Computed tomography-guided tissue engineering of upper airway cartilage.

Normal laryngeal function has a large impact on quality of life, and dysfunction can be life threatening. In general, airway obstructions arise from a reduction in neuromuscular function or a decrease in mechanical stiffness of the structures of the upper airway. These reductions decrease the ability of the airway to resist inspiratory or expiratory pressures, causing laryngeal collapse. We propose to restore airway patency through methods that replace damaged tissue and improve the stiffness of airway structures. A number of recent studies have utilized image-guided approaches to create cell-seeded constructs that reproduce the shape and size of the tissue of interest with high geometric fidelity. The objective of the present study was to establish a tissue engineering approach to the creation of viable constructs that approximate the shape and size of equine airway structures, in particular the epiglottis. Computed tomography images were used to create three-dimensional computer models of the cartilaginous structures of the larynx. Anatomically shaped injection molds were created from the three-dimensional models and were seeded with bovine auricular chondrocytes that were suspended within alginate before static culture. Constructs were then cultured for approximately 4 weeks post-seeding and evaluated for biochemical content, biomechanical properties, and histologic architecture. Results showed that the three-dimensional molded constructs had the approximate size and shape of the equine epiglottis and that it is possible to seed such constructs while maintaining 75%+ cell viability. Extracellular matrix content was observed to increase with time in culture and was accompanied by an increase in the mechanical stiffness of the construct. If successful, such an approach may represent a significant improvement on the currently available treatments for damaged airway cartilage and may provide clinical options for replacement of damaged tissue during treatment of obstructive airway disease.

Prenatal and early postnatal morphogenesis and growth of human laryngotracheal structures.

Advances in neonatal medicine have resulted in increased care of fetal and neonatal airways. These advances have required an exhaustive knowledge of fetal airway anatomy and development. The aim of this study was to determine the anatomical development of laryngotracheal structures during the fetal and immediate postnatal period and to correlate these observations with other fetal biometric parameters to estimate developmental particularities of the fetal airway. An anatomical prospective study was based on examination of larynx and trachea from 300 routine autopsies of fetuses and infants, free of malformation and never intubated. Anatomical measurements of cricoid cartilage, thyroid cartilage, glottis, arytenoid cartilage and trachea were performed using a precision calliper and precision divider. Statistical analysis was performed to represent the growth of anatomical structures and to evaluate the correlation with biometric data. Raw data and 10th and 90th percentile curves were fitted satisfactorily with a linear model for gestational age. A linear relationship between laryngotracheal measurement and body weight and height was observed except for glottis length, interarytenoid distance and anterior cricoid height. The diameter of the cricoid lumen was significantly less than that of the trachea and glottis lumen. A sexual dysmorphism was noted for thyroid cartilage measurements and interarytenoid distance, with measurements significantly smaller in females. This study reports the anatomical development of normal laryngotracheal structures during the fetal period. Despite the fact that this study was performed during postmortem examination, these observations can be useful to develop criteria, materials and surgical procedures adapted to fetal and neonatal airways as well as for the purposes of early diagnosis and management of laryngotracheal malformations.

Whole organ evaluation of collagen in the developing human larynx and adjoining anatomic structures (hyoid and trachea).

The collagen composition (types I, II, and III) of the normal developing human larynx and trachea was examined by biochemical methods. Autopsy specimens of larynges with attached upper tracheal rings were obtained from 28 humans ranging in age from birth to 44 years. The specimens were randomly collected, but excluded if laryngeal disease existed. The age, sex, and cause of death were documented. Collagen is important in the growth, development, repair, regeneration, and structural and functional integrity of the laryngeal framework. A preliminary report of selected cartilaginous components of the larynx was previously published by the authors, which studied the changes in the phenotypic expression of the collagen genes in children from the newborn period to 5 years 10 months of age. The current study included all of the functioning components of the skeletal larynx and trachea. The results of biochemical examination of these tissues are reported, and the potential clinical significance of the results of the study is discussed.

Collagen in the developing larynx. Preliminary study.

