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D J Lim

Publications and source records attributed to D J Lim.

At least 73 records · Page 4Linked to original sources
PubMed1992

Luminal development of the eustachian tube and middle ear: murine model.

The purpose of this study was to describe the luminal development of the murine eustachian tube and middle ear. Thirty specimens, aging from gestational day 11 to postnatal day 21, were investigated through the light microscopic observations. The present study also used digitizer, computer, and serially sectioned temporal bone specimens for three-dimensional reconstruction to measure the volume of the eustachian tube and middle ear cavity at different gestational and postnatal ages. The first pharyngeal pouch elongated during gestational day 12 to form the tubotympanic recess. Between gestational day 13 and 14 this tubotympanic recess extended to the middle ear area. A rapid increment in the volume of the tubotympanic recess was noted between gestational day 15 and 16. At this age, a definite division of the tubotympanic recess into the eustachian tube and middle ear cavity was observed. During the postnatal period, the maximum change of the middle ear volume was noted on postnatal day 11 when the mesenchymal tissue in the middle ear cavity disappeared completely.

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PubMed1991

Primary culture of chinchilla middle ear epithelium.

Chinchilla middle ear epithelium was successfully cultured in medium containing Dulbecco's modified Eagle's medium, Ham's F12 mixture, and fetal bovine serum. After 3 to 5 days, the explants produced outgrowths of primarily flat polygonal and ciliated cells that persisted for up to 10 days in culture. These cells in the outgrowth often formed a "dome" indicating the presence of functional polarization and fluid transportation capability. The ciliated cells were more frequently found near the explant, and were fewer in number in the area distant from the explant. This finding suggests that the ciliated cells in the outgrowth are migrated ciliated cells deriving from the explant. That secretory cells were not identified in the outgrowth indicated that the present culture technique did not support secretory activity. Using the present culture technique, we were able to maintain the explants and primary cultured cells for up to 14 days in a majority of cases; hence, these techniques appear to be applicable to a number of in vitro studies.

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PubMed1991

Glycoconjugates in the chinchilla tubotympanum.

Various biotinylated lectins were used to characterize and semiquantitate glycoconjugate residues in the tubotympanum. Epithelial goblet cells were stained predominantly by WGA, LFA, SNA, RCA-I, Con-A, LCA, SBA, PHA-E, and UEA; this finding suggests they contain alpha-neuraminic acid, beta-galactose, alpha-mannose, N-acetyl alpha-galactosamine, and alpha-fucose. Glandular mucous cells were stained predominantly by WGA, LFA, SNA, and RCA-I; this finding suggests that they contain alpha-neuraminic acid and beta-galactose. The glandular serous cells were stained predominantly by Con-A, WGA, and LFA; this finding suggests that they produced alpha-mannose and alpha-neuraminic acid that represented serum-type glycoprotein. The positive staining of epithelial goblet cells and glandular mucous cells with PNA after neuraminidase digestion suggests that they produced mucin-type glycoproteins. The staining of the mucous blanket by WGA, LFA, SNA, RCA-I, LCA, PNA, SBA, PHA-E, and UEA suggests the presence of alpha-neuraminic acid, beta-galactose, N-acetyl alpha-galactosamine, and alpha-fucose. The epithelial cell (nonsecretory) surface was stained largely by WGA, LFA, SNA, RCA-I, Con-A, and LCA; this finding suggests the presence of alpha-neuraminic acid, beta-galactose, and alpha-mannose.

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PubMed1991

Serial culture and characterization of the chinchilla middle ear epithelium.

We have successfully cultured a fibroblast-free chinchilla middle ear epithelium up to the 10th passage by using conditioned medium or using irradiated 3T3 cells as feeder cells. The cultured epithelial cells assumed a polygonal shape with a cobblestone appearance, indicating tight junction formation. A small number of the cells began to show abnormal morphology, such as indistinct cell boundaries, fibroid appearance, or giant cell formation, as the passage increased, particularly after the 5th passage. These morphologically transformed cells showed positive labeling with an anticytokeratin antibody, which indicated the epithelial origin of these cells. Neither ciliated nor secretory cells were observed in the serially cultured cells. The rate of cell growth slowed after the 7th passage, and after the 11th passage the cells no longer proliferated. Even with the above limitations, these cultured cells can be used for a number of in vitro experiments.

