Distribution of basal lamina type IV collagen and laminin in normal rat tongue mucosa and experimental oral carcinoma: Ultrastructural immunolocalisation and immunogold quantitation

The relationship of basal lamina, a form of specialised extracellular matrix which separates epithelial cells and other cell types from adjacent stroma, to the behaviour of malignant neoplasms of epithelial origin is not well understood. However, it is widely acknowledged that the properties of local invasion and metastasis of carcinomas are linked to extracellular matrix (including basal lamina) changes. In the present study, the distribution of the major basal lamina components, type IV collagen and laminin, in normal rat tongue mucosa and experimentally induced oral carcinomas was investigated using post-embedding immunogold techniques and electron microscopy. The expression of these components was also quantitatively analysed using morphometry and immunocytochemistry. Results indicated that type IV collagen and laminin were confined to the lamina densa of normal oral epithelial basal lamina, and that both components were also detected in the lamina densa of basal lamina associated with carcinomas, and in the extracellular matrix of tumours. Furthermore, laminin was detected within stromal fibroblasts in normal tissues and experimental carcinomas. Quantitative analysis indicated that expression of laminin was significantly increased in carcinomas. In contrast, type IV collagen expression was significantly decreased. The quantitative changes observed in the two basal lamina constituents may be related to the process of tumour invasion, reflecting altered metabolic activities of tumour and stromal cells. These observations may be of use in understanding the architectural characteristics of oral mucosa basal lamina and in assessing the malignant potential of epithelial dysplasias or “premalignant” lesions.     

Reactivity of monoclonal antibodies 17.13 and 63.12 with 141 oral mucosal lesions

We studied the reactivity of monoclonal antibodies (MAbs) 17.13 and 63.12 with normal and diseased human oral mucosa by means of the immunoperoxidase technique. The specimens included: 22 normal oral tissues, 20 benign tumors, 17 lichen planus, 23 focal keratosis and epithelial hyperplasias, 18 proliferative verrucous leukoplakias, 20 dysplasias, and 21 squamous cell carcinomas. In most cases of normal mucosa and benign lesions, MAb 17.13 stained basal epithelial cells only, whereas MAb 63.12 stained all cell layers above the basal cells. In the premalignant and malignant lesions MAb 17.13 stained above the basal cells and MAb 63.12 either stained areas not stained by MAb 17.13 or the staining was absent. Based on the different staining patterns observed, there appears to be a potential value of these new reagents in diagnostic histopathology regarding specimens with equivocal cellular morphology.     

Expression of integrin α9 subunit and tenascin in oral leukoplakia, lichen planus, and squamous cell carcinoma

OBJECTIVES: Integrin alpha9 subunit is a member of beta1 integrin family and binds tenascin (TN). It is expressed by stratified squamous epithelium and may be associated with cell differentiation and growth. We studied if the expression of alpha9 integrin and TN is altered in leukoplakia, lichen planus, and squamous cell carcinoma (SCC).METHODS: Frozen sections of tissue samples obtained from normal human keratinized (16 subjects) and non-keratinized (three subjects) oral mucosa, oral leukopakias with dysplasia (19 subjects), reticular type lichen planus (nine subjects), or oral mucosal SCC (23 subjects) were stained immunohistochemically with antibodies against alpha9 integrin and TN.RESULTS: In contrast to its most prominent localization at the cell membranes of the basal epithelial cells in the normal mucosa, alpha9 integrin was localized in a more diffuse pattern with focal loss of expression at the epithelial cell membranes in leukoplakic dysplasia, lichen planus, and SCC. In some areas of SCC, alpha9 integrin localized throughout all cell layers of the tumor epithelium. In most areas, alpha9 integrin colocalized with TN but in heavily inflamed areas there was focal loss of TN and alpha9 integrin at the basement membrane zone. CONCLUSIONS: The findings show that alpha9 integrin expression is altered in leukoplakic dysplasia, lichen planus, and SCC.     

In situ DNA hybridization analysis of oral papillomas, leukoplakias, and carcinomas for human papillomavirus.

Twenty-one papillomas, 23 ordinary benign keratoses, 13 smokeless tobacco keratoses, 10 verrucous hyperplasias, 10 verrucous carcinomas, 17 squamous cell carcinomas, 3 epithelial dysplasias, and 6 lichen planus lesions were evaluated for human papillomavirus (HPV) types 6/11, 16/18, and 31/33/35, with biotinylated double-stranded DNA probes by in situ hybridization. Sixty-two percent (13/21) of oral squamous papillomas were positive for HPV DNA. HPV DNA types 6 and 11 demonstrated the strongest reactivity. Of the 13 cases, 10 also showed some reactivity with HPV-16/18 and -31/33/35. None of the cases of keratoses, epithelial dysplasia, squamous cell carcinoma, verrucous hyperplasia, verrucous carcinoma, or lichen planus were positive for HPV DNA. This study confirms the consistent and frequent finding of HPV DNA in oral squamous cell papillomas and the inconsistency of being able to identify HPV DNA in keratotic, premalignant, or cancerous lesions of the oral mucous membranes.     

