The Effects of Passive Smoking on Laryngeal and Tracheal Mucosa in Male Wistar Rats During Growth: An Experimental Study.

Cigarettes contain toxic and carcinogenic substances. In this context, cigarette smoking, and similar activities, are associated with numerous pathologies, being considered a risk factor in up to 10% of the total number of deaths in adults. Recent evidence suggests that the exposure of children to smoking in the early days of their development causes many diseases. Using light microscopy, this study aims to analyze the possible histopathological effects of an experimental model of chronic inhalation of cigarette smoke (passive smoking) on the laryngeal and tracheal mucosa of young Wistar rats. A total of 24 young Wistar rats were studied for a period of 120 days. The animals were divided into two groups: passive smoking (n = 16) and control (n = 8). The level of exposure to cigarette smoke was evaluated from the urinary cotinine level. Although no cancerous lesions were identified, histopathological analysis in the laryngeal and tracheal mucosa of all the animals in the experimental group showed that the proportion of moderate and focal inflammation was higher in animals exposed to chronic inhalation of cigarette smoke (P = 0.041). Histopathologic analysis revealed moderate and focal inflammatory lesions in the region of the infraglottic mucosa in exposed animals, although without dysplastic or neoplastic lesions in the laryngeal and tracheal mucosa.

Morphological development and expression of neurotrophin receptors in the laryngeal sensory corpuscles.

Morphological development of sensory structures in the laryngeal mucosa of postnatal rats was observed by use of immunohistochemistry for protein gene-product 9.5 (PGP9.5). Moreover, expression changes of high affinity neurotrophin receptors, TrkA, TrkB and TrkC, and low affinity neurotrophin receptor p75(NTR) were examined to elucidate the relationship to morphogenesis. Intraepithelial nerve endings and parent axons of the laminar endings with immunoreactivity for PGP9.5 have already appeared in the rat on embryonic day 18 (E18) as well as solitary chemoreceptor cells in the glottic cleft. According to neurotrophin receptors, TrkA immunoreactivity were observed on and after postnatal week 3 (3W) in the nervous sensory structures, that is, free nerve endings, laminar endings and sub- and intragemmal plexuses of the taste buds. In the laminar endings, TrkC immunoreactivity was also observed on and after 3W. According to the laryngeal sensory cells, the solitary chemoreceptor cells were immunoreactive to TrkA, TrkB, and TrkC on and after postnatal day 3 (P3). In the taste buds in arytenoid region, taste cells were immunoreactive for TrkA, TrkB, and TrkC on and after 3W, P14, and 3W, respectively. Immunoreactivity for p75(NTR) was observed on the surface of taste cells on and after P9. The results of the present study suggest that sensory structures in the laryngeal mucosa were developed on perinatal days to involve respiratory reflex, and that neurotrophin receptors may take part in the regulation and maintenance of sensory structures.

Functional histology of the macula flava in the human vocal fold--Part 2: Its role in the growth and development of the vocal fold.

OBJECTIVE: This study aims to clarify the role of the maculae flavae (MFe) during growth and development of the human vocal fold mucosa (VFM). METHODS: Our current results concerning the MFe in the human newborn, infant, and child VFM are summarized. RESULTS: Newborns already had immature MFe at the same sites as adults. They were composed of dense masses of vocal fold stellate cells (VFSCs), whereas extracellular matrix components were sparse. VFSCs in the newborn MFe had already started synthesizing extracellular matrices (EM). During infancy, the EM synthesized in the MFe appeared in the VFM to initiate the formation of the three-dimensional extracellular matrix structure of the human VFM. During childhood, MFe including VFSCs continued to synthesize EM such as collagenous, reticular, and elastic fibers, and hyaluronic acid (glycosaminoglycan), which are essential for the human VFM as a vibrating tissue. The MFe in newborns, infants and children were related to the growth and development of the human VFM. CONCLUSION: Human MFe including VFSCs were inferred to be involved in the metabolism of EM, essential for the viscoelasticity of the human VFM, and are considered to be an important structure in the growth and development of the human VFM.

Local control of murine subglottis development.

