Pattern of distribution of T lymphocytes, Langerhans cells and HLA-DR bearing cells in normal human oral mucosa

The tissue distribution of helper/inducer and suppressor/cytotoxic T cells, Langerhans cells (LC) and HLA-DR bearing cells was determined in normal oral mucosa by use of monoclonal antibodies OKT4, OKT8, OK.T6 and OKIal, respectively. OKT4+ and OKT8+ cells were invariably present in normal oral epithelium and in the lamina propria. OKT8+ cells were consistently seen inside the basal cell layer of the epithelium. The distribution of LC in oral epithelium showed regional variation. In palatal epithelium LC were evenly distributed in the basal half of the epithelium, whereas in buccal mucosa the highest concentration of LC was seen in the epithelium overlying the tips of connective tissue papillae. OKIal stained dendritic cells in the epithelium and plump cells with small dendritic processes in the connective tissue. Some of the latter were located close to the basal cells of the epithelium. The consistent relationship between immunocompetent cells and the epithelium of the oral mucosa suggests the presence of a local immunologic defence barrier in the oral mucosa.     

Pattern of distribution of T lymphocytes, Langerhans cells and HLA-DR bearing cells in normal human oral mucosa

The tissue distribution of helper/inducer and suppressor/cytotoxic T cells, Langerhans cells (LC) and HLA-DR bearing cells was determined in normal oral mucosa by use of monoclonal antibodies OKT4, OKT8, OKT6 and OKIa1, respectively. OKT4+ and OKT8+ cells were invariably present in normal oral epithelium and in the lamina propria. OKT8+ cells were consistently seen inside the basal cell layer of the epithelium. The distribution of LC in oral epithelium showed regional variation. In palatal epithelium LC were evenly distributed in the basal half of the epithelium, whereas in buccal mucosa the highest concentration of LC was seen in the epithelium overlying the tips of connective tissue papillae. OKIa1 stained dendritic cells in the epithelium and plump cells with small dendritic processes in the connective tissue. Some of the latter were located close to the basal cells of the epithelium. The consistent relationship between immunocompetent cells and the epithelium of the oral mucosa suggests the presence of a local immunologic defence barrier in the oral mucosa.     

Ultrastructural examination of experimentally induced premalignant lesions

Lesions induced in hamster cheek pouch using dimethylbenz(alpha)anthracene were studied by transmission electron microscopy and compared with normal tissue. Features regarded as suggestive of progression towards malignancy were: increased numbers of membrane-bound vesicles in basal cells, gaps in the lamina densa associated with widening of the lamina lucida and an irregular epithelial-connective tissue junction, the presence of cerebriform cells and frequent close association of 2 or more intra-epithelial cells (lymphocytes, cerebriform cells, Langerhans cells).     

Regional variation in Langerhans cell distribution and density in normal human oral mucosa determined using monoclonal antibodies against CD1, HLADR, HLADQ and HLADP

The distribution, density and activation of Langerhans cells (LC) has been established in biopsies of normal human buccal mucosa, hard palate, lateral border and dorsum of tongue, floor of mouth and lip taken from sudden death post mortems. LC were identified in cryostat sections with monoclonal antibodies against CD1, HLADR, HLADQ and HLADP using an immunoalkaline phosphatase technique. The use of post mortem material was validated by comparison with biopsies taken from volunteers. LC were predominantly situated in the basal and immediately suprabasal layers of the epithelium. In floor of mouth, lip, lateral border and dorsum of tongue the cells were found along the length of the epithelium. In buccal mucosa, although LC showed fundamentally a similar distribution, a tendency to cluster around the connective tissue papillae was also noted. In hard palate LC were found parallel to the surface in the mid zone of the epithelium. No evidence of LC free areas was found. Dorsum of tongue had the highest density of LC per mm epithelial surface length (28.3 cells per mm) which was significantly greater (P less than 0.05) than buccal mucosa (25.2) which in turn had significantly more cells (P less than 0.05) than lip (22.4). The lowest density of LC was found in lateral border of tongue (17.6), hard palate (17.6) and floor of mouth (16.7). These sites had significantly fewer cells than lip, buccal mucosa and dorsum of tongue (P less than 0.05). Class II MHC molecules are necessary for antigen presentation and in all sites except buccal mucosa there was no significant difference between the number of cells expressing CD1, HLADR, HLADQ and HLADP.(ABSTRACT TRUNCATED AT 250 WORDS)     

Intraepithelial lymphocytes and Langerhans cells in the oral mucosa--dynamic aspects.