幽门螺杆菌感染与口腔扁平苔藓和鳞状细胞癌相关性的初步探讨

目的研究正常口腔黏膜、口腔扁平苔藓（OLP）和口腔鳞状细胞癌（OSCC）中幽门螺杆菌（Hmylori）感染情况及其与p53蛋白和p16蛋白表达的关系．探讨H．pylori感染与OLP和OSCC的相关性。方法OLP标本23例、OSCC标本50例和正常口腔黏膜标本10例,采用免疫组化sP法检测其H．pylori的感染、p53蛋白、p16蛋白的表达．应用SPSS13．0软件包对结果进行卡方分析检验．单因素方差分析和线性回归检验分析。结果正常口腔黏膜组织组、0LP组及0SCC组中H．pylori阳性率分别为0、47．8％和48．0％,正常口腔黏膜组与OLP组和OSCC组间差异有统计学意义（X2=7．17．P=0.013;X^2=8．00,P=0．004）;OLP标本中H．pylor／阳性组和阴性组p53蛋白表达率分别为45．5％和16．7％,p16蛋白表达率为分别36．4％和66．7％,组间差异有统计学意义（X^2=3．86,P=0．043：X^2=3．89,P=0．048）;OSCC标本中H.pylori阳性组和阴性组p53蛋白表达率分别为91．7％和84．6％,p16蛋白表达率为分别20．8％和30．8％,组间差异无统计学意义（X^2=0．58,P=0．669;x2=0．65,P=0．526）。结论H．pylori感染与OLP和OSCC存在一定的相关性。H．pytori感染在OLP和OSCC发生、发展中的作用有待于进一步研究。     

Immunohistochemical evaluation of expression of cytokeratin 19 in different histological grades of leukoplakia and oral squamous cell carcinoma.

Recent progress in understanding the biology of keratins together with the development of monoclonal antibodies to individual keratin proteins provide the foundation for studying the keratin expression in normal and pathological oral epithelia. Cytokeratin (CK) alterations have been reported in carcinomas and these have been associated with specific aspects of tumour behaviour. Immunohisto-chemistry with monospecific CK19 antibody was used to study the expression pattern in normal mucosa, dysplasias, and oral squamous cell carcinomas (OSCC). In non-keratinzed normal mucosa, CK19 was detected in the basal cell layer, while in dysplasias (diagnosed in H and E stained sections, mild-severe) stained strongly for CK 19 in the basal and supra basal cell layers indicating layer specificity for CK 19 expression. In OSCC, in the number of CK19 labelled cells increased from well to poorly differentiated tumour. Thus the results of the present study indicate an alteration in synthesis of keratin proteins when exposed to carcinogens.     

Immunohistochemical localization of growth factors fibroblast growth factor-1 and fibroblast growth factor-2 and receptors fibroblast growth factor receptor-2 and fibroblast growth factor receptor-3 in normal oral epithelium,epithelial dysplasias,and squamous cell carcinoma

OBJECTIVES: Fibroblast growth factors (FGFs) and their receptors (FGFRs) have been identified in a variety of carcinomas, but there are few studies concerning their presence in oral cancers. The objective of this study was to determine whether FGF-1, FGF-2, and high affinity receptors FGFR2 and FGFR3 are present in the pathogenesis of oral epithelial dysplasias and oral squamous cell carcinoma. STUDY DESIGN: Sections from formalin-fixed, paraffin-embedded samples of oral normal mucosa (n = 14), epithelial dysplasia (n = 20), carcinoma in situ (n = 10), and squamous cell carcinoma (n = 12) were tested for cytoplasmic staining by standard in situ immunohistochemistry with antibodies for FGF-1, FGF-2, FGFR2, and FGFR3. RESULTS: Staining for FGF-1 is decreased or lost in the development of epithelial dysplasia and carcinoma. Staining for FGF-2 showed increased intensity (although not statistically significant) in oral epithelial dysplasias and squamous cell carcinomas and showed a significant increased expression in the upper layers of dysplasias and stratum spinosum-like cells in squamous cell carcinomas. Staining for FGFR2 showed a statistically significant increase in intensity in all layers of epithelial dysplasias and squamous cell carcinomas. Staining for FGFR3 was found in the upper stratum spinosum cells of normal and dysplastic epithelium and well-differentiated squamous cells in squamous cell carcinomas, with a statistically significant increase in staining intensity in dysplastic and carcinomatous tissues. CONCLUSIONS: The loss of FGF-1 is consistent with loss of differentiation in dysplasias and some squamous cell carcinomas. Changes in the localization of FGF-2 and FGFR2 into upper epithelial layers with increasing dysplasia suggest increased mitotic potential of high level cells. The co-localization of FGF-2 and its high affinity receptors in neoplastic tissues suggests an autocrine mechanism of influence on carcinogenesis.     

