Effects of mechanical trauma to the human tympanic membrane: an experimental study using transmission electron microscopy.

HYPOTHESIS: To examine the cellular events following induced superficial lesions of the human tympanic membrane (TM). Such information could lead to enhanced appreciation of repair mechanisms and novel strategies to restore TM perforations. BACKGROUND: Persistent perforation of the TM in chronic otitis media is a major global health problem and frequently necessitates surgical intervention. Most TM perforations heal spontaneously and swiftly, but sometimes healing fails. The underlying mechanisms and the reason for incomplete repair are often elusive, although some mechanisms have been proposed. METHODS: Here, five healthy adult human TMs were sampled during vestibular schwannoma surgery. Three days before harvesting, three TMs were superficially lesioned, including the epithelial and sub-epithelial layers, using a needle and two TMs served as controls. Light and transmission electron microscopy were performed. RESULTS: Surrounding lesion showed distinct ultrastructural changes. This included a keratinocyte frontier with electron-dense cells with abundant ribosomes and nuclei metamorphosis. Beneath, were activated fibroblasts and invaded/transformed free cells and signs of increased transcellular activity of adjacent blood vessels. CONCLUSIONS: The study describes dynamic morphological events of a human lesioned TM. The human model may be used for further investigations and understanding of TM healing mechanisms.

Emergence of Refractory Otomastoiditis Due to Nontuberculous Mycobacteria: Institutional Experience and Review of the Literature.

BACKGROUND: Nontuberculous mycobacteria (NTM) infection has attracted increasing attention in recent years; however, NTM otomastoiditis is extremely rare. Surgery combined with antibiotic therapy is the current mainstay of treatment; however, the reported duration of medication still varies. In this study, we aimed to analyze patients with NTM otomastoiditis and establish a more efficient treatment strategy. METHODS: Medical records and temporal bone images of patients with NTM otomastoiditis were retrospectively analyzed. In addition, a comprehensive review of cases with NTM otomastoiditis in the literature was also performed. RESULTS: Twenty-two patients were identified in our institution, and all patients had refractory otorrhea. The rates of granulation tissue, otalgia, and facial palsy were 90.9%, 31.8%, and 9.1%, respectively. Soft tissue attenuation via imaging studies was demonstrated in all of the middle ear cavities. All patients received medical treatment, 20 (90.9%) underwent surgery, and 4 (18.2%) underwent revision surgery. The median time to cure was similar between the "prolonged-course" and "standard-course" antibiotic groups (3.0 vs 3.3 months; P = .807). However, the former had a longer median duration of antibiotic therapy (6.0 vs 3.0 months; P = .01). In the literature review, 54 (96.4%) patients received medical treatment, 51 (91.1%) underwent surgery, and 27 (48.2%) underwent revision surgery. CONCLUSIONS: NTM otomastoiditis should be suspected if a patient has chronic refractory otorrhea and ear granulation tissue. Surgery, which is the mainstay of treatment, should be complemented with antibiotics. In those without temporal bone osteomyelitis, antibiotic treatment can be stopped after a dry ear is achieved.

Morphology and ciliary motion of mucosa in the Eustachian tube of neonatal and adult gerbils.

The Eustachian tube is a small canal that connects the tympanic cavity with the nasal part of the pharynx. The epithelial lining of the Eustachian tube contains a ciliated columnar epithelium at the tympanic cavity and a pseudostratified, ciliated columnar epithelium with goblet cells near the pharynx. The tube serves to equalize air pressure across the eardrum and drains mucus away from the middle ear into the nasopharynx. Blockage of the Eustachian tube is the most common cause of all forms of otitis media, which is common in children. In the present study, we examined the epithelial lining of the Eustachian tube in neonatal and adult gerbils, with a focus on the morphological and functional development of ciliated cells in the mucosa. The length of the tube is ∼8.8 mm in adult gerbils. Scanning electron microscopy showed that the mucosal member near the pharyngeal side contains a higher density of ciliated cells and goblet cells than that near the tympanic side. The cilia beat frequency is 11 Hz. During development, the length of the Eustachian tube increased significantly between postnatal day 1 (P1) and P18. Scanning electron microscopy showed that the mucosa contained a high density of ciliated cells with a few goblet cells at P1. The density of ciliated cells decreased while the density of goblet cells increased during development. At P18, the mucosa appeared to be adult-like. Interestingly, the ciliary beat frequency measured from ciliated cells at P1 was not statistically different from that measured from adult animals. Our study suggests that the Eustachian tube undergoes significant anatomical and histological changes between P1 and P18. The tube is morphologically and functionally mature at P18, when the auditory function (sensitivity and frequency selectivity) is mature in this species.