In the oral epithelium a constant population of non-epithelial immunocompetent cells--lymphocytes and Langerhans cells exists. 3025 such cells of the murine and 542 of the normal human oral mucosa were documented photographically at the ultrastructural level. Location within the epithelium, morphological signs of locomotion, membrane configuration and labeling of the intercellular substance by ruthenium red were registered. About 40 per cent of the lymphocytes and 70 per cent of the Langerhans cells show signs of locomotion, predominantly in a lateral and basal direction. Occasional cells crossing the basal lamina are seen. Close apposition, or rarely a denticulated surface and lysis of desmosomes were present, both probably indications of cellular interactions. Ruthenium red labeling of the glycocalix is continuous forming a homogeneous layer between epithelial and non-epithelial cells and between intraepithelial cells, suggesting an even distribution of antigens and the lack of preformed spaces for intraepithelial cells. Murine and human intraepithelial cells exhibited no major differences.     

Integrin expression in oral hairy leukoplakia and normal tongue epithelium

OBJECTIVE: To describe the expression of integrins in the epithelium of oral hairy leukoplakia (HL) and compare to that of normal lateral tongue epithelium. MATERIALS AND METHODS: Immunohistochemistry to identify integrins (alpha 2, alpha 3, alpha 5, alpha 6, alpha v, beta 1) was performed, using a standard biotin-streptavidin-peroxidase technique on five clinically and histologically confirmed frozen biopsy specimens of HL and five normal lateral tongue control tissues. RESULTS: Expression of integrins alpha 2, alpha 3, alpha 6, alpha v, beta 1 was seen both in HL epithelium and in normal control tissue. alpha 5 expression was not seen in HL or in control tissue epithelium. alpha 2 and alpha 3 were expressed mainly in the basal and suprabasal layers; alpha 6 expression was most intense on the basal surface of the basal cells, alpha v was expressed in the basal and suprabasal layers with more expression seen in the higher differentiated cell layers than the other integrins. beta 1 expression was seen in the basal and suprabasal layers only. No apparent difference between HL and normal oral mucosa was noted in the staining pattern of the various integrins. CONCLUSION: Integrins alpha 2, alpha 3, alpha 6, alpha v, beta 1 are expressed in HL and the expression pattern is not different from that of normal oral mucosa. alpha 5 is not expressed in HL or in normal oral epithelium.     

Human gingival Langerhans cells in health and disease

Epithelial Langerhans cells in samples of healthy and diseased gingival tissue were studied using ATPase histochemistry and the monoclonal antibodies OKT6 and anti HLA-DR. In healthy gingiva Langerhans cells were seen in both oral and sulcular epithelium; they were generally positioned in the basal layers. No Langerhans cells were seen in junctional epithelium. In diseased tissue there was a large increase in the number of Langerhans cells in both oral and sulcular epithelium with many more being situated in the stratum spinosum. There was an increase in the expression of the Class 2 antigen, HLA-DR, and morphological polarization occurred with dendrites preferentially orientated towards the surface. No Langerhans cells were seen in the pocket lining epithelium of periodontally diseased gingiva.     

Effects of a 10% carbamide peroxide bleaching agent on rat oral epithelium proliferation

The purpose of the present study was to evaluate the influence of short course topical application of carbamide peroxide on proliferating cell nuclear antigen (PCNA) immunohistochemical expression in the oral tongue mucosa of rats. Twelve male Wistar rats were submitted to topical application of 10% carbamide peroxide on one side of the dorsal tongue once a week for three consecutive weeks. Only distilled water was applied on the control side. The animals were killed on days 0, 10, and 20 after the last application. The tongue was fixed in buffered formalin for 24 h and embedded in paraffin. Tissue blocks (3 microns) were subjected to the biotin-streptavidin amplified system for identification of PCNA. The percentage of epithelial-positive basal cells in each side of the tongue mucosa was calculated. The results demonstrated that topical application of 10% carbamide peroxide increases PCNA immunohistochemical expression on the basal layer of the oral mucosa epithelium of rats on day 0 after treatment. In conclusion, short-course use of carbamide peroxide induces transient epithelial cell proliferation of the oral mucosa of rats.     