Identification of the AgNORs, PCNA and ck16 proteins in oral lichen planus lesions

The expression of proliferating cell nuclear antigen (PCNA), cytokeratin 16 (ck16) and Ag nucleolar organizer regions (AgNORs) were assessed in 20 cases of lichen planus, 20 cases of keratosis and 20 cases of normal oral mucosa in order to evaluate the rate of keratinocyte proliferation in these tissues. Three hundred cells were counted in each sample: 100 basal cells, 100 suprabasal cells and 100 squamous cells. The mean number of AgNORs and the percentage of PCNA positive cells were calculated. Except from similar staining of suprabasal cells of lichen planus and keratosis, PCNA and AgNORs values were higher in all layers of lichen planus than in both keratosis and normal oral mucosa. The three groups showed similar ck16 immunostaining: all of the cells were positive, except those of the basal layer. The results suggest that the keratinocyte proliferation index is higher in lichen planus than in keratosis and normal mucosa. Besides, ck16 should not be used to differentiate the entities studied.     

PCNA在口腔扁平苔藓变诊断中的可行性研究

为了探讨客观、准确的方法诊断口腔扁平苔藓癌变，本研究应用S－P法显示PCNA在口腔扁平谷藓、扁平谷藓上皮异常增生及鳞癌中的分布及表达。结果可见：PCNA在不同的组织中分布及表达不同，随着病变程度的增加，PCNA分布由基底细胞层逐渐累及到棘细胞层，各组间PCNA阳性细胞百分数有显著性差异。表明将PCNA作为判断口腔扁平苦藓癌变的辅助参考指标具有一定可行性。     

口腔黏膜癌前病变及口腔鳞癌中bFGF的表达及意义

目的　探讨口腔黏膜癌前病变及口腔鳞癌的发生、发展过程中bFGF的表达及意义。方法　应用免疫组织化学方法对 10例正常口腔黏膜、2 7例口腔扁平苔藓、2 4例口腔白斑及 3 0例口腔鳞癌分别进行检测。结果　口腔鳞癌组织中bFGF高表达 ,明显高于正常口腔黏膜、口腔扁平苔藓和口腔白斑组织 (P <0 .0 5 ) ;口腔扁平苔藓和口腔白斑组织中bFGF表达高于正常口腔黏膜 (P <0 .0 5 )。结论　bFGF的过表达在口腔鳞癌的发生、发展过程中起着十分重要的作用 ,可以将其作为预测口腔黏膜恶变潜能的重要标志物     

Intraepithelial expression of perlecan, a basement membrane-type heparan sulfate proteoglycan reflects dysplastic changes of the oral mucosal epithelium

BACKGROUND: Intercellular deposition of perlecan, a major heparan sulfate proteoglycan (HSPG) of the basement membrane, is known to result in characteristic stellate reticulum-like structures in ameloblastomas or tooth germs. Although enlargement of the intercellular space is one of the histological characteristics of epithelial dysplasia of oral mucosa, the mode of expression of perlecan is poorly understood in these epithelial lesions. METHODS: Eighty-two biopsy specimens consisting of normal and hyperplastic epithelium, epithelial dysplasia, and squamous cell carcinomas were examined for both perlecan core protein and heparan sulfate (HS) chains by immunohistochemistry and in situ hybridization. RESULTS: In normal and hyperplastic epithelium, perlecan core protein and HS chains were localized in the cell border of parabasal cells and lower prickle cells, and HS chains were also found in basal cells. With an increase in the severity of epithelial dysplasia, the core protein was heavily and extensively deposited in the interepithelial space as well as in the cytoplasm of epithelial cells from the basal to the surface layers. Its gene expression was confirmed in the cells around the protein deposits. On the other hand, HS chains were enhanced in mild dysplasia, but decreased in moderate and severe dysplasias. In squamous cell carcinomas, either the core protein or HS chains were found scarcely in tumor cells but abundantly in the stromal space. CONCLUSIONS: The findings indicate that perlecan is localized in the intercellular space of the oral epithelia, and that it is over-expressed in dysplastic epithelial cells and is deposited in their interepithelial space, which results in the histology of reduction of cellular cohesion.     

