Acta Oto-Laryngologica
ISSN: 0001-6489 (Print) 1651-2251 (Online) Journal homepage: http://www.tandfonline.com/loi/ioto20
Middle Ear Mucosa in Cleft Palate Children: A Scanning Electron Microscopic Study Per Møller & Helge Dalen To cite this article: Per Møller & Helge Dalen (1978) Middle Ear Mucosa in Cleft Palate Children: A Scanning Electron Microscopic Study, Acta Oto-Laryngologica, 86:sup360, 198-203, DOI: 10.3109/00016487809123516 To link to this article: http://dx.doi.org/10.3109/00016487809123516
Published online: 08 Jul 2009.
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Full Terms & Conditions of access and use can be found at http://www.tandfonline.com/action/journalInformation?journalCode=ioto20 Download by: [RMIT University Library]
Date: 02 May 2016, At: 00:45
Acta Otolaryngol, Suppl. 360: 198-203, 1979
MIDDLE EAR MUCOSA IN C L E F T PALATE CHILDREN A Scanning Electron Microscopic Study
Per Mraller and Helge Dalen From the Depurtment of Otorhinolaryngology, Hrrukeland sykehus. and Luborrrtor>~of
Clinictrl Electron Microscopy. The Medical Fncrrlty, University of Bergen, Norway
Acta Oto-Laryngologica 1978.86:198-203.
Ahstrcrct. Twenty-three promontory mucosal specimens
from cleft palate children were studied by scanning electron microscope. Comparative studies were made, using light and transmission electron microscopy. The mucosal lining was pseudostratified columnar with microvilli, a few ciliated cells with secretory granules and numerous goblet cells. A few mucous glands could be seen in certain areas. Microvilli with bulging tips and cilia with microknobs were common phenomena. The submucosa were inflamed, with leukocytes, macrophages, plasma cells and lymphocytes, indicating the presence of both cellular and humeral defence mechanisms.
Most children with cleft palate have from birth onward a serous or secretory otitis media (SOM). The main reason is, it seems, dysfunction of the Eustachian tube due to muscle incompetence. Periodically the hearing can be affected, but the long-term hearing results are favourable (Mraller, 1975). In order to study the middle ear mucosa in cleft palate children, biopsies were taken from the promontory area during myringotomy and observed with a scanning electron microscope (SEM). Our results were compared with the findings reported by Lim (1976), Shimada et al. (1972) and Hentzer (19701, describing the normal epithelium to be simple squamous, cuboidal or columnar with microvilli and ciliated cells on the promontory, indicating that even normal epithelium can differ considerably. MATERIAL AND METHODS Twenty-three cleft palate patients underwent bilateral myringotomy with general anesthesia. Biopsies were taken from the lower part of the promontory and specimens were proAcrtr Otolor\~iigo/Suppl360
cessed for scanning electron microscopy. The material consisted of 18 patients (under 2 years) with bilaterally mucoid effusions, 2 patients ( 2 4 years) with bilaterally mucoid effusions and 1 patient (2 years) with bilaterally serous effusions. One patient ( 2 months) had bilaterally dry ears and 3 patients (8-10 years) had dry ears the last 2 4 weeks prior to myringotomy. In order to remove cellular debris from the surface of the specimens, the material was briefly rinsed with physiological saline prior to fixation with 2 % glutaraldehyde and 1% OsO,. Both fixatives were made up in 0.1 M cacodylate buffer at pH 7.2 and kept ice-cold during fixation. The tissue was dehydrated in increasing concentrations of ethanol, transferred to acetone and critical point dried using CO,. After coating the specimens with a thin film of gold, using the diode-sputter technique, they were examined in a Philips (PSEM 500) scanning electron microscope operated at 20 kV. Specimens which had been studied in the SEM were embedded in Epon 812. After polymerization the blocks were cut with glass knives on a Reichert ultramicrotome. Thick sections (1-2 pm) were stained with Toluidine blue for observation in the light microscope, while ultrathin sections were stained with uranyl acetate and lead citrate according to standard techniques for correlative studies in the transmission electron microscope (TEM). In addition, biopsy samples from 3 cleft palate children (2, 7 and 11 years of age) with mucoid effusions were fixed as described
Middle ear mucosa in cleft palate children above and directly processed for TEM investigation in a Philips 300 electron microscope operated at 80 kV.