Quantitative analysis of immunocompetent cells in human normal oral and uterine cervical mucosa, oral papillomas and leukoplakias

Using monoclonal antibodies reacting with T-cell subpopulations, Langerhans cells and macrophages, the number and distribution of cells of the immune system in normal oral and cervical mucosa was determined and statistically compared with that in oral papillomas and oral leukoplakias. Increased numbers of labelled cells were found in oral leukoplakias and particularly in oral papillomas. In the epithelium of all specimens, Langerhans cells and T-lymphocytes of the suppressor/cytotoxic phenotype as well as of the helper phenotype were seen. Suppressor/cytotoxic and helper T-lymphocytes were in equal numbers in the epithelium of oral papillomas, but were about 2:1 in all other lesions. In normal oral epithelium, macrophages were rare but were in greater numbers in leukoplakias and papillomas. In the connective tissue of all lesions, more labelled cells were present than in epithelium with T-lymphocytes predominant. Although Langerhans cells were rare in connective tissue, many were seen in oral papillomas.     

Langerhans cell surface densities in rat oral mucosa and human buccal mucosa

We determined surface densities of Langerhans cells (LCs) in rat oral mucosa and human buccal mucosa by enumerating ATPase-positive dendritic cells in epithelial whole mounts. For the rat, mean surface densities per mm² were 160 in anterior buccal mucosa, 640 in posterior buccal mucosa, 430 in the palate and 340 in the tongue. Human buccal mucosa showed a density of 890 cells per mm². We conclude that LC densities in oral mucosa approximate those of external body sites, making them available in numbers sufficient to accomplish their postulated antigen-presenting functions.     

Extent and diversity of inflammatory cell infiltrates in squamous cell carcinomas and basal cell epitheliomas of the head and neck

Using monoclonal antibodies reactive with Langerhans' cells (LCs), macrophages, and T cell subpopulations, the density and proportions of cells of the immune system of the normal oral mucosa were determined immunohistochemically, and compared with findings in oral squamous cell carcinomas (SCC) and basal cell epitheliomas (BCE). In normal oral epithelia, the dominant cell type was the LC, positive for CD 1, and expressing HLA-DR antigens (DR+). Many intraepithelial cells were lymphocytes of the suppressor/cytotoxic phenotype (CD 8+), which was also the most prominent cell type in the normal mucosal stroma. Significant differences were observed for the content of CD 8-, OKM 1-, and CD 4-positive cells in the epithelium of normal oral mucosa, SCC, and BCE, and for the amount of CD 1-positive Langerhans cells in the connective tissue of the different groups of tissues. When CD 4/CD 8 ratios were calculated, differences between SCC and BCE became most evident. A CD 4/CD 8 ratio greater 0.5 was seen to be characteristic for BCE. Thus, in contrast to the striking preponderance of suppressor/cytotoxic lymphocytes (CD 8+) in SCC, BCE showed typically almost balanced numbers of suppressor/cytotoxic (CD 8+) and helper/inducer (CD 4+) lymphocytes. This finding further underlines the biological differences recognized between these most common neoplasias of the head and neck.     

Interepithelial lymphocytes in experimental gingivitis in young and elderly individuals

Experimental gingivitis was induced and monitored in young and elderly subjects. After 9 days of oral hygiene abstention, a gingival biopsy was obtained from the labial surface of the lower right cuspid. The distribution of lymphocytes within the crevicular epithelium was assessed, and the ultrastructural morphology of the lymphocytes was studied. From day 0 to day 9 the increase in gingival inflammation (Gingival Index) was greater in elderly than in young subjects. The number of lymphocytes per 100 epithelial cells was greater within the crevicular epithelium of elderly subjects. However, this difference was not statistically significant. The ratio of lymphocytes was significantly higher in the coronal than in the apical portion of the epithelium in both age groups. On an ultrastructural level two types of interepithelial lymphocytes, inactive and intermediate, were recognized. The intermediate cell types displayed morphological characteristics typical of cells responding to antigen stimulation.     