Assessment of the ototoxicity of almond oil in a chinchilla animal model.

OBJECTIVES/HYPOTHESIS: Almond oil is frequently prescribed as a ceruminolytic, to soften ear wax or relieve ventilation tube occlusion. Ceruminolytics could lead to ototoxicity in the presence of a tympanic perforation. Reports on the safety of almond oil as a ceruminolytic is limited. The present study aimed to assess the effect of ototopic almond oil on hearing. STUDY DESIGN: Prospective, randomized, controlled trial in a chinchilla animal model. METHODS: Bilateral myringotomies were performed in 19 female chinchilla. One randomly selected ear received almond oil, whereas the other ear received saline applied transtympanically. Auditory Brainstem Response (ABR) testing was performed prior to application and at 14 and 30 days following application. Postmortem Scanning Electron Microscopy (SEM) images were obtained to assess cochlear hair cell status. RESULTS: At 30 days following application, there was no significant change in ABR thresholds at 16, 20, or 25 kHz. No cochlear hair cell loss was observed with SEM. CONCLUSIONS: In the chinchilla, when a tympanic perforation is present, almond oil does not seem to cause ototoxicity. Further studies are needed to better assess the effect of almond oil on hearing in humans.

The posterosuperior quadrant of the tympanic membrane.

OBJECTIVE: To document the anatomic relationship of the posterosuperior segment of the tympanic membrane. STUDY DESIGN: Point-prevalence study of postmortem material. SUBJECTS AND MATERIALS: One hundred seventy temporal bones from adults without history of otologic pathology were studied. The anatomic details of the tympanic membrane were observed, histology was documented on 30 sectioned bones, and, with the use of the scanning electron microscope, lamina propria was studied in 40 bones. RESULTS: We established that the posterosuperior quadrant was the largest, representing 28.7 percent of the total area of the eardrum. This feature implies that this quadrant is more likely than are the other quadrants to exhibit changes prompted by pressure challenges. We also verified that the posterosuperior part of tympanic sulcus, above the chorda tympanic nerve, lost its depth in an unexpected way in 60 percent of the cases. In association, we established that at this level there was a decrease in annulus caliber. In 25 percent of cases, by the absence of the circular fibers of lamina propria, the posterosuperior quadrant had only the radial fiber layer. CONCLUSIONS: Anatomic and morphologic details in posterosuperior quadrant were newly described and may explain the greater incidence of retraction pockets and marginal perforations that provide origin for the cholesteatoma.

[Reconstruction of three different kinds of tympanic membrane in vitro by tissue engineering technique].

OBJECTIVE: To reconstruct three different kinds of tympanic membrane in vitro by tissue engineering technique, and to examine their histological structures and mechanical properties. METHODS: The skin and dura of pig (weight 30 kg) were processed with high satuated saline and enzymes to make extracellular matrix. Meanwhile, fibroblasts (1 x 10(6)/mL, 0.2 mL) were seeded on the surface of these two scaffolds and collagen. The composite tissues were cultured in vitro for 1 week and examined in histological structure and mechanical properties. RESULTS: Fibroblasts cultured were spindle-shaped and could grow and attach to these scaffolds with a arrangement of sarciniform and parallel. The reconstructed tissue of ECM and collagen appeared to integrate well and had better bio-compatibility. The mean thickness of the collagen, the skin and the dura (all covered with fibroblasts) were 9.4, 10.0 and 10.4 microm respectively. The tension of the collagen was (1.417+/-0.030) N/mm2, of the acellular dermal matrix was (24.500+/-2.040) N/mm2 (being close to the tension of normal tympanic membrane, 26.700 N/mm2), of the acellular dura was (53.300+/-2.600) N/mm2. CONCLUSION: The results suggest that the tension and the thinkness of acellular dermal matrix is similar to the normal tympanic membrane of guinea pig, it is an ideal material for tympanoplasty.

Stem cells and enhanced healing of chronic tympanic membrane perforation.