Evaluation of myofibroblasts in oral epithelial dysplasia and squamous cell carcinoma

Background:  Carcinogenesis is accompanied by a number of changes in the adjacent stroma including the appearance of myofibroblasts. The purpose of this study was to evaluate and compare the presence of myofibroblasts in normal mucosa, oral epithelial dysplasia, and different grades of oral squamous cell carcinoma.
Methods:  The study sample consisted of three groups, including 40 oral squamous cell carcinomas, 15 dysplasias, and 15 sections of normal oral epithelium. Vimentin, desmin, and alpha-smooth muscle actin were used to identify myofibroblasts.
Results:  The percentage and intensity of alpha-smooth muscle actin were examined, and positive immunostaining was observed in the myofibroblasts of all squamous cell carcinomas; however these cells did not stain in the dysplasias or normal epithelium specimens. The presence of myofibroblasts was significantly higher in oral squamous cell carcinomas compared to both, dysplasias and normal mucosa cases ( P  < 0.001). A significant difference was not observed between the different grades of oral squamous cell carcinoma ( P  = 0.2).
Conclusions:  These findings show the presence of myofibroblasts in the stroma of oral squamous cell carcinoma but not dysplasia and normal mucosa, suggesting further investigation to clarify the role of myofibroblasts in the carcinogenesis of this tumor.     

Oral dysplasia and in situ carcinoma: clinicopathologic correlations of eight patients.

Eight patients with multiple oral dysplastic epithelial lesions were followed by clinical examinations and serial biopsies for periods varying from four to 22 years. The dysplasias and in situ carcinomas were characterized by persistence, recurrence, and eventual progression to invasive squamous cell carcinoma. It could not be determined whether dysplasia and in situ carcinoma were separate clinical-pathologic entities with similar end points or whether they were part of a continuum in a spectrum of epithelial neoplasia. The need for close clinical observation and local excision was emphasized because of the multiplicity of lesions and because of the protracted clinical course. Treatment of these patients was problematic because of similarities of the disease to lichen planus. It is possible that they had a premalignant disease process that mimicked lichen planus, or that they had an unusual form of lichen planus for which criteria have not been established. The progressive nature of the disease was exemplified by one death, one patient with cervical metastasis, and one with generalized remote metastatic disease.     

The immunohistology of CD40 in human oral epithelium in health and disease

BACKGROUND: CD40 has a role in the regulation of immune responses, cell proliferation and migration, and apoptosis. Little is known of its distribution in oral mucosal pathology. METHODS: Oral keratinocyte lines were tested for CD40 protein by Western blotting. Immunohistochemistry was used to stain paraffin sections of oral mucosa in health and in inflammatory, reactive, dysplastic and malignant disease. RESULTS: Western blotting confirmed the presence of CD40 in oral keratinocytes. CD40 was generally expressed by keratinocytes in the basal layer, with variable parabasal expression. Langerhans cells also stained positively. Expression was lost in nine of 33 (27%) epithelial dysplasias, seven of which were severe. Eighty-one percent of well, 69% of moderately and 50% of poorly differentiated oral squamous cell carcinomas (OSCC) expressed CD40. Overall, 45 of 65 (69%) OSCC were positive. The pattern of expression was unrelated to tumour differentiation. CONCLUSION: CD40 expression by basal and parabasal oral keratinocytes is physiological. Expression is lost in approximately one-third of oral epithelial dysplasias and OSCC. The significance of such loss remains unknown, but may be related to immunological or other abnormalities of keratinocyte homeostasis.     

MAGE-A antigens in lesions of the oral mucosa

Oral squamous cell carcinoma develops continuously out of predamaged oral mucosa. For the physician and pathologist, difficulties arise in distinguishing precancerous from cancerous lesions. MAGE-A antigens are tumor antigens that are found solely in malignant transformed cells. These antigens might be useful in distinguishing precancerous from cancerous lesions. The aim of this study was to verify this assumption by comparing MAGE-A expression in benign, precancerous, and cancerous lesions of the oral mucosa. Retrospectively, biopsies of different oral lesions were randomly selected. The lesions that were included are 64 benign oral lesions (25 traumatic lesions (oral ulcers), 13 dental follicles, and 26 epulis), 26 oral lichen planus, 123 epithelial precursor lesions (32 epithelial hyperplasia found in leukoplakias, 24 epithelial dysplasia found in leukoplakias, 26 erythroplasia with oral epithelial dysplasia, and 41 carcinomas in situ in erythroleukoplakias). The lesions were immunohistochemically stained with the poly-MAGE-A antibody 57B, and the results were compared. Biopsies of oral lichen planus, oral ulcers, dental follicles, epulis, and leukoplakia without dysplasia showed no positive staining for MAGE-A antigens. Leukoplakia with dysplasia, dysplasia, and carcinomata in situ displayed positive staining in 33%, 65%, and 56% of the cases, respectively. MAGE-A antigens were not detectable via immunohistochemistry in benign lesions of the oral mucosa. The staining rate of dysplastic precancerous lesions or malignant lesions ranged from 33% to 65%. The MAGE-A antigens might facilitate better differentiation between precancerous and cancerous lesions of the oral mucosa.     

