Epithelial cells of nasal mucosa express functional gap junctions of connexin 43

OBJECTIVE: Recent studies have suggested that inositol triphosphate is transmitted through the gap junction of adjacent epithelial cells and communicates the intracellular calcium wave that controls the beating of cilia. Therefore, gap junction-mediated intercellular communication may modulate the metachronal ciliary beating of respiratory mucosa. MATERIAL AND METHODS: The expression and function of gap junctions were evaluated in nasal mucosa using immunofluorescent staining, Western blotting and Lucifer Yellow dye coupling in both whole nasal mucosa and cultured nasal epithelial cells. RESULTS: Among connexins (Cxs) 26, 32 and 43, only Cx43 was found in the nasal mucosa, in both the epithelial cells and submucosa tissue. Using Western blotting, Cx43 protein was found in protein extracts of both whole mucosa and cultured epithelial cells. The Cx43 protein content increased twofold between the 3rd and 6th days of culture. Lucifer Yellow dye, microinjected into single cultured epithelial cells, had spread to the adjacent cells. CONCLUSION: Cx43 is present in nasal mucosa. The expression of Cx43 on epithelial cells increases after culture. The dye-coupling study in epithelial cells indicated that the signal may be transmitted to neighboring cells via a gap junction composed of Cx43. Thus, Cx43 may be involved in the regulation of metachronal ciliary beating in ciliated epithelial cells and growth or differentiation in nasal tissue.     

Epithelial Cells of Nasal Mucosa Express Functional Gap Junctions of Connexin 43

Objective—Recent studies have suggested that inositol triphosphate is transmitted through the gap junction of adjacent epithelial cells and communicates the intracellular calcium wave that controls the beating of cilia. Therefore, gap junction-mediated intercellular communication may modulate the metachronal ciliary beating of respiratory mucosa. Material and Methods—The expression and function of gap junctions were evaluated in nasal mucosa using immunofluorescent staining, Western blotting and Lucifer Yellow dye coupling in both whole nasal mucosa and cultured nasal epithelial cells. Results—Among connexins (Cxs) 26, 32 and 43, only Cx43 was found in the nasal mucosa, in both the epithelial cells and submucosa tissue. Using Western blotting, Cx43 protein was found in protein extracts of both whole mucosa and cultured epithelial cells. The Cx43 protein content increased twofold between the 3rd and 6th days of culture. Lucifer Yellow dye, microinjected into single cultured epithelial cells, had spread to the adjacent cells. Conclusions—Cx43 is present in nasal mucosa. The expression of Cx43 on epithelial cells increases after culture. The dye-coupling study in epithelial cells indicated that the signal may be transmitted to neighboring cells via a gap junction composed of Cx43. Thus, Cx43 may be involved in the regulation of metachronal ciliary beating in ciliated epithelial cells and growth or differentiation in nasal tissue.     

Expression of the the cyclin-kinase inhibitors p21(WAF1) and p27(Kip1) and the p53 tumor suppressor genes in adult-onset laryngeal papillomas

Different types of human papilloma virus are known to be closely associated with laryngeal papillomas. On the other hand, the proliferation of epithelial cells is associated with various abnormalities in the mechanisms of cellular regulation. In this study, we detected the expressions of p53, p21 and p27 proteins in adult-onset laryngeal papillomas by immunohistochemical techniques. The objective of this study is to evaluate the expression of these factors in adult-onset laryngeal papillomas and to determine whether such expression is correlated with the existence of dysplastic epithelium covering the papillomas. Eighteen patients with adult-onset papillomas who were surgically treated at the Department of Otolaryngology at the University of Istanbul between January 1994 and December 1999 were included in this study. Anti-p21, -p27 and -p53 antibodies were used to perform immunostaining. Positive nuclear staining for p21 was detected in 14 of the 18 (78%) cases, especially in the parabasal layer. Also, in 78% of the cases, weak to strong immunoreactivity was observed for p27. In all cases, negative immunoreactivity was observed for p53 throughout the epithelium except for the basal and parabasal cells. A negative correlation was observed between the existence of dysplastic epithelium and p21 expression (P=0.02). In conclusion, variable p21 and p27 expression was detected by immunohistochemistry in our series of 18 cases of adult-onset laryngeal papillomatosis, and a statistically significant inverse correlation was detected between p21 expression and the existence of dysplastic epithelium covering the papillomas. Further prospective studies are warranted to determine the prognostic values of these variables and to evaluate their role in the pathogenesis of adult-onset laryngeal papillomas.     