Identity of TUNEL-positive cells in the oral buccal epithelium of normal mucosa and lichen lesions

BACKGROUND: In situ detection of DNA fragmentation by TdT (terminal deoxynucleotidyl transferase)-mediated dUTP nick end labeling (TUNEL) is a widely used technique to identify apoptotic cells in the terminal phases of cell death. Several studies have shown that there are statistically increased numbers of TUNEL(+) cells within the epithelium of oral lichen (OL). It was suggested that this indicates an increased rate of apoptosis among basal and suprabasal keratinocytes in OL epithelium. The aim of this study was to identify the TUNEL(+) cells in the epithelium of erythematous (ERY) OL and normal oral mucosa (NOM). METHODS: Sections of biopsies from NOM and ERY OL were processed for TUNEL combined with immunostaining for pan-cytokeratin or for cell markers specifically expressed by different leukocytes. RESULTS: In NOM, TUNEL(+) keratinocytes were almost exclusively seen in the outermost epithelial layers. This labeling was absent in ERY OL. In the basal and lower spinous layers, more TUNEL(+) cell nuclei were seen in ERY OL as compared with NOM, in accordance with previous studies. The present observations show, however, that only very few of these cells were keratinocytes, but rather were CD4(+) lymphocytes and CD68(+) macrophages. There was no difference between the numbers of TUNEL(+) keratinocytes in basal and lower spinous layers in ERY OL and NOM epithelium. No intraepithelial CD8(+) lymphocytes, Langerhans cells, or mast cells were found to be TUNEL(+). CONCLUSION: The findings indicate that the pathologic changes in ERY OL epithelium cannot be explained by increased prevalence of terminal keratinocyte cell death identified by TUNEL.     

Hairy leukoplakia: Epstein-Barr virus receptors on oral keratinocyte plasma membranes

Hairy leukoplakia (HL) is an oral white lesion associated with, and probably caused by, the Epstein-Barr virus (EBV) among persons who are seropositive for infection with human immunodeficiency virus. A unique feature of HL is its localization to the lateral portion of the tongue. To determine site differences for EBV receptors according to epithelial phenotype, these receptors were mapped in oral mucosa with the use of monoclonal antibodies HB5 and B2(specific for the Complement Fraction 3d/EBV receptor on B lymphocytes). Immunoperoxidase and immunofluorescence techniques were employed with the use of both cytologic suspensions and frozen tissue sections of oral epithelium. Pericellular plasma membrane immunoreactants were localized to upper spinous layer cells of the parakeratin phenotype; basal and parabasilar layers as well as all strata of orthokeratinized epithelia were negative. Those cells harboring EBV DNA as detected by in situ hybridization corresponded to cells with C3d/EBV receptors.     

Langerhans cells: structure, function and role in oral pathological conditions

Langerhans cells (LCs) are dendritic bone marrow derived cells situated suprabasally in most stratified squamous epithelia, such as the epidermis and the epithelium of oral mucosa, including the gingiva. Langerhans cells are thought to act as antigen-presenting cells (APC) during induction of immune responses. The exact role of Langerhans cells in the oral mucosa is not fully understood although several investigations suggest that these cells are involved in reactions to antigen challenge under both normal and pathological situations. In this paper the structure, phenotypic markers and derivation of Lungerhans cells are reviewed. In view of recent findings, the immunological characteristics and the implications of Langerhans cells in pathologic oral reactions are discussed.     

Macro- and microanatomy of the lateral border of the tongue with special reference to oral hairy leukoplakia

The macro- and microanatomy of the marginal tongue mucosa were studied. Tissues were harvested from 12 individuals at autopsy. Random sections from six different locations along the margin and serial sections representing three defined section planes to the margin of the tongue were evaluated. Vertical, parallel mucosal folds alternating with shallow grooves were a characteristic macroscopical findings on the lateral border of the tongue. The mucosa presented a non-keratinized epithelium with PAS-positive, lightly stained spinous cells and no or slight inflammatory reaction in the connective tissue. Changes in the epithelium mimicking hyperplasia, acanthosis, keratin projections, and focal parakeratosis could be produced by changing the direction of tissue sectioning. The macro- and microscopical parameters recorded in normal marginal tongue mucosa are among other included in criteria for diagnosing oral hairy leukoplakia. The results emphasize the importance of a thorough knowledge of the normal anatomy of a mucosal site to arrive at reliable diagnostic criteria.     