Acta Oto-Laryngologica 1978.86:198-203.
RESULTS With one exeption, all of the specimens were covered in part with a mucous blanket. This was even true in the clinically dry ears. Light microscopy revealed that the epithelium was pseudostratified columnar and the apical surface of most cells was covered with numerous microvilli. Occasional ciliated cells were also noted (Fig. 1). A 2-year-old girl had the soft palate closed 6 days prior to myringotomy. She experienced serous fluid in the middle ears for a few postoperative weeks before the ears became airfilled again. The epithelium in this case was found to be flat (Fig. 2). No sign of inflammation was found in the submucosa. Secretory doplets could be seen, typical of goblet cell secretion as described by Tos et al. (1975) and Lim (1976). Bulging of the microvilli was apparent in some areas (Fig. 3). We found both goblet cells and epithelial cells-as well as ciliated cells-to have microvilli at the surface. Some of the ciliated cells had large numbers of microknobs on the ciliar surface (Fig. 4). These structures were mainly located at the tip and upper part of the cilia. Strands from mucous glands were observed in a few places. The presence of such glands, as has been described by Tos et al. (1973), was confirmed by our correlative light microscopy and TEM studies. The submucosa were dominated by numerous large capillaries. A striking feature was the large quantities of inflammatory cells (Fig. 5 ) . In most specimens leukocytes and lymphocytes dominated the picture, along with plasma cells and macrophages.
DISCUSSION The promontory mucosa in cleft palate patients with SOM is considerably thickened,
199
with a pseudostratified columnar epithelium and with numerous inflammatory cells in the submucosa. A considerable proportion of the epithelial cells are goblet cells and a few mucous glands could be seen in our specimens. The microvilli in some areas revealed considerable bulging of their tips, which may be the morphological manifestation of secretory activity andfor fluid exchange. In some ciliated cells with a large number of microknobs covering the ciliar surface, TEM studies showed that these structures contained an electron-dense material and that the wall was an extension of the ciliar membrane. A few other knobs ballooned from the cilia and resembled more the kind described by Dahlgren et al. (1972) and others (Mecklenburg et al., 1973; Baldetop et al., 1977). Support for the view that the microknobs represent a local secretory activity is found in the appearance of numerous vesicles in the apical cytoplasm of the ciliated cells, as was also reported by Hentzer (1972). However, until the true nature of the microknobs is elucidated, the possibility remains that they may be degenerative phenomena. The number of atypical cilia seems to be high in the present material. The majority of cilia had a normal 9+2 pattern, but compound cilia were frequently observed. Such cilia have previously been found in the middle ear mucosa by Kawabata et al. (1969). One patient had transient serous fluid in the middle ear, due to impaired Eustachian tube function alone. In this patient the promontory epithelium was flat and no secretory capacity could be demonstrated. The serous fluid was most likely a transudate, as described by Gundersen et al. (1971). Interestingly enough, a 2-month-old girl had air in both middle ears, but our microscopical studies showed a high mucosal lining with a picture indistinguishable from the children with mucous-filled middle ears. Within a few months, during which no acute infection occurred (to our knowledge), the girl developed secretory otitis media in both ears. Actti
Otu/orsngo/ Srrppf 360
P . M d l e r rind H . Dalen
Acta Oto-Laryngologica 1978.86:198-203.
200
Fig. I . SEM, X 5 850. Promontory middle ear rnucosa from a cleft palate child with secretory otitis media. Numerous microvilli cover the surface ( M ) . A single ciliated cell ( C ) and the mucous blanket (S) can be seen.
Fig. 2 . SEM, x 11 700. Flat epithelium on the promontory in a cleft palate child with serous otitis media lasting a few weeks, due to soft palate closure.
Acta Oto-Laryngologica 1978.86:198-203.
Middle ear mucosa in cleft palate children
Fig. 3. SEM, x I 1 700. Promontory surface. Numerous microvilli ( M ) with tip bulging. a ciliated cell ( C ) and secretory granule ( S ) .