The primary purpose of this study was to determine the types of collagen in the developing human larynx that contribute to the structural framework and function of various components of this organ. The infant larynx is much more than a mere miniature of the adult "voice box." There are many age-related differences that occur in the larynx from the newborn period to the adult period of life. While collagen has been studied in numerous tissues, both normal and diseased, there have been no studies of the whole organ content, types, and/or changes of collagen in the developing human larynx that may account for many of the clinical findings. This study may at least in part explain whether collagen differences may account for the structural changes and responses that are seen in clinical practice.

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.

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.

A morphometric study of the growth of the subglottis after endolaryngeal trauma.

In young rabbits the growth of the subglottis was studied over a period of 20 weeks following an endolaryngeal trauma. This lesion of the larynx at the age of 4 weeks resulted in the development of a stenosis in the adult stage. The stenosis appeared to be caused by a specific change in the pattern of growth leading up to an anterior narrowing ('pear'-like) of the cricoid ring and a conspicuous thickening of the subepithelial layer.

Growth of the human prepubertal and pubertal larynx.

Prepubertal, pubertal, and adult measurements of human larynges were used to describe growth of the larynx from prepuberty to adulthood. Linear and weight measurements made of 10 Caucasian male and 10 Caucasian female prepubertal and pubertal cadaveric larynges were compared to adult measurement data obtained from another study on 20 male and 20 female larynges of adults, aged 37-70. In both studies measurements and experimental protocols were similar, allowing direct comparisons to be made between samples. Results highlight specific quantitative characteristics and trends in circumpubertal growth of the laryngeal cartilages and the vocal folds for each sex and with respect to sex differences. The results are discussed with respect to the literature on postnatal laryngeal development and to published research on adolescent voice change.

Does tracheostomy in children retard the growth of trachea or larynx?

Tracheostomy in children has been reported to cause tracheal stenosis, laryngeal stenosis and subsequent difficulties in decannulation. No long term follow-up studies appear to be in the literature describing the events following decannulation. This study compared the radiological size of the cricoid and tracheal lumens with that of normals. Except in special cases tracheostomy did not affect the subsequent growth of the trachea or cricoid. It is suggested that traumatic tracheal narrowing in children should be managed conservatively.

The growth of cartilage from a free perichondrial graft in the larynx.

An attempt was made to promote the growth of new cartilage from a frlage. Three dogs had the perichondrial grafting alone, while two had both the perichondrial grafting and a full thickness buccal mucosal graft. In all these dogs new cartilage was found as early as four weeks postoperative.

Infant primate larynx: developmental histology.

Laryngeal epithelium of monkeys (M. mulatta, M. nemestrina) ranging in age from late fetal to adult was found to undergo the following developmental changes: (1) progressive elaboration of commissural epithelium prenatally, reaching the adult form of pseudostratified columnar ciliated epithelium early in the postnatal period;(2) vocal process epithelium changing from cuboid to stratified squamous tissue during the late prenatal period, and (3) progressive increments in overall epithelial thicknees during development, reaching a maximum in the early postnatal period. Chondrocyte density of the cricoid and thyroid cartilage decreased during the late prenatal period, reaching the adult form at the neonatal stage. The evidence suggests that mechanical or metaplastic stresses do not determine the observed tissue changes.

Effects of highly dosed ultrasound on the growing rabbit's larynx. Scanning electron microscopic and reflecting ligth microscopic findings.

The morphologically recognizable effects of highly dosed ultrasound application on the growing rabbit's larynx (5 Watt/cm2 for 10 min) were studied by means of scanning electron microscopic (SEM) and reflecting light microscopic (RLM) methods. It could be shown that the area of cartilaginous necrosis corresponded exactly to the location of ultrasound application and was sharply limited by the normal cartilage. After a survival period of six weeks, areas of chondroneogenesis became visible on the endolaryngeal perichondrium. After a survival period of twelve weeks, an outer zone of cartilaginous necrosis was still to be seen. Connective tissue directly affected by ultrasound and the endolaryngeal muscles did not show any retarded damage. The results of our experiments permit to state that highly dose ultrasound application on the cartilaginous framework leads to a growth retardation with a subsequent alteration in form and shape.