Effects of 5 -fluorouracil on mitogen -induced costimulatory capacity of accessory cells from rat oral mucosa and dental pulp

The present study was designed to investigate the effect of the antimetabolite 5-fluorouracil (5-FU) on the capacity of the oral epithelium and the dental pulp to induce a mitogen-driven T-cell proliferation. Inbred Lewis rats were given 6 i.v. injections of 5-FU (30 mg/kg or 50 mg/kg) over a period of 8 days. Suspensions of oral epithelial and dental pulpal cells were prepared. The costimulatory capacity of the accessory cells from treated animals was monitored by their ability to induce a mitogen (ConA)-mediated proliferation of T cells isolated from regional lymph nodes of untreated animals. Accessory epithelial cells from rats treated with the high dose of 5-FU, but not the low dose, induced a decreased T-cell proliferation compared to controls. Accessory pulpal cells from rats, treated with 30 mg/kg or 50 mg/kg of 5-FU, induced a lower T-cell proliferation. When MHC class II molecule depleted T-cell suspensions from lymph nodes of 5-FU-injected animals were incubated with ConA, a significant proliferative response was observed. This finding correlated with an increase of MHC class II molecule expressing cells detected after incubation, although no such cells were observed immediately following the initial purification step of T cells. This finding demonstrates that the accessory cells could partly restore their expression of MHC class II molecules during incubation. The results of the study suggest that the function of immunocompetent cells of the oral mucosa and dental pulp is influenced by treatment with 5-FU and that the function of accessory cells of the pulp is affected more than the function of accessory cells derived from the oral mucosa.     

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.     

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.     

Ontogeny of class II antigen expressing cells in rat incisor pulp.

Class II antigen expressing cells are generally associated with the early phase of the immune response. Dendritic cells and macrophages expressing these cell surface antigens have recently been demonstrated and characterized in the dental pulp. The present study was undertaken to determine when the pulp receives its immunologic defense potential by examining the temporal appearance of class II antigen expressing cells in the rat incisor pulp. Pulp tissue specimens obtained at various time periods from a gestational age of 16 days to 14 wk after birth were examined by immunohistochemistry using O x 6 as a primary antibody and the ABC-technique. Comparisons were made with tissue samples from the spleen, intestine, skin and oral mucosa. At birth, all tissues, except for the pulp, presented cells expressing class II antigen with a dendritic appearance in a number and orientation resembling the mature tissue. A complete distribution of these cells was not seen in the dental pulp until 7 weeks following birth. Data show that the dental pulp acquires its ultimate structural arrangement of immune cells later than other tissues.     

Ontogeny of class II antigen expressing cells in rat incisor pulp

Abstract— Class II antigen expressing cells are generally associated with the early phase of the immune response. Dendritic cells and macrophages expressing these cell surface antigens have recently been demonstrated and characterized in the dental pulp. The present study was undertaken to determine when the pulp receives its immunologic defense potential by examining the temporal appearance of class II antigen expressing cells in the rat incisor pulp. Pulp tissue specimens obtained at various time periods from a gestational age of 16 days to 14 wk after birth were examined by immunohistochemistry using O x 6 as a primary antibody and the ABC-technique. Comparisons were made with tissue samples from the spleen, intestine, skin and oral mucosa. At birth, all tissues, except for the pulp, presented cells expressing class II antigen with a dendritic appearance in a number and orientation resembling the mature tissue. A complete distribution of these cells was not seen in the dental pulp until 7 weeks following birth. Data show that the dental pulp acquires its ultimate structural arrangement of immune cells later than other tissues.     

Expression of adhesion and activation molecules in human buccal epithelial cell lines and normal human buccal epithelium in situ

An in vitro culture system may be used to analyse interactions between T cells and epithelium. We have analysed two human buccal squamous epithelial cell lines: SqCC/Y1, derived from a buccal carcinoma, and SVpgC2a, an SV-transformed healthy buccal squamous epithelium. Under serum-free culture conditions, the cell lines resemble normal buccal squamous epithelium in situ in their expression of MHC class I, CD29 (beta1 integrin), CD40, CD44, CD54 (ICAM-1), CD58 (LFA-3), CD95 (Fas), and E-cadherin. Furthermore, when the SVpgC2a cell line was treated with T-cell derived cytokines, upregulation of CD40 expression was observed when the cells were treated with IL-4, whereas IFN-gamma caused upregulation in expression of CD40, CD54 and MHC class II. These buccal squamous epithelial cell lines, therefore, provide a good tool for analysing interactions between epithelium and T cells in vitro.     

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.     