Fig. 4.SEM, x 1 1 700. Cilia ( C ) with microknobs.
201
Acta Oto-Laryngologica 1978.86:198-203.
202
P . M d l e r and H . Dalen
Fig. 5 . SEM, X 2 925. Submucosa near the mucosal lining. Inflammatory cells ( L ) can be seen. Microvilli ( M ) and secretory granule (S) on the surface.
It is important to note that patients selected for this material had no history of acute otitis media in the previous months and no clinical signs of acute infection at the time of myringotomy. Nevertheless all promontory specimens (except the case of the patient with transient serous fluid) showed a marked inflammatory response indicating both cellular and humoral defence systems to be intact. The high secretory capacity of the middle ear epithelium in our cases is associated with inflammation in the thickened submucosal layer.
ZUSAMMENFASSUNG Eine Studie uber die promontoriale Schleimhaut bei Kindern mit Gaumenspalte. 23 Praparate der Promontonalschleimhaut wurden mit einem Scanningelektronenmikroskop untersucht. Vergleichbdre Studien wurden transmissionsmikroskopisch und lichtmikroskopisch durchgeAcfo Ofdn,go/, S ~ i p p360 /
fiihrt. Die Schleimhaut zeigte ein hohes Zylinderepithel mit Mikrovilli und wenigen Zilien. Es wurden sekretorische Tropfchen von Becherzellen gefunden und auch einige Schleimhautdriisen. Die Mikrovilli waren ,,bulging" und die Zilien mit ,,Mikroknobs" bedeckt. In der Submucosa wurden Zeichen einer chronischen Entzundung mit Granulozyten, Plasmazellen, Lymphozyten und Makrophagen gefunden.
REFERENCES Baldeterp, L . , Mecklenburg, C. V. & Hakansson, C. H. 1977. Ultrastructural alterations in ciliary cells exposed to ionizing radiation. Cell Tiss Res 180, 421. Dahlgren, S . E., Dalen, H. & Dahlhamn, T. 1972. Ultrastructural observations on chemically induced inflammation in guinea pig trachea. Arch abt. B . Zellpathol. 11. 211. Gundersen, T. 1971. Serm otitis media. Nord Med 86, 945. Hentzer, E . 1970. Ultrastructure of the normal mucosa in the human middle ear, mastoid cavities and Eustachian tube. Ann Otol-Rhino/ Luryngol79, 1143.
Middle eur mucosa in cleft palate children
Shimada, T. & Lim, D. 3. 1972. Distribution of ciliated cells in the human middle ear. Ann OfolRhinol Laryngo181. 203. Tos, M. & Bak-Pedersen, K . 1973. Density of mucous glands in a biopsy material of chronic secretory otitis media. Acta Otolaryngol (Stockh) 75, 55. - 1975. Density of goblet cells in chronic secretory otitis media. Findings in a biopsy material. Laryngoscope 85, 377.
Per M d l e r Department of O t ~ ~ ~ h i ~ ~ ~ l a r y n ~ o l o g y 5016 Haukeland sykehus Bergen, Norway
Acta Oto-Laryngologica 1978.86:198-203.
- 1972. Ultrastructure of the middle ear mucosa in secretory otitis media. Acta Orolaryngol (Stockh) 73. 394. Kawabata. I . & Paparella, M. 1969. Atypical cilia in normal human and guinea pig middle ear rnucosa. Acra Otolaryngol (Stockh) 67, 5 1 I . Lim, D. J . 1976. Functional morphology of the mucosa of the middle ear and Eustachian tube. Ann Otol Rlrinol Laryngol85, Suppl. 25, 36-43. Maeda, S . , Mogi, G. & Motow, 0. 1976. Fine structures of the normal mucosa in developing rat middle ear. Ann Otol Rhinol Laryngol85, Suppl. 31. Mecklenburg, C. V . , Merche, U . , Hakonsson, C. H. & Toremalm, N . G. 1974. Morphological changes in ciliary cells due to heat exposure. Cell Tiss R e s 148, 45. Mnller, P. 1975. Long term otologic features of cleft palate patients. Arch Otolnryngol 101. 605.