Ultrastructural findings in the oral mucosa of betel chewers

Eighteen biopsies of the oral mucosa of northern Thai hilltribe betel chewers were studied histologically and by transmission and scanning electron microscopy (TEM, SEM). Clinically, varying stages of epithelial atrophy and one case of submucous fibrosis were observed. Histologically, epithelial atrophy with marked reduction of the rete pegs, hyperortho- and/or parakeratosis, and subepithelial edema and inflammatory changes were the prominent findings. On the ultrastructural level, cytoplasmic projections of the basal cells into the subepithelial stroma were seen. The basal membrane frequently revealed gaps; the interepithelial space was widened and unusual microvilli were observed on cell surfaces (SEM). Intercellularly, cristalloid material of unknown origin was also seen. The subepithelial connective tissue was characterized by dense bundles of collagen fibres adjacent to which masses of amorphous material were located. While some of the ultrastructural findings in the epithelium of betel chewers are indicative of early dysplastic changes, the nature of the juxta-epithelial stromal alterations is still unknown. Current hypotheses regarding the etiology of oral submucous fibrosis are briefly discussed.     

Extent and diversity of inflammatory cell infiltrates in squamous cell carcinomas and basal cell epitheliomas of the head and neck

Using monoclonal antibodies reactive with Langerhans' cells (LCs), macrophages, and T cell subpopulations, the density and proportions of cells of the immune system of the normal oral mucosa were determined immunohistochemically, and compared with findings in oral squamous cell carcinomas (SCC) and basal cell epitheliomas (BCE). In normal oral epithelia, the dominant cell type was the LC, positive for CD 1, and expressing HLA-DR antigens (DR⁺). Many intraepithelial cells were lymphocytes of the suppressor/cytotoxic phenotype (CD 8⁺), which was also the most prominent cell type in the normal mueosal slroma. Significant differences were observed for the content of CD 8-, OKM 1-, and CD 4-positive cells in the epithelium of normal oral mucosa, SCC, and BCE, and for the amount of CD 1-positive Langerhans cells in the connective tissue of the different groups of tissues. When CD 4/CD 8 ratios were calculated, differences between SCC and BCE became most evident. A CD 4/CD 8 ratio greater 0.5 was seen to be characteristic for BCE. Thus, in contrast to the striking preponderance of suppressor/cytotoxic lymphocytes (CD 8⁺) in SCC, BCE showed typically almost balanced numbers of suppressor/cytotoxic (CD 8⁺) and helper/induced (CD 4⁺) lymphocytes. This finding further underlines the biological differences recognized between these most common neoplasias of the head and neck.     

Ultrastructural findings in the oral mucosa of betel chewers

Eighteen biopsies of the oral mucosa of northern Thai hilltribe betel chewers were studied histologically and by transmission and scanning electron microscopy (TEM, SEM). Clinically, varying stages of epithelial atrophy and one case of submucous fibrosis were observed. Histologically, epithelial atrophy with marked reduction of the rete pegs, hyperortho- and/or parakeratosis, and subepithelial edema and inflammatory changes were the prominent findings. On the ultrastructural level, cytoplasmic projections of the basal cells into the subepithelial stroma were seen. The basal membrane frequently revealed gaps; the interepithelial space was widened and unusual microvilli were observed on cell surfaces (SEM). Intercellularly, cristalloid material of unknown origin was also seen. The subepithelial connective tissue was characterized by dense bundles of collagen fibres adjacent to which masses of amorphous material were located. While some of the ultrastructural findings in the epithelium of betel chewers are indicative of early dysplastic changes, the nature of the juxta-epithelial stromal alterations is still unknown. Current hypotheses regarding the etiology of oral submucous fibrosis are briefly discussed.     

Mucosal changes at erupting molars in germfree rats

An accumulation of inflammatory cells in the tissue adjacent to the newly established dentogingival junction of erupting teeth has been reported by several investigators. In order to elucidate the role played by bacteria in the induction of this presumed defence reaction against irritants from the oral cavity, the first and second molars were studied in seven germfree rats aged 14 to 20 days and in age-matched control animals. After previous demineralisation and embedding in paraffin the molars on one side were serially sectioned sagittally.
Inspection of the molar areas revealed no differences in the stage of eruption between the two groups. Also, the general histologic picture was almost identical in the two groups. Before fusion of the dental and oral epithelia, only the ordinary cells of loose connective tissue were present. As soon as the molar appeared in the oral cavity, both germfree and control animals regularly showed some accumulation of inflammatory cells in the vicinity of the epithelial confluence.
The greatest number of cells as well as an absolute dominance of leucocytes was noted at the onset of the reaction. Later, lymphocytes became frequent, but then the total number of inflammatory cells, and the proportion of lymphocytes, was smaller in the germfree animals than in the controls. At this stage the numbers of inflammatory cells also varied considerably in different areas around each molar.