Immunohistologic analysis of epithelial cell populations in oral lichen planus

Previous studies have demonstrated heterogeneity within lesional lymphocytes in drug-related oral lichen planus (D-LP) and idiopathic lichen planus (I-LP). This study examined the phenotype of Langerhans cells (LC) and keratinocytes in non-erosive D-LP and I-LP. In I-LP, keratinocytes expressed HLA-DR antigens whilst LC co-expressed CDIa, MHC Class II and CD4 antigens. The high levels of expression of MHC Class II antigens by LC were maintained during short term organ culture. In I-LP, the epithelium contained occasional CD25+ dendritic cells (putative activated LC). These cell phenotypes are suggestive of cell activation and likely result from local production of gamma interferon. In D-LP, expression of MHC Class II antigens on LC was reduced and no CD25+ cells were detected. The epithelium contained an increased number of CD45RA+ dendritic cells. While no differences between the production of interleukin-1 and interleukin-1 inhibitors by tissue samples could be detected, it is likely that the variations in epithelial cell phenotypes in I-LP and D-LP reflect altered cytokine production.     

Oral mucosal Langerhans cells express DR and DQ antigens

The expression of the Class II antigens HLA-DR and HLA-DQ in human oral mucosa was examined in health and in the presence of inflammation. DQ antigens were detected on dendritic intra-epithelial cells which expressed the Langerhans cells (LC) phenotype T6+, DR+. In healthy gingiva, DR and DQ were co-expressed on Langerhans cells, whereas in an experimental gingivitis (day 8), more LC expressed DR than DQ. Absolute LC numbers were increased in inflammation. Traumatic ulceration of the buccal mucosa resulted in a decrease in the density of T6-, DR-, and DQ-positive cells. Repopulation of migrating and regenerated epithelium was complete 10 days after ulcer induction. Disparity between DR and DQ expression was seen in both normal buccal mucosa and throughout the ulcer healing period. These results are in agreement with the reported sequence of Class II antigen expression on lymphoid 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)     

A comparative light-microscopic, electron-microscopic and chemical study of human vaginal and buccal epithelium

The scarcity of sizeable specimens of normal oral mucosa for experimental purposes has hampered research on oral epithelium. Because large specimens of viable human vaginal mucosa are readily available and because vaginal and buccal epithelia are microscopically similar, vaginal mucosa has been used successfully to establish a human cyst model in experimental animals. The ultrastructure and distribution of keratin filaments in these epithelia are also similar, as is their permeability to water and a number of chemical substances. Therefore, if vaginal mucosa could be substituted for buccal mucosa it would expedite research on the epithelium of buccal mucosa. To strengthen further the concept that vaginal epithelium could replace buccal epithelium in certain experimental studies, the thickness of these epithelia, their patterns of surface keratinization, the presence or absence of intercellular lipid lamellae and their lipid contents were now compared. Thirty-three specimens of vaginal mucosa from postmenopausal women and 36 of buccal mucosa were investigated. To compare the thickness of the epithelial layers the number of cell layers in sections of 20 vaginal and 20 buccal mucosal specimens were counted in the three thickest and three thinnest regions of each specimen. Surface keratinization was evaluated on sections stained with the Picro-Mallory method. To demonstrate lipid lamellae two vaginal and two buccal mucosa specimens were examined electron microscopically after normal fixation and postfixation in ruthenium tetroxide. Following solvent extraction of 11 vaginal and 14 buccal epithelia, quantitative lipid analyses were performed using thin-layer chromatography. No statistically significant differences were found between the maximum and minimum number of epithelial cell layers. The patterns of surface keratinization and the distribution and appearance of the lipid lamellae in the intercellular spaces were similar. The lipid composition of the two epithelia corresponded, except for the cholesterol esters and glycosylceramides, which were higher in buccal epithelium. Ceramides for vaginal epithelium and triglycerides for buccal epithelium were not determined. Based on structural similarities, a similar lipid composition and earlier findings, it is concluded that vaginal epithelium can be used as a substitute for buccal epithelium in certain in vitro, and possibly for in vivo, studies.     

An immunohistochemical study of the distribution of immunocompetent cells, especially macrophages and Ia antigen-expressing cells of heterogeneous populations, in normal rat molar pulp.

The precise distribution of various immunocompetent cells in rat molar pulp was immunohistochemically examined by use of seven anti-rat monoclonal antibodies. It was demonstrated that rat molar pulp contained many OX6 (anti-Ia antigen)-positive cells and a large number of ED1 (anti-monocytes, macrophages, and dendritic cells)-positive, ED2 (anti-tissue macrophages)-positive, and/or OX35 (anti-macrophages and CD4+ lymphocytes)-positive cells. Macrophage-like cells predominated in the central portion of the pulp, while cells of dendritic appearance usually existed in the periphery of the pulp. Double-immunoperoxidase staining revealed that these cells showed some heterogeneity, but the majority could be classified as ED1+/OX6-/ED2+ cells, which may be Ia-histiocytes. Findings also suggested that true dendritic cells may be included in the ED1+/OX6+/ED2- category of cells. A small number of T lymphocytes and plasma cells were also detected. These results suggest that the normal dental pulp contains a variety of immunocompetent cells, with macrophages as the most dominating. Following the exogenous invasion of pathogenic stimuli in the pulp, these cells may participate in the defense reaction by acting as phagocytes or antigen-presenting cells, which are essential for the initiation of immune responses.     