缝隙连接蛋白43和E-钙黏附素表达与喉癌生物学行为关系的研究

OBJECTIVE: To investigate expression of connexin43 (Cx43) and E-cadherin (E-cad) and its relationship with the biological behavior in Laryngeal carcinomas. METHODS: Immunohistochemistry staining (PicTure two steps method) was used to detect connexin 43 and E-cadherin expression in 60 paraffin-embedded laryngeal carcinomas and 10 normal epithelia around tumors. All the patients were followed up more than five years. RESULTS: Cx43 and E-cad were expressed by normal epithelia with typical membranous staining 10 normal epithelia strongly expressed Cx43 and E-cad. The reduction rate of Cx43 expression in laryngeal carcinomas was 30.0%. The significant relationship was observed between low Cx43 expression and differentiation, lymph node metastasis and recurrence (chi2 = 15.09, 6.41, 3.86, P < 0.01, 0.05, 0.05), but there was no significance between low Cx43 expression and prognosis (chi2 = 2.65, P > 0.05). 41.7% laryngeal carcinoma showed reduced or no expression of E-cad. Tumor cell dedifferentiation correlated with reduced expression for E-cad (chi2 = 15.07, P < 0.01). Absent or low E-cad expression was observed more frequently in patients with local recurrence and lymph node metastasis and with less than 5-year survival period (chi2 = 5.35, 6.65, 5.14; P < 0.05, 0.01, 0.05). There was a positive relationship between the expression of Cx43 and E-cad in same sample (r = 0.63, P < 0.0001). CONCLUSIONS: Laryngeal cancer presented inactivation of Cx43 gene and E-cad gene and down regulation of Cx43 and E-cad proteins. The level of Cx43 and E-cad may be a sensitive predictor of differentiation, invasion, lymph node metastasis and recurrence. E-cad is also a prognostic factor for patients with laryngeal cancer.     

Green tea prevents down-regulation of gap junction intercellular communication in human keratinocytes treated with PMA

It has been suggested that connexin (Cx) gap junction proteins act as tumor suppressors and green tea has a potential to prevent tumor development, however, the studies on their association with human keratinocytes were rare. We evaluated the effects of a tumor promoter, phorbol-12-myristate-13-acetate (PMA), on the expression of Cxs and gap junction intercellular communication (GJIC) in human keratinocytes (HaCaT cells) and explored the preventive effects of green tea extracts-epicatechin (EC) and epigallocatechin-3-gallate (EGCG). We performed neutral red dye uptake assay to determine the optimal concentrations of PMA, EC, and EGCG for this study and confirmed the expression of Cx mRNAs using RT-PCR. We evaluated GJIC quantitatively using the ‘scrape-loading dye transfer (SLDT)’ technique after 24-h culture of HaCaT cells treated with agents. To analyze the expression change of Cxs, we also performed Western blot and immunocytochemistry. HaCaT cells were found to express Cx26, Cx30, Cx31, and Cx43, but not Cx29. In ‘scrape-loading dye transfer’ for functional study for GJIC, EC and EGCG significantly prevented PMA-induced down-regulation of GJIC. Western blot analyses revealed that EC and EGCG prevented down-regulation of Cx26 and Cx43 proteins in HaCaT cells treated with PMA. Immunocytochemistry showed decreased expression and abnormal location of Cx26 and Cx43 in HaCaT cells when treated with PMA, and EC and EGCG inhibited its effect. These results suggest an important role of GJIC played in carcinogenesis involving human keratinocytes and green tea as a useful anticancer diet.     

Cellular retinol-binding protein in mucosa, in benign and malignant non-squamous cell tumours of the head and neck

The measurement and localisation of cellular retinol-binding protein (CRBP), in samples of normal oral mucosa, larynx papilloma and malignant lymphoma of the oropharynx, was performed with a radioimmunoassay and immunolocalisation techniques. As compared with CRBP concentration in normal mucosa, those in laryngeal papilloma were significantly higher, but those of malignant lymphoma were similar. CRBP concentrations were highest in maturing keratinocytes within the prickle cell layers of normal mucosa and in laryngeal papillomas, as estimated on the basis of immunoreactivity to CRBP. The retinyl palmitate concentrations in extracts of oral mucosa correlated to the retinol concentrations, both in plasma and mucosal extracts, but not to the CRBP content in mucosal extracts. The immunolocalisation of a cellular retinoic acid-binding protein (CRABP) like antigen in normal oral mucosa showed much the same picture with strongest staining of the maturing keratinocytes of the prickle cell layers.     