CONCLUSIONS: Important information about the basic reparative process of tympanic membrane (TM) healing is shown, which can be incorporated for further clinical understanding. This provides a basis for the exploration of stem cell treatment for TM perforations and holds promise for future improvements. OBJECTIVES: This study aimed to analyse the healing of TM perforation by using stem cells and the stiffness of the membrane was tested in an acute and long-term study. MATERIALS AND METHODS: Sprague-Dawley rats were used in a model of TM perforation. The perforation was performed with a laser system. Stem cells were applied and the healing time and morphological analysis were performed with light and transmission electron microscope. Stiffness was examined by moiré interferometry. RESULTS: The stiffness of the perforated and healed TM was restored after just 2 weeks. In the chronic perforation model, mesenchymal stem cells enhanced the healing.

Biocompatibility of polylactides in the middle ear: an experimental animal study.

OBJECTIVE: To study the suitability of resorbable polylactides for reconstruction in middle ear surgery. EXPERIMENTAL ANIMALS: Twenty-four chinchillas. INTERVENTIONS: Twenty chinchillas were operated on using 2 types of polylactides inserted into the middle ear and outer ear canal, 10 animals in each group. At 6 months, the animals were killed, and their temporal bones were removed for sectioning. Degree of inflammation, fibrosis, degradation of polylactide material, and new bone formation were assessed histologically. MAIN OUTCOME MEASURES: Polylactide implants caused only a mild local inflammation, with no difference between the 2 materials tested. CONCLUSION: Polylactide acid-based middle ear implants were successfully used in the reconstructive middle ear surgery. Resorption occurred without complications.

Efecto del factor de crecimiento endotelio vascular (VEGF) sobre perforaciones timpánicas en ratas Long-Evans / Effect of vascular endothelial growth factor (VEGF) on tympanic perforations in Long-Evans rats

Introducción: La anglogenesis es el desarrollo de nuevos vasos sanguíneos desde una red vascular existente, contempla una secuencia de eventos complejos y es fundamental en el proceso reparativo. Existen múltiples factores estimulantes de la angiogénesls, entre ellos se encuentran factores de crecimiento como el VEGF (factor de crecimiento endotelio vascular). Debido a su rol reparativo se han utilizado factores proanglogénicos para reparar perforaciones timpánicas. Objetivo: Estudiar el efecto del VEGF sobre perforaciones timpánicas de ratas Long-Evans. Material y método: Se usan 15 ratas adultas, se realizan perforaciones timpánicas bilaterales, se instilan al azar las perforaciones con solución fisiológica y VEGF, se realiza visualización microscópica de los tímpanos a los días 9,15 y 21 posperforación. Las ratas son sacrificadas el día 21 y se realiza estudio histológico del grosor timpánico. Resultados: No se aprecia un efecto inductivo del VEGF sobre el cierre de las perforaciones timpánicas, se produce un aumento en el grosor timpánico de las ratas tratadas con VEGF.

Utilization of fresh human tympanic membranes for structural analysis and cytokeratin immunocytochemistry implementing resin techniques.

CONCLUSIONS: The results of this study have demonstrated for the first time that tympanic membrane (TM) structure is preserved following removal of fresh, normal tissue from patients undergoing surgery. Greater clarity has been demonstrated using resin sections than in previous studies on paraffin sections. Of particular note, cytokeratin (CK) immunocytochemistry was successfully performed on resin sections, which has not been previously reported. This may have potential applications for future work involving tissues that express CKs. OBJECTIVES: To analyse the structure of normal, fresh human TM specimens after surgical removal and to evaluate their CK immunocytochemistry using resin techniques, neither of which have been demonstrated previously. MATERIAL AND METHODS: Seven TM specimens were removed during surgery and then preserved in a modified Karnovsky's fixative. Semi-thin and thin sections were examined by means of light and electron microscopy, respectively. For comparison purposes, paraffin block-embedded specimens were also sectioned. CK immunocytochemistry was performed on semi-thin sections using standard immunoperoxidase techniques, with expression being demonstrated using light microscopy. RESULTS: The three-layer architecture of the TM was preserved. The morphology of the TM was vastly superior in the semi-thin resin sections than in the thicker paraffin sections. The outer, middle and inner layers were clearly demonstrated. The integrity of the outer epithelial layer was maintained, with an outer keratinizing stratum corneum and underlying stratum granulosum, stratum spinosum and stratum basale layers resting on the basal lamina. The thin inner mucosal layer was also viable, consisting of simple squamous or cuboidal cells. Preservation of the middle lamina propria was achieved, with demonstration of the outer radial and inner circular fibres. CK immunocytochemistry utilizing resin techniques provided excellent staining of CK 7 and 8 in the inner layer, with positive staining of CK 5 and 10 in the outer layer.