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PubMed1991

Heterogeneity of glycoconjugates in the secretory cells of the chinchilla middle ear and eustachian tubal epithelia: a lectin-gold cytochemical study.

The present study was conducted to characterize and localize the glycoconjugates in the tubotympanum (auditory or eustachian tube and middle ear cavity) of chinchilla on an ultrastructural level, using lectin-gold complexes with six different lectins: BPA, ConA, RCA-1, WGA, LFA, and SNA. A comparison of the affinity of these lectins demonstrated the heterogeneity of secretory cells. The glandular serous cells and epithelial dark granulated cells produced "serum"-type glycoprotein. The glandular mucous cells and goblet cells produced dominantly "mucin"-type glycoprotein in the light granules, but "serum"-type glycoprotein in the dark cores. The labeling of LFA and SNA showed that sialic acids existed mainly in the mucinous granules of secretory cells and ciliated epithelium glycocalyx, and in the mucous blanket. The results also suggested that the dominant linkage of sialic acids of mucin is a Neu5Ac(alpha 2-6)Gal/GalNAc sequence. Furthermore, the data obtained from ConA and BPA suggested that initial O-glycosylation of mucin took place in the cis side of the Golgi apparatus and that initial N-glycosylation of the serum occurred in the rough endoplasmic reticulum.

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PubMed1991

Surface active substances in the chinchilla tubotympanum. A biochemical study.

It has been demonstrated that the eustachian tube and middle ear epithelium produce Tubal Surface Active Substances (TSAS), which facilitate the opening of the eustachian tube. In order to characterize the biochemical contents of chinchilla TSAS, the tubal washings were analyzed using 2-D thin layer chromatography. The results indicate that phosphatidylcholine was the predominant phospholipid, followed by sphingomyelin, phosphatidylinositol phosphatidylethanolamine, and phosphatidylserine. In comparison, pulmonary lavage showed phosphatidylcholine to be highest allowed by phosphatidylethanolamine and sphingomyelin. Phosphatidylcholine/phosphatidylethanolanim ratios were 5:1 in the tubal lavage, and 8:1 in the pulmonary lavage. Phosphatidylcholine/sphingomyelin ratios were 2:1 in the tubal lavage, and 67:1 in the pulmonary lavage. It is concluded that the biochemical content of TSAS is similar but not identical to that of pulmonary surfactants.

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PubMed1991

[Glycoconjugates in the chinchilla tubotympanum: a lectin histochemical study].

Glycoconjugates are essential for normal functioning of the mucociliary defence system of the tubotympanum. Recently, terminal glycosylation sequences of glycoconjugates have been recognized for their role in mediating biological recognition, such as serving as a receptor for bacterial adherence. In this study, neuraminidase and 6 different lectins: Wheat germ agglutinin (WGA), Limax flavus agglutinin (LFA), Sambucus nigra agglutinin (SNA), Peanut agglutinin (PNA), Ricinus communis agglutinin-I (RCA-I), and Concanavalin A (Con A), were used to characterize histochemically the carbohydrate structures of glycoconjugates of the chinchilla tubotympanum. WGA, LFA, SNA, and RCA-I strongly labeled epithelial goblet cells, glandular mucous cells, cell surfaces, and mucous blanket. PNA weakly labeled only a small number of epithelial goblet cells and glandular mucous cells, and did not label cell surfaces. After neuraminidase treatment, PNA labeled large number of these secretory cells and cell surfaces. Con A predominantly labeled glandular serous cells, epithelial dark granulated cells, and cell surfaces, and also labeled dark cores of mucinous granules. These results revealed that: sialomucin are produced from glandular mucous cells and epithelial goblet cells and are present on cell surfaces and within the mucous blanket; their terminal trisaccharide linkage appears to be the sequence Neu5Ac (alpha 2-6) Gal (beta 1-3) GalNAc and serum type glycoproteins are produced from glandular serous cells and dark cores of mucinous granules.

