with at least one of the three isoforms. In situ hybridization revealed that those fibers previously typed as liB actually expressed the IIX isoform. This study provides previously unavailable basic information about the MHC composition of normal laryngeal muscles, which is intimately linked with their function. This work was supported by a grant from the American Laryngologic Association. 8:45 AM
A Rat Model of Wound Repair after Laryngotracheal Injury DAVID G. DILLARD, MD (presenter), IAN N. JACOBS, MD, and ANTHONY A. GAL, MD, Atlanta, Ga,
Acquired subglottic stenosis occurs in a small percentage of premature infants who require chronic ventilation. Endotracheal tube injury to the mucosal surface of the airway may result in cartilage exposure and subsequent chondritis. This leads to cartilage remodeling and collagen deposition. The new scar tissue may narrow the subglottic airway, resuiting in airway compromise. Wound repair after injury is a complex process that involves epidermal cells, monocytes, fibroblasts, cytokines, and extracellular matrix proteins. To better understand airway wound healing we investigated sequential changes in wound repair and scar tissue formation after posterior cricoid injury in rats. Twenty-eight male adult rats underwent a midline laryngofissure to expose the posterior wall of the cricoid. A rectangular area (1 x 3 mm) of mucosa, submucosa, and
P 115
perichondrium was removed under the microscope. The underlying cricoid cartilage was exposed. The laryngofissure site was then closed. Four rats were sacrificed at each time period (0, 1, 2, 4, 7, 14, and 21 days). Their larynges were harvested, and 5-p.m thick sections were stained with hematoxylin and eosin. Additional sections underwent immunehistochemical staining for fibronectin, cellular fibronectin, procollagen, and tenascin. Optical density measurements of staining intensity were performed on computer-aided microscopy equipment. The wound sites reepithelialized in 4 days by migration and proliferation of basal epithelial cells. Epithelial maturation was complete by 7 days. Fibroblasts infiltrated into the submucosa by 2 days. Fibronectin staining intensity peaked between two and four and returned to normal by 21 days after injury. Procollagen appears later and peaks at 7 days. Tenascin appears between 2 and 4 days at the periphery of the injury site. The expression of these extracellular matrix proteins and cellular elements results in the scar tissue in the airway. This immunohistochemical model of wound repair will serve as a basis for our future studies on the effect of growth factors in the etiology and prevention of subgiottic stenosis. 9:00 AM
Tracheal Reconstruction: Long-term Growth Potential of Auricular Perichondrium and Utility of a Tubed Myoperiosteal Free Flap PAUL T. HOFF, MD (presenter), a n d RAMON M. ESCLAMADO, MD, Ann Arbor, Mich., and Cleveland, Ohio
Reconstruction of extensive laryngotracheal stenoses continues to pose a significant challenge. Previous work from our laboratory has shown the utility of vascularized perichondrium for airway reconstruction. Preliminary work by other investigators suggests that vascularized periosteum may also provide suitable grafting material. We propose in this study to address two different questions using separate animal models. In the first study we propose to examine long-term cartilage growth, epithelial growth, and luminal patency in our previously described rabbit model. Second, we propose to use a canine model to evaluate the efficacy of vascularized periosteum as a donor material to reconstruct a segmental tracheal defect so that this may soon be applied in the clinical setting. 9:15 AM
The Effect of Platelet Derived Growth Factor in Laryngotracheal Surgery Utilizing a Rabbit Model DAVID L. WALNER, MD (presenter), ROBIN T. COTTON, MD, DEAN M. TORIUMI, MD, a n d J. PAUL WILLGING, MD, Cincinnati, Ohio, a n d Chicago, III.
Growth factors are proteins that help regulate the inflammatory response and wound healing in tissues. Multiple growth factors including platelet-derived growth factor (PDGF) have been identified as important cell mediators