Achados otomicroscópicos e histológicos da miringoesclerose induzida em ratos: estudo crítico de um modelo experimental / Otomicroscopic and histologic findings of induced myringosclerosis in rats: a critical study of an experimental model

A miringoesclerose é caracterizada por hialinização da lâmina própria da membrana timpânica. Trabalhos experimentais têm utilizado a otomicroscopia ou a histologia para estudar a miringoesclerose em animais, porém sem correlacionar precisamente estes dois métodos de avaliação. OBJETIVO: Este estudo procura avaliar a acurácia da otomicroscopia no diagnóstico de miringoesclerose em modelo experimental com ratos. FORMA DE ESTUDO: experimental. MATERIAL E MÉTODO: Foi induzida miringoesclerose por inoculação transtimpânica de Streptococcus pneumoniae em 25 ratos Wistar, que foram examinados quinzenalmente por otomicroscopia e sacrificados após oito semanas, tendo suas membranas timpânicas examinadas por histologia. RESULTADOS: Da confrontação das alterações otomicroscópicas com os achados histológicos, houve uma sensibilidade de 80 por cento e especificidade de 75 por cento. CONCLUSÃO: Pelos resultados obtidos neste estudo, a otomicroscopia não representou um bom método para avaliar a miringoesclerose neste modelo experimental.

[Microstructure of tympanic membrane in chronic otitis media: immunohistochemical evaluation]. / Mikrostruktura blony bebenkowej w przewleklym zapaleniu ucha srodkowego w badaniach immunohistochemicznych.

Chronic otitis media (COM) is an inflammatory process involving the middle ear mucosa and the tympanic membrane. The healing and epidermization is mostly impaired by immunological response of the host. Investigating the activity and the function of immunological response elements one can learn the immunological mechanisms taking place in chronic otitis media. The ultrastructural investigations of the tympanic membrane were done on its fragments obtained from 19 patients with COM during middle ear surgery, performed at ENT Department of Medical University of Gdansk in the years 1997-1999. Immunohistochemical investigations were performed using monoclonal antibodies against tenascin, S-100 protein, Ki 67, CD 31, F VIII, HLA-DR, TGFbeta1 and EGFR. The control group was 11 healthy tympanic membranes from cadavers. The presence of tenascin was proven in all COM tympanic membranes and in 45.5% of those from control group. S-100 protein was present in 88.9% of the patients with COM and absent in control group. Ki 67 was observed in 44.4% of the patients with COM and in 27.3% of the healthy tympanic membranes. Angiogenesis factors (CD 31 and FVIII) were present in 77.8% of the investigated COM tympanic membranes, in control group in 45.5%. HLA-DR expression was observed in 90% COM patients, in control group in 72.7%. Growth factor TGFbeta1 was present in the all cases in mucous and fibrous layer and in 54.5% of healthy tympanic membranes. EGF receptor was present in 60% of COM patients, mainly in epithelial layer of tympanic membrane and in 54.5% of those from control group. The presented investigations confirm the immunological activity of tympanic membrane in chronic otitis media.

The effects of incisional myringotomy and CO2 laser myringotomy on rat tympanic membranes.

OBJECTIVE: The purpose of this study was to compare closure rates and histopathological findings of carbon dioxide (CO(2)) laser myringotomies to those of incisional myringotomy. STUDY DESIGN: We performed CO(2) laser round myringotomy on left ears and incisional round myringotomy on the right ears of 34 rats on the same day. The incisions were 2mm in size on both ears. Examination was made with otoendoscope on days 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 25, 30, 35, 40, 45, 50 with videorecording. Two rats were sacrificed every 2 days for 25 days and every 5 days thereafter, randomly. The tympanic membranes (TM) were excised and hyalinization, fibrosis and inflammation were assessed with light microscope. RESULTS: Most of the CO(2) laser myringotomies healed after day 15, and all of them healed by day 50. Three of the incisional myringotomy perforations were closed at day 3, and the rest by day 15. Patency of CO(2) laser myringotomies was significantly longer than that of incisional myringotomies. Hyalinization, fibrosis and inflammation of the incisional myringotomy group were significantly less than those of the laser myringotomy group. CONCLUSIONS: CO(2) laser myringotomies remain patent for a longer period of time than the incisional procedure, however, they cause more tissue inflammation. We believe that CO(2) laser myringotomy is an effective method, however, additional studies are needed to identify its complications.