203
Actci O/u/urviigo/Sirppl.360
ISSN: 0001-6489 (Print) 1651-2251 (Online) Journal homepage: http://www.tandfonline.com/loi/ioto20
Middle Ear Mucosa in Cleft Palate Children: A Scanning Electron Microscopic Study Per Møller & Helge Dalen To cite this article: Per Møller & Helge Dalen (1978) Middle Ear Mucosa in Cleft Palate Children: A Scanning Electron Microscopic Study, Acta Oto-Laryngologica, 86:sup360, 198-203, DOI: 10.3109/00016487809123516 To link to this article: http://dx.doi.org/10.3109/00016487809123516
Published online: 08 Jul 2009.
Submit your article to this journal
Article views: 1
View related articles
Full Terms & Conditions of access and use can be found at http://www.tandfonline.com/action/journalInformation?journalCode=ioto20 Download by: [RMIT University Library]
Date: 02 May 2016, At: 00:45
Acta Otolaryngol, Suppl. 360: 198-203, 1979
MIDDLE EAR MUCOSA IN C L E F T PALATE CHILDREN A Scanning Electron Microscopic Study
Per Mraller and Helge Dalen From the Depurtment of Otorhinolaryngology, Hrrukeland sykehus. and Luborrrtor>~of
Clinictrl Electron Microscopy. The Medical Fncrrlty, University of Bergen, Norway
Acta Oto-Laryngologica 1978.86:198-203.
Ahstrcrct. Twenty-three promontory mucosal specimens
from cleft palate children were studied by scanning electron microscope. Comparative studies were made, using light and transmission electron microscopy. The mucosal lining was pseudostratified columnar with microvilli, a few ciliated cells with secretory granules and numerous goblet cells. A few mucous glands could be seen in certain areas. Microvilli with bulging tips and cilia with microknobs were common phenomena. The submucosa were inflamed, with leukocytes, macrophages, plasma cells and lymphocytes, indicating the presence of both cellular and humeral defence mechanisms.
Most children with cleft palate have from birth onward a serous or secretory otitis media (SOM). The main reason is, it seems, dysfunction of the Eustachian tube due to muscle incompetence. Periodically the hearing can be affected, but the long-term hearing results are favourable (Mraller, 1975). In order to study the middle ear mucosa in cleft palate children, biopsies were taken from the promontory area during myringotomy and observed with a scanning electron microscope (SEM). Our results were compared with the findings reported by Lim (1976), Shimada et al. (1972) and Hentzer (19701, describing the normal epithelium to be simple squamous, cuboidal or columnar with microvilli and ciliated cells on the promontory, indicating that even normal epithelium can differ considerably. MATERIAL AND METHODS Twenty-three cleft palate patients underwent bilateral myringotomy with general anesthesia. Biopsies were taken from the lower part of the promontory and specimens were proAcrtr Otolor\~iigo/Suppl360
cessed for scanning electron microscopy. The material consisted of 18 patients (under 2 years) with bilaterally mucoid effusions, 2 patients ( 2 4 years) with bilaterally mucoid effusions and 1 patient (2 years) with bilaterally serous effusions. One patient ( 2 months) had bilaterally dry ears and 3 patients (8-10 years) had dry ears the last 2 4 weeks prior to myringotomy. In order to remove cellular debris from the surface of the specimens, the material was briefly rinsed with physiological saline prior to fixation with 2 % glutaraldehyde and 1% OsO,. Both fixatives were made up in 0.1 M cacodylate buffer at pH 7.2 and kept ice-cold during fixation. The tissue was dehydrated in increasing concentrations of ethanol, transferred to acetone and critical point dried using CO,. After coating the specimens with a thin film of gold, using the diode-sputter technique, they were examined in a Philips (PSEM 500) scanning electron microscope operated at 20 kV. Specimens which had been studied in the SEM were embedded in Epon 812. After polymerization the blocks were cut with glass knives on a Reichert ultramicrotome. Thick sections (1-2 pm) were stained with Toluidine blue for observation in the light microscope, while ultrathin sections were stained with uranyl acetate and lead citrate according to standard techniques for correlative studies in the transmission electron microscope (TEM). In addition, biopsy samples from 3 cleft palate children (2, 7 and 11 years of age) with mucoid effusions were fixed as described
Middle ear mucosa in cleft palate children above and directly processed for TEM investigation in a Philips 300 electron microscope operated at 80 kV.