Involucrin in laryngeal dysplasia. A marker for differentiation

Involucrin is a major structural subunit of the cross-linked protein envelope that encases keratin in maturing squamous cells. Intracytoplasmic involucrin is identifiable via immunoperoxidase techniques as these cells migrate from the basal layer to the more superficial layers of the stratified epithelium. Normal squamous epithelia and mildly dysplastic epithelia show uniform staining in the suprabasal and superficial layers of the mucosa but show no staining in the basal layer. Moderate to severe dysplasias and invasive carcinomas demonstrate irregular or focal staining in all three layers. Thirty-three microscopic samples from 27 glottic laryngeal biopsy specimens were reviewed. The histochemically abnormal differentiation identified via involucrin staining correlated with accepted histologic criteria for dysplasia. Involucrin staining may provide objective information to assist the pathologist in differentiating degrees of dysplasia in laryngeal biopsy specimens.     

内耳间隙连接和连接蛋白基因家族与遗传性聋

耳蜗有非感觉上皮细胞和结缔组织细胞2套间隙连接系统,内耳前庭的感觉上皮也有间隙连接。已发现在内耳表达的连接蛋白有Cx26、Cx30、Cx31、Cx32、Cx43和Cx45。连接蛋白是同一基因家族编码,其中GJB2(Cx26)、GJB3(Cx31)、GJB6(Cx30)、GJA1(Cx43)、GJB1(Cx32)都与遗传性聋有关。GJB2基因敲除小鼠在胚胎期致死。GJB3敲除小鼠60%在胚胎死亡,成活小鼠无耳聋和皮肤疾病等表型。GJA1裸鼠表现心脏发育缺陷与膜内和软骨内成骨延迟。GJB1敲除小鼠表型无明显异常,但外周神经表现脱髓鞘改变。不同间隙连接的缺陷导致听力下降的机制可能不同。间隙连接与耳聋关系的深入研究,可能有助于阐明听觉和耳聋的部分机制。     

Lactate dehydrogenase activity in laryngeal carcinoma and surrounding mucosa

The total lactate dehydrogenase (LDH) activity and LDH isoenzyme pattern were measured in tissue taken from laryngeal cancers and from the laryngeal mucosa both close to and distant from the tumour margin. An increase in the total LDH activity and a cathodic shift in the isoenzyme pattern were found in tumour tissue compared to the close and distant mucosa. However, significant LDH alterations in the mucosa close to the tumour were shown in comparison to the distant mucosa and to the laryngeal mucosa of controls. Histochemical studies showed the LDH activity to be located in the basal and parabasal layers of the epithelium and in the cancer foci.     

Study on ceramide expression and DNA content in patients with healthy mucosa, leukoplakia, and carcinoma of the larynx

OBJECTIVE: To investigate the expression of ceramide produced by sphingomyelin and DNA content in patients with healthy laryngeal mucosa, leukoplakia, and laryngeal carcinoma. DESIGN: A retrospective review of the clinical and surgical records of 178 patients with leukoplakia of the larynx; 23 of them developed laryngeal carcinoma. SETTING: University medical center. PATIENTS: One hundred seventy-eight consecutive patients with leukoplakia of the larynx were identified from the archived pathology files of the Eye Ear Nose and Throat Hospital of Fudan University from January 1, 1990, to December 30, 2001. Among them, 23 developed laryngeal carcinoma. Flow cytometry and immunohistochemistry were performed to test DNA content and ceramide expression in healthy tissue, tissue with leukoplakia, and tissue with laryngeal carcinoma from the same patient. RESULTS: Among 23 patients with leukoplakia, 20 had aneuploidy and 3 had diploidy. The healthy tissues were all diploids, and the tissues with laryngeal carcinoma were all aneuploids. The expression of ceramide decreased gradually from healthy tissue to tissue with leukoplakia to tissue with laryngeal carcinoma (0, no staining; 1+, weak staining; 2+, mild staining; 3+, moderate staining; 4+, strong staining; and 5+, the highest staining intensity). The expression of ceramide in DNA diploid cells is stronger than in aneuploid cells. CONCLUSIONS: Ceramide, the second messenger in apoptosis, may associate with the progress of leukoplakia to carcinoma of the larynx. The reduction of ceramide may contribute to laryngeal carcinogenesis.     