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PubMed1990

An ultrastructural study of the guinea pig tectorial membrane 'type A' protofibril.

Fine structural features of the 'type A' protofibrils in the guinea pig tectorial membrane were examined using negative staining and deep-etching techniques. Negative-stained samples of fragmented tectorial membrane were composed of several fine filamentous subunits showing the clear banding pattern of the type A protofibrils. Deep-etched replicas of the EGTA (ethylene glycol bis-N,N,N',N'-tetraacetic acid)-treated samples showed fine surface structure consisting of several linear arrays of filamentous elements on the extracellular fibrils, which is interpreted to be type A protofibrils.

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PubMed1990

Role of middle ear endotoxin in inner ear inflammatory response and hydrops: long-term study.

The permeability of the round window membrane for Salmonella typhimurium-derived endotoxin was examined with use of a total of 33 chinchillas. One milligram of each endotoxin was instilled into the tympanic cavities via the superior bullae. The endotoxin activities in middle ear effusions (MEEs), perilymph, and sera were determined by limulus amebocyte lysate assay. Endotoxin was detected in perilymph on the inoculated side by 12 hours after endotoxin instillation and persisted for up to 3 weeks. Endotoxin level peaked at 24 to 48 hours postinstillation, and it steadily declined afterward. This result suggests that the maximum penetration occurred during the active inflammatory stage. Histologic evidence demonstrated remarkable pathologic changes in the inner ear, including bleeding and inflammatory cell recruitment, mostly in the perilymphatic spaces (eg, scalae tympani, scalae vestibuli, spiral ligament), strial swelling, and sensory cell degeneration. This result suggests that endotoxin present in the middle ear can permeate the round window membrane, causing inner ear tissue damage in this animal model.

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PubMed1989

Hazard from weapons impulses: histological and electrophysiological evidence.

Current methods of rating the hazard of weapons impulses for the ear have recently been challenged by electrophysiological data from experiments with animal ears which indicate that the hazard from low-frequency impulses is much lower than the hazard from higher frequency impulses (Dancer et al., 1981; Price, 1986b). To supplement these data, histological data are reported here for 51 cats that were exposed on one occasion to either rifle or howitzer impulses at peak pressures from 145 to 155 dB or 153 to 166 dB, respectively. Histological procedures (scanning electron and light microscopy) were carried out over 2 months after the exposure and after electrophysiological measures had been made. For both types of impulse the losses tended to be in the middle of the cochlea in focused lesions, even though the spectral peaks of the acoustic stimuli had been at about 80 Hz (howitzer) and 1000 Hz (rifle). Outer hair cells were more susceptible than the inner hair cells and interindividual differences in effects were large. Furthermore, the two impulse sources were equally hazardous when the peak pressure of the rifle impulse was lower than the peak pressure of the howitzer impulse by about 9 dB. In terms of A-weighted energy, the exposures were equally hazardous when the rifle exposure contained about 35 times less energy than the howitzer exposure. The histological data are thus consistent with the electrophysiological data, which indicate that present standards for impulse noise exposure may overrate the hazard of low-frequency impulses relative to impulses in the midrange.

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PubMed1989

Immunological responsiveness of chinchillas to outer membrane and isolated fimbrial proteins of nontypeable Haemophilus influenzae.

Thin, nonhemagglutinating fimbriae have been demonstrated on 100% of the clinical isolates of nontypeable Haemophilus influenzae recovered from children with chronic otitis media tested in this laboratory (L. O. Bakaletz, B.M. Tallan, T.M. Hoepf, T.F. DeMaria, H.G. Birck, and D.J. Lim, Infect. Immun. 56:331-335, 1988). Chinchillas with induced otitis media responded to this surface-located antigen of both the infecting and a heterologous strain. Antibodies were found in both serum and middle ear fluids.