Acta Oto-Laryngologica 1978.86:198-203.
RESULTS With one exeption, all of the specimens were covered in part with a mucous blanket. This was even true in the clinically dry ears. Light microscopy revealed that the epithelium was pseudostratified columnar and the apical surface of most cells was covered with numerous microvilli. Occasional ciliated cells were also noted (Fig. 1). A 2-year-old girl had the soft palate closed 6 days prior to myringotomy. She experienced serous fluid in the middle ears for a few postoperative weeks before the ears became airfilled again. The epithelium in this case was found to be flat (Fig. 2). No sign of inflammation was found in the submucosa. Secretory doplets could be seen, typical of goblet cell secretion as described by Tos et al. (1975) and Lim (1976). Bulging of the microvilli was apparent in some areas (Fig. 3). We found both goblet cells and epithelial cells-as well as ciliated cells-to have microvilli at the surface. Some of the ciliated cells had large numbers of microknobs on the ciliar surface (Fig. 4). These structures were mainly located at the tip and upper part of the cilia. Strands from mucous glands were observed in a few places. The presence of such glands, as has been described by Tos et al. (1973), was confirmed by our correlative light microscopy and TEM studies. The submucosa were dominated by numerous large capillaries. A striking feature was the large quantities of inflammatory cells (Fig. 5 ) . In most specimens leukocytes and lymphocytes dominated the picture, along with plasma cells and macrophages.
DISCUSSION The promontory mucosa in cleft palate patients with SOM is considerably thickened,
199
with a pseudostratified columnar epithelium and with numerous inflammatory cells in the submucosa. A considerable proportion of the epithelial cells are goblet cells and a few mucous glands could be seen in our specimens. The microvilli in some areas revealed considerable bulging of their tips, which may be the morphological manifestation of secretory activity andfor fluid exchange. In some ciliated cells with a large number of microknobs covering the ciliar surface, TEM studies showed that these structures contained an electron-dense material and that the wall was an extension of the ciliar membrane. A few other knobs ballooned from the cilia and resembled more the kind described by Dahlgren et al. (1972) and others (Mecklenburg et al., 1973; Baldetop et al., 1977). Support for the view that the microknobs represent a local secretory activity is found in the appearance of numerous vesicles in the apical cytoplasm of the ciliated cells, as was also reported by Hentzer (1972). However, until the true nature of the microknobs is elucidated, the possibility remains that they may be degenerative phenomena. The number of atypical cilia seems to be high in the present material. The majority of cilia had a normal 9+2 pattern, but compound cilia were frequently observed. Such cilia have previously been found in the middle ear mucosa by Kawabata et al. (1969). One patient had transient serous fluid in the middle ear, due to impaired Eustachian tube function alone. In this patient the promontory epithelium was flat and no secretory capacity could be demonstrated. The serous fluid was most likely a transudate, as described by Gundersen et al. (1971). Interestingly enough, a 2-month-old girl had air in both middle ears, but our microscopical studies showed a high mucosal lining with a picture indistinguishable from the children with mucous-filled middle ears. Within a few months, during which no acute infection occurred (to our knowledge), the girl developed secretory otitis media in both ears. Actti
Otu/orsngo/ Srrppf 360
P . M d l e r rind H . Dalen
Acta Oto-Laryngologica 1978.86:198-203.
200
Fig. I . SEM, X 5 850. Promontory middle ear rnucosa from a cleft palate child with secretory otitis media. Numerous microvilli cover the surface ( M ) . A single ciliated cell ( C ) and the mucous blanket (S) can be seen.
Fig. 2 . SEM, x 11 700. Flat epithelium on the promontory in a cleft palate child with serous otitis media lasting a few weeks, due to soft palate closure.
Acta Oto-Laryngologica 1978.86:198-203.
Middle ear mucosa in cleft palate children
Fig. 3. SEM, x I 1 700. Promontory surface. Numerous microvilli ( M ) with tip bulging. a ciliated cell ( C ) and secretory granule ( S ) .