Regulators of Proliferation and Apoptosis in Carcinoma of the Larynx

Expression of interrelated gene products regulating cell proliferation and apoptosis may be disordered in squamous cell carcinoma (SCC) of the larynx compared with normal squamous mucosa. Certain of these abnormalities, alone or in combination, may be of prognostic significance in low-stage carcinomas of the larynx. A retrospective study of archival material was made. Expression of the Bcl-2 family of apoptosis-related genes (bcl-2, bcl-X, mcl-1, and bax) and the proliferation- and apoptosis-related genes p53 and cyclin D-1 were determined in 40 low-T-stage laryngeal carcinomas and in uvular epithelium from patients without SCC. Among the antiapoptotic members of the Bcl-2 family, Bcl-X and Mcl-1 showed more intense and widespread staining than Bcl-2 itself in both normal squamous mucosa and SCC. The well-ordered expression patterns of Bcl-2-related proteins found in normal epithelium were lost in SCC, and patterns of expression varied widely among individual tumors. Also, mean expression levels for Bax and cyclin D-1 were significantly lower than in normal epithelium (P = .036 and P = .009, respectively), whereas expression of p53 was higher in tumors (P = .034). Expression of Bcl-X and Mcl-1 was greater in poorly differentiated than in well-differentiated tumors (P = .014 and P = .031, respectively). No associations were seen between marker expression patterns and clinical outcome in this group of patients. Bcl-x and Mcl-1 appear to be the most abundantly expressed antiapoptotic proteins of the Bcl-2 family in both normal squamous mucosa and SCC of the larynx. Multiple genes regulating proliferation and apoptosis are expressed abnormally in laryngeal SCC compared with normal epithelium. In particular, loss or measurable decrease in expression of the proapoptotic protein Bax in tumors may contribute to the deranged growth control of SCC. Further study is needed to evaluate the prognostic significance of particular patterns of disordered expression of proteins regulating proliferation and apoptosis in SCC of different head and neck sites.     

c-myc癌基因蛋白在喉癌及癌旁不同距离组织中的表达研究

应用枸橼酸－ＬＳＡＢ－微波免疫组化法，检测ｃ－ｍｙｃ癌基因蛋白在喉癌、边缘区及癌旁０．５、１．０、１．５、２．０ｃｍ处粘膜和４例正常喉粘膜中的表达。结果：ｃ－ｍｙｃ蛋白在喉癌组织中的阳性表达率为９６．７％（２９／３０），与正常组织相比有极显著性差异（Ｐ〈０．０１）；在癌旁２．０ｃｍ处与１．５ｃｍ以内组织的表达存在着显著性差异（Ｐ〈０．０５）。ｃ－ｍｙｃ表达依正常粘膜→癌旁组织→癌呈逐步递增趋势。ｃ     

Electron microscopic study of intercellular junction in nasal mucosa of nasal allergy by lectin histochemistry

To explain of the mechanism of the enhanced nasal epithelial permeability to HRP in patients with nasal allergy, the inferior turbinate mucosa was removed from 6 normal adults and 7 adults with nasal allergy. Difference of the fine structure of the intercellular junction was compared between normal mucosa and mucosa of nasal allergy by electron microscope. Staining pattern of four kinds of HRP-conjugated lectin (HRP-WGA, PNA, UEA-I and RCA-I) was also studied by electron microscope. There was no significant difference in the intercellular space of the mucosa between the normal mucosa and mucosa of nasal allergy. In the epithelial cell membrane, pattern of HRP-lectin staining was almost similar in both groups. In normal nasal epithelium, the intercellular junction consisted of junctional complex; adherent junction, desmosome and gap junction. The intercellular space was approximately 150-250 A in width. The tight junction was located beneath the luminal surface of the epithelium, and belt-like continuation connecting the adjacent cells. It was concluded that enhanced permeability to HRP in nasal allergy was not morphologic changes of the intercellular junction and component and distribution of the glycoconjugates in epithelial cellular membrane, but this may be based on functional changes.     