CD146 expression in oral lichen planus and oral cancer

Clinical Oral Investigations - To examine the CD146/METCAM expression on keratinocytes in normal oral mucosa (NOM), oral lichen planus (OLP), oral epithelial dysplasia (OED), and oral squamous cell...     

Induction and upregulation of adhesion receptors in oral and dermal lichen planus

The expression pattern of well-defined cell surface adhesion receptors called VLA-family, LFA-1 and ICAM-1 was determined semiquantitatively in biopsies of oral (n = 12) and dermal lichen planus (n = 5) and compared to normal uninvolved human oral mucosa (n = 12) and skin (n = 12) using an indirect immunoperoxidase technique. In both oral and dermal lichen planus, an induction of the beta 1-integrins VLA-1 and VLA-3 and an upregulation of VLA-6 was found in T cells infiltrating the basement membrane zone. These cell surface molecules function as receptors for collagen, fibronectin and laminin. A focal induction of ICAM-1 on basal keratinocytes could be detected at sites of intramucosal T cells. These results suggest that investigated adhesion receptors are crucially involved in the aggregation of T cells in both conditions. Further investigations have to be done to determine the functional role of these adhesion receptors in lichen planus.     

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目的研究鸟氨酸脱羧酶（ODC）与口腔白斑的发生、发展乃至癌变潜能的关系。方法选取2009—2010年大连医科大学附属第一医院口腔科收治且经病理确诊的口腔白斑患者22例,以其病变黏膜组织为研究对象,同时以11例正常口腔黏膜、22例口腔扁平苔藓组织和22例口腔鳞癌组织为对照。采用免疫组化法检测ODC在各种组织中的表达,并对阳性率进行比较分析。结果在口腔正常黏膜、扁平苔藓、白斑和鳞癌组织中ODC的阳性率依次增加;ODC在口腔扁平苔藓中的阳性率高于正常黏膜,但差异无统计学意义（P〉0.05）,ODC在口腔白斑中的阳性率与正常黏膜和扁平苔藓的差异均有统计学意义（P〈0.05）,口腔鳞癌组织中ODC的阳性率显著高于其余组（P〈0.05）。结论 ODC表达程度可用于判断口腔白斑的恶变倾向及口腔鳞癌的恶性程度。     

单层上皮细胞角蛋白在口腔粘膜癌变过程中的表达

目的：探讨单层上皮细胞角蛋白ＣＫ１８和ＣＫ１９作为口腔癌前病变标志的可能性。方法：用ＬＳＡＢ免疫组化染色方法检测ＣＫ１８和ＣＫ１９在福尔马林固定,石蜡包埋的口腔正常粘膜,上皮单纯增生,轻度上皮导演增生,中度上皮异常增生,重度上皮异常增生和口腔鳞癌组织中的分布和表达强度,光镜观察染色切片,结果用秩和检验分析。     

口腔黏膜癌前病变及鳞状细胞癌中VEGF的表达及意义

目的 :探讨VEGF在口腔扁平苔藓、口腔白斑、口腔鳞状细胞癌中的表达及其在口腔鳞癌发生、发展过程中的意义。方法 :应用VEGF多克隆抗体对 10例正常黏膜、2 7例口腔扁平苔藓、2 4例口腔白斑及 30例口腔鳞状细胞癌分别进行免疫组化检测。结果 :VEGF在口腔鳞状细胞癌组织中高表达 ,与口腔扁平苔藓、口腔白斑、正常口腔黏膜组织存在着显著性差异 (P <0 .0 5 ) ;在口腔扁平苔藓和口腔白斑中VEGF有一定的表达 ,且表达强度强于正常口腔黏膜组 (P <0 .0 5 )。结论 :VEGF的过表达与口腔鳞癌的发生、发展有关 ,可以将其作为预测口腔黏膜恶变潜能的重要标志物。