Fig. 4.SEM, x 1 1 700. Cilia ( C ) with microknobs.
201
Acta Oto-Laryngologica 1978.86:198-203.
202
P . M d l e r and H . Dalen
Fig. 5 . SEM, X 2 925. Submucosa near the mucosal lining. Inflammatory cells ( L ) can be seen. Microvilli ( M ) and secretory granule (S) on the surface.
It is important to note that patients selected for this material had no history of acute otitis media in the previous months and no clinical signs of acute infection at the time of myringotomy. Nevertheless all promontory specimens (except the case of the patient with transient serous fluid) showed a marked inflammatory response indicating both cellular and humoral defence systems to be intact. The high secretory capacity of the middle ear epithelium in our cases is associated with inflammation in the thickened submucosal layer.
ZUSAMMENFASSUNG Eine Studie uber die promontoriale Schleimhaut bei Kindern mit Gaumenspalte. 23 Praparate der Promontonalschleimhaut wurden mit einem Scanningelektronenmikroskop untersucht. Vergleichbdre Studien wurden transmissionsmikroskopisch und lichtmikroskopisch durchgeAcfo Ofdn,go/, S ~ i p p360 /
fiihrt. Die Schleimhaut zeigte ein hohes Zylinderepithel mit Mikrovilli und wenigen Zilien. Es wurden sekretorische Tropfchen von Becherzellen gefunden und auch einige Schleimhautdriisen. Die Mikrovilli waren ,,bulging" und die Zilien mit ,,Mikroknobs" bedeckt. In der Submucosa wurden Zeichen einer chronischen Entzundung mit Granulozyten, Plasmazellen, Lymphozyten und Makrophagen gefunden.
REFERENCES Baldeterp, L . , Mecklenburg, C. V. & Hakansson, C. H. 1977. Ultrastructural alterations in ciliary cells exposed to ionizing radiation. Cell Tiss Res 180, 421. Dahlgren, S . E., Dalen, H. & Dahlhamn, T. 1972. Ultrastructural observations on chemically induced inflammation in guinea pig trachea. Arch abt. B . Zellpathol. 11. 211. Gundersen, T. 1971. Serm otitis media. Nord Med 86, 945. Hentzer, E . 1970. Ultrastructure of the normal mucosa in the human middle ear, mastoid cavities and Eustachian tube. Ann Otol-Rhino/ Luryngol79, 1143.
Middle eur mucosa in cleft palate children
Shimada, T. & Lim, D. 3. 1972. Distribution of ciliated cells in the human middle ear. Ann OfolRhinol Laryngo181. 203. Tos, M. & Bak-Pedersen, K . 1973. Density of mucous glands in a biopsy material of chronic secretory otitis media. Acta Otolaryngol (Stockh) 75, 55. - 1975. Density of goblet cells in chronic secretory otitis media. Findings in a biopsy material. Laryngoscope 85, 377.
Per M d l e r Department of O t ~ ~ ~ h i ~ ~ ~ l a r y n ~ o l o g y 5016 Haukeland sykehus Bergen, Norway
Acta Oto-Laryngologica 1978.86:198-203.
- 1972. Ultrastructure of the middle ear mucosa in secretory otitis media. Acta Orolaryngol (Stockh) 73. 394. Kawabata. I . & Paparella, M. 1969. Atypical cilia in normal human and guinea pig middle ear rnucosa. Acra Otolaryngol (Stockh) 67, 5 1 I . Lim, D. J . 1976. Functional morphology of the mucosa of the middle ear and Eustachian tube. Ann Otol Rlrinol Laryngol85, Suppl. 25, 36-43. Maeda, S . , Mogi, G. & Motow, 0. 1976. Fine structures of the normal mucosa in developing rat middle ear. Ann Otol Rhinol Laryngol85, Suppl. 31. Mecklenburg, C. V . , Merche, U . , Hakonsson, C. H. & Toremalm, N . G. 1974. Morphological changes in ciliary cells due to heat exposure. Cell Tiss R e s 148, 45. Mnller, P. 1975. Long term otologic features of cleft palate patients. Arch Otolnryngol 101. 605.
203
Actci O/u/urviigo/Sirppl.360