细胞周期调控因子与喉癌癌变的相关性研究

目的 :探讨喉癌癌变过程中 Cyclin E、p2 1WAF1 /CIP1和 p5 3表达的临床病理学意义。方法 :用免疫组化检测 2 0例喉正常粘膜、40例喉不典型增生病变和 6 0例喉癌组织中 Cyclin E、p2 1WAF 1 /CIP1和 p5 3的表达。结果 :1Cyclin E阳性表达率在正常组、不典型增生组和喉癌组分别为 5 .0 % (1/ 2 0 )、2 0 .0 % (8/ 40 )、45 .0 % (2 7/ 6 0 ) ,喉癌组与正常组比较 ,其差异有极显著性意义 (P<0 .0 1) ;p2 1WAF1 /CIP1阳性表达率分别为 95 .0 % (19/ 2 0 )、75 .0 % (30 /40 )和 6 3.3% (38/ 6 0 ) ,喉癌组与正常组比较 ,其差异亦有显著性意义 (P<0 .0 5 ) ;p5 3阳性表达率分别为 0 (0 /2 0 )、30 .0 % (12 / 40 )和 6 1.7% (37/ 6 0 ) ,喉癌组与正常组比较 ,其差异有极显著性意义 (P <0 .0 1)。 2 p2 1WAF 1 /CIP1在高、中、低分化的喉癌中阳性表达率逐渐降低 ,其差异有显著性意义 (P <0 .0 5 )。 3 Cyclin E和 p5 3阳性表达显著相关 ,p2 1WAF 1 /CIP1与 p5 3阳性表达无相关性。结论 :1Cyclin E和 p5 3异常表达是喉癌发生中早期分子事件。 2p2 1WAF 1 /CIP1 表达与喉癌细胞分化程度有关。 3细胞周期因子相互调控机制在喉癌发展中起重要作用。     

Immunohistochemical detection of epidermal growth factor receptor in laryngeal squamous cell carcinomas.

Laryngeal squamous cell carcinomas from 15 consecutive preoperatively irradiated patients were investigated for the expression of epidermal growth factor receptor (EGF receptor). The study was performed on frozen sections by means of the 5-layer APAAP technique employing an antibody recognizing the extracellular part of the EGF receptor. In sections from 9 of the patients with laryngeal squamous cell carcinoma, normal differentiated epithelia were included. Sections from 6 of these patients, in addition, contained dysplastic epithelia. Expression of EGF receptor-like material was demonstrated in the basal cell layer of normally differentiated laryngeal epithelial and in dysplastic epithelia. Fourteen of the squamous cell carcinomas proved EGF receptor positive. Nearly all cells in the poorly differentiated carcinomas showed positive staining with the antibodies. In moderately to well differentiated carcinomas a reduction in the extent of staining was seen in certain areas. Especially for the epithelial pearls, the staining reaction was localized to the undifferentiated cells in the periphery. This finding corresponds to the staining pattern observed in the basal cell layers of normal epithelial. The present investigation confirms the expression of EGF receptor-like material in normal laryngeal epithelial, dysplastic epithelial and squamous cell carcinoma. The staining pattern was similar to that observed in oral squamous cell carcinomas, predominantly varying inversely with cellular differentiation.     

Unique expression of connexins in the human cochlea

Mutations in the genes GJB2 and GJB6, which encode the proteins Connexin 26 (Cx26) and Connexin 30 (Cx30), have been linked to nonsyndromic prelingual deafness in humans. These proteins may form so-called gap junctions (GJ) or transcellular pathways between cells. The pathogenesis of deafness due to GJ Connexin mutations remains unclear partly because examinations performed in the human ear are infrequent. Here we analysed the expression and distribution of Cx26 and Cx30 in five fresh normal human cochleae taken out at occasional surgery. Immunohistochemistry including confocal microscopy in decalcified specimen showed that these proteins are widely expressed in the human cochlea. In the lateral wall there was strong antibody co-labeling for Cx26 and Cx30 that support the existence of channels comprising heteromeric Cx26/Cx30 connexons. In the organ of Corti there were some co-labeling in the supporting cell area including mainly the Claudius cells and Deiter cells of these two Cxs, apart from isolated Cx26 and Cx30 labeling in the same area, suggestive of both homomeric/homotypic pattern and hybrid pattern (heteromeric or heterotypic). Cx30, Cx26 and Connexin 36 (Cx36) immunoreactivity was also associated with spiral ganglion type I neurons, the latter being a gap junction protein specific to neurons. Gap–junction-based electrical synapses are not known to occur in mammalian auditory system other than in bats where they may play a role for fast electrical nerve transmission useful for echolocation. Their potential role in the processing of human auditory nerve signaling as well as non-GJ roles of the connexins in human cochlea is discussed.     