OBJECTIVE: To determine if subglottic development is at least partially under local control and to determine which tissue layer(s) is predominantly responsible. DESIGN: The suglottises of 12 day-3 CD1 mice were grown in whole organ culture. The 12 subglottises were divided into 3 individual groups: +++, -++, and ---. Group+++ had all tissue layers of the subglottis intact: luminal epithelium, cricoid cartilage, inner and outer perichondrium. Group-++ had all layers intact with the exception of luminal epithelium. Group--- had all layers removed (luminal epithelium, inner and outer perichondrium) resulting in cricoid cartilage-only rings. All rings were grown in basic medium without the use of growth factors or serum for 15 days. Measurements of the rings were taken before and after organ culture growth. RESULTS: Group+++ was the only group that experienced growth. Only luminal growth was statistically significant although all rings experienced growth in both the luminal and external diameter. Group-++ did not experience any growth. Group--- lost structural integrity with collapse of the ring and did not experience growth of any dimension of the cartilage. CONCLUSIONS: Growth of the subglottis is under local control but may have additional influences from the outside that were not investigated here. Removal of just the epithelium stunts growth of the entire ring, but preferentially the lumen more so than the external diameter. Removal of all tissue layers around the cricoid cartilage results in a structural collapse of the ring, suggesting that the cartilage in this age group is dependent on surrounding tissues for structural integrity.

Age related changes in the distribution of laryngeal glands in the human adult larynx.

OBJECTIVE: To understand the defense status of glandular tissue in the human adult larynx, we examined the glandular distribution, concentration and the ratio of the mucous glands in the subglottic and supraglottic larynx. METHODS: One-hundred and seven adult larynges were examined. The area of the mucosa and glands were measured using an image analyzing system. The correlations of the concentration as well as the ratio of the mucous glands with age were analyzed. RESULTS: The ratio of the areas occupied by the glandular acini of the subglottis was almost equal to that of infants as we reported earlier. The concentration of the laryngeal glands decreased in higher-aged adults at the level of the supraglottis, but not at the level of the subglottis. CONCLUSIONS: In comparison to infants, the ratio of laryngeal glands of the human subglottic larynx is rather constant in human adults. However, the supraglottic larynx seems to be influenced by increasing age. The changes deserved in the laryngeal glands predominantly occur in the supraglottic larynx.

Regenerative medicine of the trachea: the first human case.

OBJECTIVES: The objective of the present study was to demonstrate regenerative medicine of the tracheal tissue by using an in situ tissue engineering technique for airway reconstruction. METHODS: Based on the previous successful experimental animal studies, the current regenerative technique was applied to repair of the trachea of a 78-year-old woman with thyroid cancer. A Marlex mesh tube covered by collagen sponge was used as a tissue scaffold. The operative intervention included right hemithyroidectomy, resection of the trachea, and tracheoplasty using the scaffold. The right half of three rings of the trachea was resected, and the scaffold material was sutured to the defect of the trachea. RESULTS: After 2 weeks, the mesh collagen structure of the artificial material could be seen with endoscopy in most of the implanted area. The artificial material was covered with epithelial growth after 2 months. Epithelialization continued to cover the artificial material completely for 2 years without any complications. CONCLUSIONS: The current regenerative technique avoided tracheotomy, a second operation, and deformity. Good epithelialization has been observed on the tracheal luminal surface without any complications for 2 years. Although long-term observation is required, regenerative medicine of the tracheal tissue appears feasible for airway reconstruction.

Gross and histologic changes in the developing rabbit subglottis in response to a controlled depth of injury.

OBJECTIVE: Our goal was to determine the effects of both perichondrial and intracartilaginous injury in the developing rabbit subglottis versus normal development. DESIGN: We conducted a descriptive, pilot study of changes in the shape and histology of the subglottis after a controlled depth of injury in 27 New Zealand White rabbits, ages 4 weeks, 8 weeks, and 1(1/2) years. INTERVENTION: Within each age group, 3 animals underwent no surgery, 3 underwent perichondrial injury, and 3 underwent intracartilaginous injury. RESULTS: Perichondrially injured animals in the 4-week age group developed a marked abnormality in the shape of the cricoid cartilage in the injured region. Cartilage of the perichondriallly injured animals in the 8-week and 1(1/2)-year groups became histologically consistent with fibrous tissue. The cartilage of all animals that underwent intracartilaginous injury was replaced with fibrous tissue. CONCLUSION: In this observational study, we identified 3 relevant findings. First, the responses of the cartilage to a perichondrial injury suggest that the luminal soft tissues may exert some morphologic control in developmentally young animals. Second, only the 4-week-old group's cartilage was tolerant of a perichondrial injury with continued growth of the ring. Third, no animal's cartilage could withstand an intracartilaginous injury regardless of age.