Expression of Class II transplantation antigens by epithelial cells in oral candidosis, oral lichen planus and gingivitis

Biopsies from normal oral mucosa and oral mucosa affected by candidosis, lichen planus or gingivitis were compared with respect to the expression of two Class II transplantation antigens, HLA-DR and HLA-DQ, by epithelial cells and the relationship of these antigens to the distribution and frequency of T-lymphocytes. Indirect immunohistochemistry with different mouse monoclonal antibodies was used on frozen and acetone-fixed sections. To evaluate the results, a score system based upon the expression of the Class II transplantation antigens by epithelial cells and the frequency of T-lymphocytes was used. In oral candidosis there was a marked expression of HLA-DR antigens throughout the epithelium. In addition, this type of epithelium was the only one that expressed HLA-DQ antigens. An intense intraepithelial infiltration of T-lymphocytes was observed. Oral lichen planus and gingivitis did, to a much lesser extent, cause the expression HLA-DR antigens by the epithelial cells. In both lesions, the number of T-lymphocytes within the epithelium did not exceed the number found in epithelium of normal mucosa. In these types of lesions, the subepithelial infiltrate varied in intensity but was mainly composed of T-lymphocytes reactive with anti-Leu 3a antibodies. The results of the present study imply that epithelial expression of the two different Class II antigens are related to the frequency of the T-lymphocytes and to the proximity of these cells to the epithelial cells.     

Expression of Class II transplantation antigens by epithelial cells in oral candidosis, oral lichen planus and gingivitis

Biopsies from normal oral mucosa and oral mucosa affected by candidosis, lichen planus or gingivitis were compared with respect to the expression of two Class II transplantation antigens, HLA-DR and HLA-DQ, by epithelial cells and the relationship of these antigens to the distribution and frequency of T-lymphocytes. Indirect immunohistochemistry with different mouse monoclonal antibodies was used on frozen and acetone-fixed sections. To evaluate the results, a score system based upon the expression of the Class II transplantation antigens by epithelial cells and the frequency of T-lymphocytes was used. In oral candidosis there was a marked expression of HLA-DR antigens throughout the epithelium. In addition, this type of epithelium was the only one that expressed HLA-DQ antigens. An intense intraepithelial infiltration of T-lymphocytes was observed. Oral lichen planus and gingivitis did, to a much lesser extent, cause the expression HLA-DR antigens by the epithelial cells. In both lesions, the number of T-lymphocytes within the epithelium did not exceed the number found in epithelium of normal mucosa. In these types of lesions, the subepithelial infiltrate varied in intensity but was mainly composed of T-lymphocytes reactive with anti-Leu 3a antibodies. The results of the present study imply that epithelial expression of the two different Class II antigens arc related to the frequency of the T-lymphocytes and to the proximity of these cells to the epithelial cells.     

Cultured pulp fibroblasts: are they suitable for in vitro cytotoxicity testing?

The use of cell cultures to test the biocompatibility of dental materials is gaining in importance. Any cytotoxic effects that restorative materials may have will be on the dental pulp and for that reason cultured pulp cells should be the model of choice for biocompatibility testing. The aim of this investigation was to study the growth and morphologic characteristics and toxic response of human pulp lines and to compare these parameters to those of human buccal mucosa fibroblasts. Twenty-one specimens of pulp tissue and six from buccal mucosa were cultured using standard techniques. Six pulp cell lines were cultured successfully as were all six from the buccal mucosa specimens. From these specimens, 12 growth curves were computed. To study the morphology of the cultured cells, they were observed microscopically and classified into three morphological types: slender elongated cells (type I), epithelioid shaped cells (type II) and large stellate cells (type III). Their numbers and proportions were determined for each cell line and compared statistically. To gauge sensitivity to toxic materials, cells were exposed to concentrations of arecoline. An analysis of the growth curves showed no statistical difference between pulp cells and buccal mucosa cells; the slopes of the curves, however, differed significantly between individual cell lines, and these individual differences were greater among pulp cell lines. The morphology of the pulp and mucosa fibroblasts was similar microscopically. There was no significant difference between the number and proportion of the cell types in the two groups, but there were significant differences between the individual cell lines. Pulp cells showed a greater inhibition of growth when exposed to arecoline. Because pulp fibroblasts are difficult to culture, their reported survival rate is poor; due to the differences that exist between individual cell lines, we conclude that pulp cells when used as single cell lines or even pooled may not be ideal for testing biocompatibility, especially if reproducibility is a prerequisite. Any evaluation will require tests on not one, but several cell lines in order to minimize the effect of inter-cell-line differences. Their greater sensitivity to toxic substances, on the other hand, may show that pulp cells could be more sensitive indicators of cytotoxicity.     