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PubMed1989

Effect of prostaglandin E2 and bacterial endotoxin on the rate of dye transport in the eustachian tube of the chinchilla.

To determine the effects of various biologic agents on the rate of fluid transport in the eustachian tube of the chinchilla, we have established an in situ method of measuring dye transport in which the bulla remains intact. The normal rate of dye transport from an injection site in the superior bulla to the nasopharyngeal orifice of the eustachian tube was 130 +/- 10 seconds. Inhibition of ciliary activity with the local anesthetic bupivacaine resulted in a saturable delay of transport (greater than 15 minutes), while exposure to the beta-adrenergic stimulator isoproterenol caused a significant increase in transport rate (79 +/- 7 seconds). Two inflammatory mediators commonly found in chronic middle ear effusions were also tested for their effect on fluid transport by the mucociliary apparatus. Bacterial endotoxin reduced transport rates at high concentrations, while prostaglandin E2 had no effect at any concentration tested.

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PubMed1989

Endotoxin permeability through the round window.

The permeability of the round window membrane to Salmonella typhimurium derived endotoxin was examined using a total of 17 chinchillas. One mg of endotoxin was instilled into the tympanic cavity via the superior bulla. Endotoxin activity in middle ear effusions (MEEs), perilymph (both inoculated and non-inoculated side), and sera was determined by Limulus lysate assay after 12, 24, 48, 72, and 120 h following endotoxin instillation. Endotoxin was detected in perilymph on the inoculated side by 12 h after endotoxin instillation and persisted for 5 days during the present measurement period. Endotoxin level peaked at 24-48 h post-instillation, and steadily declined afterwards. This result suggests that the maximum penetration occurred during the active inflammatory stage. Histologic investigation revealed marked pathological changes in the inner ear, including bleeding and inflammatory cell recruitment, mostly in the perilymphatic spaces (e.g. scalae tympani and vestibuli, spiral ligament), strial swelling, and sensory cell degeneration. These results suggest that endotoxin, when introduced into the middle ear, can permeate through the round window membrane and can cause inner ear tissue damage in this animal model.

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PubMed1988

Beta 2-microglobulin in otitis media with effusion.

beta 2-Microglobulin (beta 2M) is a low-molecular-weight protein present in serum and other fluids during various autoimmune or chronic inflammatory diseases. beta 2-Microglobulin was measured in middle ear effusion (MEE) and serum samples obtained from 36 patients with chronic otitis media with effusion. Using a quantitative competitive enzyme immunoassay, we were able to demonstrate beta 2M in 98% of the MEE samples. The mean concentration of beta 2M was higher in the MEE samples than in the serum samples. There was considerable variability between ears in those patients with bilateral MEE. There was no correlation between beta 2M concentration and the patients' age, sex, MEE type, and culture results, or cytologic profiles of the MEEs. This increased beta 2M may reflect earlier lymphocyte activity during the inflammatory process.

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PubMed1988

Frequency of fimbriation of nontypable Haemophilus influenzae and its ability to adhere to chinchilla and human respiratory epithelium.

To date, we have examined nearly 60 clinical isolates of nontypable Haemophilus influenzae (26 nasopharyngeal, 33 from middle ear effusions) and have found that 100% were fimbriated. The percentage of cells bearing fimbriae within each isolate varied from less than 10 to 100%, with fimbriae being either peritrichous or bipolar in distribution. Fimbriae were approximately 2.4 to 3.6 nm in width; however, there was a high degree of variability in both length and number of fimbriae per individual bacterial cell among these isolates. All isolates tested adhered to both human oropharyngeal cells and chinchilla tracheal epithelium regardless of the degree to which the particular isolate was fimbriate. The level or degree of fimbriation did not correlate with either site of isolation, biotype, strength of hemagglutination reaction, or type of effusion present in the ear. These appendages appear to be quite different from those described for type b H. influenzae in which the ability to adhere and strength of ability to hemagglutinate correlated strongly with degree of fimbriation.

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