Pathology of aural abscesses in free-living Eastern box turtles (Terrapene carolina carolina).

Aural abscess or abscess of the middle ear is common in free-living Eastern box turtles (Terrapene carolina carolina) of Virginia (USA) and elsewhere. Although its etiology remains unknown, hypovitaminosis A has been suggested on the basis of similar lesions occurring in captive chelonians fed diets that are deficient in vitamin A. This hypothesis was supported by significantly greater body burdens of organochlorine compounds (reported disruptors of vitamin A metabolism) and a nonsignificant trend toward lower serum and hepatic vitamin A levels in free-living box turtles with this lesion. The tympanic epithelium was evaluated in 27 box turtles (10 with aural abscesses and 17 without). Lesions of the tympanic epithelium of box turtles with aural abscesses included hyperplasia, squamous metaplasia, hyperemia, cellular sloughing, granulomatous inflammation, and bacterial infection. These changes were more severe in turtles with aural abscesses than in those without and were more severe in tympanic cavities that had an abscess compared to those without when the lesion was unilateral. Organs from 21 box turtles (10 with aural abscesses and 11 without) from the study population were examined for microscopic lesions, and minimal histopathologic changes were found, none of which were similar to those found in the tympanic epithelium. Histopathologic changes in box turtles with aural abscesses were consistent with a syndrome that may involve hypovitaminosis A.

Tympanic membrane vessel revisited: a study in an animal model.

HYPOTHESIS: The present study aimed at elucidating whether there are blood vessels in the semitransparent portion of the tympanic membrane. BACKGROUND: The normal semitransparent portions of pars tensa show strikingly few, small-caliber vessels under the otomicroscope. The major portion of a pars tensa seems to be devoid of blood vessels. In inflammatory conditions of the middle ear, the vascular pattern of the tympanic membrane is dramatically altered, and blood vessels traversing the pars tensa can be discernable. METHODS: The study was performed in rats with healthy tympanic membranes and in tympanic membranes obtained from animals with purulent otitis media evoked by inoculation of Str. pneumoniae. The tympanic membrane vessels were dilated by injection of adenosin, and directly afterwards the animal was perfused with china ink. Vessels were also revealed by immunohistochemistry with antibodies for Thy-1 and the von Willebrand factor as well as by detection of carbon particles at an ultrastructural level. RESULTS: Adenosin caused a marked dilation of the mallear and annular vessels. However, no preexisting vasculature was revealed in the normally transparent portions of the pars tensa except single vessels in the posterior quadrant and in the lower quadrants. In Str. pneumoniae-induced acute otitis media, the tympanic membrane thickened, bulged, and discolored. Even then, at 12 hours after inoculation, no vessels could be distinguished in the normally transparent portions of the tympanic membrane. However, at 4 and 7 days of acute purulent otitis media, vessels developed in those areas, most probably through ingrowth of newly formed vessels. CONCLUSION: The results support the view that the semitransparent portions of the pars tensa lack vascularity. In inflammation, new vessels are formed in pars tensa to meet the demand for an increased blood supply.

Tympanic membrane changes in experimental cholesteatoma in the gerbil.

HYPOTHESIS: The current study aimed to analyze changes of the acoustic stiffness properties and coincident morphologic changes of the tympanic membrane (TM) in early stages of cholesteatoma. The results were compared with those obtained in previous studies of otitis media with effusion and purulent otitis media. BACKGROUND: The pathophysiology of the cholesteatoma development process remains unclear. Changes of the physical properties of the TM seem to play an important role. In chronic middle ear disease, atrophic regions and retraction pockets of the TM are often present. Such changes of the TM may facilitate the development of cholesteatoma. METHODS: Early stages of external ear canal cholesteatoma were produced in the Mongolian gerbil by closing the external meatus with a suture. Acoustic admittance measurements were performed after 1, 2, or 3 months after surgery. After completed measurements, the TMs were analyzed morphologically. RESULTS: The acoustic stiffness of the TM was significantly increased in all cholesteatoma ears compared to normal control ears. A pronounced thickening and proliferative activity of the entire TM was observed in the early stage of cholesteatoma. The outer keratinizing epithelium showed an increased number of cell layers and an increased keratin production. The fibrous layer was thickened because of an increased amount of collagen fibers combined with minor edema. CONCLUSIONS: The thickness of the fibrous layer was almost doubled, mostly because of an increased amount of collagen fibers. The acoustic stiffness was significantly increased in all cholesteatoma ears. The results of the acoustic admittance measurements are comparable with those obtained in previous studies on purulent otitis media and otitis media with effusion.