Quantification and distribution of lymphocyte subsets and Langerhans cells in normal human oral mucosa and skin

Normal human oral (check) mucosa was studied to discover whether the oral cavity resembles the Mucosal Immune System (MIS) or the Skin Immune System (SIS). Immunophenotypes of lymphocyte subsets and Langerhans cells (LC) with their exact locations in the epithelium and papillary layer of the normal buccal mucosa were determined and compared with data of normal human skin. In a double staining procedure, the distribution of T-lymphocytes in relation to blood and lymph vessels was determined. Immunophenotyping of LC was done with a CD1a monoclonal antibody. In contrast to the skin, T-lymphocytes in buccal mucosa are not primarily perivascular in location. They are more or less randomly distributed on both sides of the basement membrane. The epithelium of the buccal mucosa contains about 37 times as many T-lymphocytes as the epidermis of normal skin. T-cell numbers in the papillary layer are more or less comparable. The CD4/CD8 ratios of about 1/2 in the epithelium of buccal mucosa and 1/4 in the skin indicates preferential presence of the CD8 subset in both sites, but the helper/inducer T-lymphocytes play a much greater role in the epithelium of the buccal mucosa when compared with skin. B-lymphocytes were not found in the epithelium and papillary layer of the buccal mucosa. Thus, immune response associated cells in buccal mucosa do not show the MIS pattern since B cells are absent. It has more in common with SIS but differences are also apparent. In the epithelium of the buccal mucosa the density of LC does not differ significantly from that of the skin, but the papillary layer of the buccal mucosa contains significantly fewer LC than the skin. As in the skin most of the LC of the buccal mucosa are found in the epithelium.     

Scanning electron microscopic study of surface of human oral mucosa

The surface ultrastructure of the healthy oral mucosa of humans was studied using SEM as follows: dorsum of the tongue (10 specimens), buccal mucosa (5), floor of the mouth (3), hard palate (5), and gingiva (10). One part of each formalin-fixed sample was processed routinely using the system of critical point drying for scanning electron microscopy. The other part of the specimen was embedded in paraffin blocks and stained with hematoxylin-eosin for light microscopy. With SEM at low magnification, the surface structure of the oral mucosa at different areas of the oral cavity was smooth with some desquamating cells. Only the tongue mucosa with its papillae formed a specialized mucosa. The hairs of the filiform papillae were covered by microorganisms, whereas on the oral mucosa there usually was little or no colonization by microorganisms. At high magnification, the surface structure of the superficial epithelial cells was pitted or microplicated. On keratinized epithelium the surface structure was pitted, whereas on non-keratinized epithelium it was microplicated. On cell boundaries some variation could also be seen; in keratinized epithelium these boundaries were overlapping and in non-keratinized epithelium they were tight.     

Scanning electron microscopic study of surface of human oral mucosa

The surface ultrastructure of the healthy oral mucosa of humans was studied using SEM as follows: dorsum of the tongue (10 specimens), buccal mucosa (5), floor of the mouth (3), hard palate (5), and gingiva (10). One part of each formalin-fixed sample was processed routinely using the system of critical point drying for scanning electron microscopy. The other part of the specimen was embedded in paraffin blocks and stained with hematoxylin-eosin for light microscopy. With SEM at low magnification, the surface structure of the oral mucosa at different areas of the oral cavity was smooth with some desquamating cells. Only the tongue mucosa with its papillae formed a specialized mucosa. The hairs of the filiform papillae were covered by microorganisms, whereas on the oral mucosa there usually was little or no colonization by microorganisms. At high magnification, the surface structure of the superficial epithelial cells was pitted or microplicated. On keratinized epithelium the surface structure was pitted, whereas on non-keratinized epithelium it was microplicated. On cell boundaries some variation could also be seen; in keratinized epithelium these boundaries were overlapping and in non-keratinized epithelium they were tight.     

槟榔提取物抑制人类口腔粘膜角朊细胞生长的实验研究

目的：建立人类口腔粘膜上皮角朊细胞体外原代培养方法：探讨口腔粘膜下纤维性变患者上皮厚度改变的机理;方法;先对人类口腔粘膜上皮角朊细胞进行分离,培养和鉴定,然后用四唑盐比色试验检测ＯＳＦ患者和正常人口腔粘膜上皮ＫＣ增殖状况,并且观察槟榔提取物对ＫＣ增殖的影响。