Growth of laryngeal glands in infants: a comparative study of the sub- and supraglottic larynx.

It is well established that laryngeal secretions, produced in the glandular tissue, contribute to the local defence system of the larynx. In this study, the distribution of glandular tissues in the subglottic and supraglottic larynx was examined in 55 infants, using an image analysing system. The average area of the mucosa or glandular acini at the level of the false vocal fold was almost three times larger than that of the subglottis. At both levels, the area occupied by the glands increased according to the age at death and became more evident according to the total development score (TDS) which was estimated by adding the age at death and the gestational period. The results indicate that the glandular structure of the level of the sub- or supraglottic larynx develops constantly and equally after birth.

Fibroblasts in human vocal fold mucosa.

Fibroblasts in two portions of the vocal fold mucosa, the macula flava (MF) and Reinke's space (RS), were investigated by means of transmission electron microscopy. Five adult and five newborn larynges from autopsy cases were subjected to study. Both in adults and newborns, fibroblasts in the MF tended to be stellate in shape, to have a small nucleus/cytoplasm (N/C) ratio, and to have a well developed rough endoplasmic reticulum (rER) and Golgi apparatus (GA). They were active in producing collagenous and elastic fibres. Most fibroblasts in RS were oval in newborns and spindle-shaped in adults. They had a large N/C ratio and less developed rER and GA, indicating that they were inactive in producing fibres. We postulate that many fibroblasts in the newborn and infant MF contribute to the development of the vocal ligament, many of those in the adult MF participate in the metabolism of the vocal ligament, and many of those in RS of both age groups are activated when the vocal fold tissue is injured.

[Age-related development of the arrangement of connective tissue fibers in the lamina propria of the human vocal folds--scanning electron microscopic examination with digestion method].

The lamina propria of the human vocal fold consists of a superficial, intermediate, and deep layer. This stratified structure is thought to facilitate phonation. Each layer has different physical properties based on different alignment and distribution of collagen and elastic fibers. In the present study, developmental changes in vocal fold structure were studied in human fetuses, infants, and children, with special reference to the pattern of distribution of collagen and elastic fibers. Vocal fold specimens were obtained at autopsy from 5 fetuses, 7 neonates, 3 infants, 3 children at the age of 1 year, 3 children at 3 years, 3 children at 5 years, 3 children at 12 years, and 5 subjects at ages ranging from 15 to 22 years. Prior to the examination of collagen fibers, elastic fibers and cells were dissolved with 10% sodium hydroxide treatment. Prior to the examination of elastic fibers, collagen fibers and cells were dissolved by treatment with 90% formic acid. The specimens were then dehydrated, dried, ion-coated with platinum, and examined with a scanning electron microscope. In fetuses and infants, thin, coiled fibers were found distributed densely in the anterior, posterior, and deep parts of the lamina propria, while irregular thick fibers were sparsely seen in the superficial layer of the vocal fold. In children aged 1 to 3 years, the dense fibers in the deep part decreased, and the longitudinal fibers in the superficial layer increased. In children at 5 years of age, longitudinal collagen and elastic fibers were noted in all of the layers of the vocal fold. The distribution of fibers was uniform irrespective of the depth. At 12 years of age, thin, coiled fibers were noted in the superficial layer, while thin, irregular fibers were found in the deep layer. At 17 years, differentiation of the superficial and deep layers was more evident. In male subjects after adolescence the curvature of curly collagen fibers decreased, and the diameter of fibers increased. The present findings suggest that the development of the vocal fold in childhood occurs in two steps. In the first step, dense fibers in the anterior, posterior, and deep parts of the lamina propria found in fetuses and infants shift to the anterior and posterior ends of the vocal fold, between which longitudinal fibers appear. During this step only simple phonation is possible. In the second step, differentiation of the superficial and deep layers occurs, and the stratified structure of the vocal fold appears in the teens. This step is probably related to the complicated modality of phonation in this age group. In males, the development of the vocal fold is completed after changes in collagen fibers during the mutation.