Titanium as an ossicular replacement material: results after 336 days of implantation in the rabbit.

OBJECTIVE: Titanium in other parts of the body, well known for its biocompatibility, was examined in an animal model for its use as an ossicular replacement material. STUDY DESIGN: The biocompatibility of titanium was studied in the middle ear of rabbits using light and scanning electron microscopy. Titanium pins were placed as middle ear prostheses or as free implants and were examined after 28, 84, 168, and 336 days. RESULTS: After 28 days, the prostheses were covered by regular mucosa. The free implants took up to 336 days to be totally epithelialized. There were no inflammatory cells observed on the surface of the material nor were unusual amounts of fibrous tissue seen. In addition, the titanium material exhibited an affinity toward bone. CONCLUSIONS: The results of this animal experiment indicate that titanium is a useful material for ossicular replacement prostheses.

Clinical-morphological correlation of nasal obstruction with skull base development and otitis media. An experimental study.

Morphological changes of the middle ear were investigated by means of microotoscopy and scanning electron microscopy in 40 developing Albino rats. In 20 one nostril (group B) and in 20 both nostrils (group C) had been experimentally obstructed. 20 rats were used as controls (group A). Microotoscopy and scanning electron microscopy on the middle ear mucosa were performed when the animals were 2 months old, i.e. at 90% of their development. At microotoscopy the tympanic membranes, homolateral to the obstructed nostril, proved to be abnormal in 81% of group B rats and in almost all rats belonging to group C. Scanning electron microscopy, performed after the bullae had been removed, showed the following epithelial surface changes of the middle ear homolaterally to the obstructed nostril: (a) a significant increase of nonciliated elements with a secretory behavior as compared to controls, even of the floor where normally a large number of ciliated cells is present; (b) several ciliated cells were destroyed and their cilia were irregular and untidy; (c) thick mucous secretions covered the floor of the tympanic bulla; and (d) squamous metaplasia of the hypotympanic epithelium. Extrarotation of the eustachian tubes and developmental abnormalities of the skull base in growing rats, caused by nasal obstruction, seem to be able to induce pathological events of the middle ear.

[Changes in the middle ear mucosa of the Wistar rat after experimental resection and obstruction of the eustachian tube]. / Modificaciones en la mucosa del oído medio de la rata Wistar tras sección y obstrucción experimental de la trompa de Eustaquio.

Eustachian tube (ET) disfunction, alone or combined with other factors, is the main source of middle ear pathogenesis and can lead to other physiopathological events that originate disease (serous otitis media, adhesive otitis, cholesteatoma). In seven Wistar rats (study group), experimental mechanical obstruction of the left Eustachian tube was performed. Using an anterior cervical incision, the osteocartilaginous junction of the ET was severed and obliterated with a bit of muscle in order to prevent rechanneling. After a period of six months, the middle ear mucosa was removed for histological study. Comparisons were made of the experimental ear and the opposite ear, as well as the ears of three rats in the control group. We compared our findings with those of other authors and reviewed experimental animal models of serous otitis media and cholesteatoma that have been used to study the influence of ET in middle ear disease.

Prussak's space: chronological development and routes of aeration.

Investigation of Prussak's space and its relationship to adjacent spaces is important in elucidating the cause of retraction pocket and cholesteatoma formation in this space. This study was designed to quantitatively characterize the chronological development of Prussak's space and its relationship to adjacent spaces in temporal bones. One-hundred and forty-nine human temporal bone slides (115 normal, 28 with otitis media with effusion, three with retraction pockets and three with attic type cholesteatoma) including specimens ranging from fetal to adult bones were studied. Prussak's space was formed and sufficient aeration routes established by 4 years of age in normal temporal bones. In temporal bones with otitis media with effusion, however, the growth of Prussak's space was suppressed and few routes for aeration established until 10 years of age. In normal temporal bones, Prussak's space developed with aeration routes sufficient to avert the negative pressure which can result in retraction pocket formation in the pars flaccida of the tympanic membrane.