Decreased connexin 43 expression correlated with eosinophil infiltration in nasal polyps

BACKGROUND: A nasal polyp usually is characterized by eosinophil infiltration. Eosinophil-fibroblast interaction is an important event of persistent inflammation in airways. We have found abundant connexin 43 (Cx43) expression in subepithelial fibroblasts of nasal mucosa. Thus, we aim to analyze the relationship of Cx43 expression and eosinophil in nasal polyps. METHODS: In 25 nasal polyps and 19 inferior turbinates, indirect immunofluorescent and hematoxylin and eosin staining were performed in adjacent sections. We calculated the density of Cx43 staining and eosinophil individually by fluorescent and light microscope. RESULTS: Positive Cx43 staining under confocal microscope was shown as punctated spots on cell margin. The density of Cx43 and eosinophil staining was significantly different between groups of inferior turbinate and nasal polyp (p = 0.01 and 0.03, respectively). Decreased Cx43 expression in the subepithelial fibroblast was correlated with eosinophil infiltration in nasal polyps. Spearman rank order coefficient was equal to -0.43 (p < 0.05). CONCLUSION: This is the first demonstration of decreased Cx43 expression related to eosinophil infiltration. To the best of our knowledge, interleukin-8 may be a link between Cx43 and eosinophil and orchestrating both in developing nasal polyps.     

Cell biology of laryngeal epithelial defenses in health and disease: preliminary studies.

Esophageal epithelium has intrinsic antireflux defenses, including carbonic anhydrases (CAs I to IV) that appear to be protective against gastric reflux. This study aimed to investigate the expression and distribution of CA isoenzymes in laryngeal epithelium. Laryngeal biopsy specimens collected from the vocal fold and interarytenoid regions were analyzed by Western blotting and immunofluorescence. Carbonic anhydrases I and II were expressed by the majority of samples analyzed. In contrast, CA III was differentially expressed in the interarytenoid samples and was not detected in any vocal fold samples. The expression of CA III was increased in esophagitis as compared to normal esophageal tissue. Carbonic anhydrase I and III isoenzymes were distributed cytoplasmically in the basal and lower prickle cell layers. The laryngeal epithelium expresses some CA isoenzymes and has the potential to protect itself against laryngopharyngeal reflux. Laryngeal tissue may be more sensitive to injury due to reflux damage than the esophageal mucosa because of different responses of CA isoenzymes.     

Acute laryngotracheitis in the rat induced by Sendai virus: the influx of six different types of immunocompetent cells into the laryngeal mucosa differs strongly between the subglottic and the glottic compartment

OBJECTIVES: Acute laryngotracheitis is a disease in which mainly the subglottic area is infected, whereas adjacent parts of the larynx, especially the narrow glottic fold, remain unaffected. The reason for the difference between these two directly adjacent regions is unknown. Therefore, in the present study the influx of dendritic cells, neutrophils, T and B lymphocytes, natural killer cells, and macrophages into the mucosa of different laryngeal compartments was investigated after Sendai virus infection in the rat. The aims were to study both the influx of immunocompetent cells and the adhesion of the pathogen and to correlate them to the different reactions of the laryngeal areas during pseudocroup. METHODS: Acute laryngotracheitis was induced by intranasal application of Sendai virus in brown Norway rats. This virus is exclusively pneumotropic in rodents and belongs to the parainfluenza virus type 1, the main pathogen of acute laryngotracheitis in children. The numbers of dendritic cells, neutrophils, T and B lymphocytes, natural killer cells, and macrophages were determined in the supraglottic, glottic, subglottic, and tracheal mucosa on days 2, 5, 7, and 14 after virus application. Furthermore, the nucleoprotein of the virus and major histocompatibility complex (MHC) Class II expression were detected immunohistologically on the laryngeal epithelium. RESULTS: All cell subsets entered the laryngeal mucosa during inflammation. The highest influx was detected among dendritic cells subglottically. This was accompanied by a strong virus adhesion and MHC Class II expression on the subglottic epithelium. In contrast, only a few immunocompetent cells entered the adjacent glottic mucosa, and on the glottic epithelium staining for virus nucleoprotein and MHC Class II expression was weak. CONCLUSIONS: The inflammatory response of the laryngeal mucosa shows great regional differences in this animal model during experimental viral infection. The response was characterized by a strong subglottic and a weak glottic reaction. A possible reason for this difference might be region-specific viral adhesion on the epithelium of the laryngeal areas, as well as differences in MHC Class II expression. Thus, these data agree with the clinical observation during acute laryngotracheitis and may explain why the subglottic part of the larynx is affected preferentially during pseudocroup. The molecular mechanisms mediating the different reactions await clarification.