Expression of apoptotic signaling proteins in leukoplakia and oral lichen planus: quantitative and topographical studies

Although the pathogenesis of leukoplakia has been unclear, carcinogenic transformation is postulated to result from alterations of apoptotic signal transduction proteins in epithelial cells. The pathogenesis of oral lichen planus (OLP) has also been unclear, but apoptotic changes of the epithelial cells in OLP have been reported. In the present study, we used a histochemical approach to describe human keratinocyte-expression of several apoptotic signaling proteins in leukoplakia, in OLP, and in normal oral mucosa as a control. Mucosal biopsies from patients with leukoplakia (n=13), OLP (n=10), and normal oral mucosa (n=9) were frozen, sectioned and immunostained with monoclonal antibodies to wild-type (wt) tumor suppressive protein p53, cyclin-dependent kinase inhibitor p21WAF1/CIP1 and the oncoproteins MDM2, and Bcl-2. Apoptosis was assessed in all cases by the TUNEL method. MDM2 and Bcl-2 expression in keratinocytes were quantitatively greater in leukoplakia than in OLP. Wt-p53 and p21WAF1/CIP1 expression was quantitatively greater in keratinocytes in OLP than in leukoplakia. Keratinocyte maturation appeared histologically normal in OLP, even though wt-p53 and p21WAF1/CIP1 were expressed in these cells. Altered keratinocyte maturation was seen in leukoplakia lesions expressing MDM2 and Bcl-2. No significant difference for the number of apoptotic epithelial cells was observed between leukoplakia and OLP, in spite of the divergent outcomes of the apoptotic signaling proteins.     

P53 and bcl-2 immunoexpression in patients with oral lichen planus and oral squamous cell carcinoma

Objective: The aim of this study was to determine by immunohistochemistry the presence and significance of p53 and bcl-2 proteins in oral lichen planus (OLP) and oral squamous cell carcinoma (OSCC). Study Design: We used 21 cases diagnosed as OLP 16 diagnosed as OSCC and four normal gingival biopsies taken from healthy patients were used as controls. Slides were processed for immunohistochemistry using anti-p53 and anti-bcl-2 monoclonal antibodies. Results: We found p53 immunoexpression in 71.4% OLP cases and 68.7% OSCC cases, with no immunoexpression in control cases. Bcl-2 was negative for all OLP and OSCC cases, and mild positivity was observed in normal tissue. We found significant correlation among p53 expression and OSCC malignancy. Conclusions: Our results suggest that TP53 system mainly promotes a hyperproliferative state by cell cycle arrest of the OLP epithelial cells for repairing damaged DNA nor apoptosis and that anti-apoptotic action of bcl-2 is not important in this disease. Key words:Oral lichen planus, oral squamous cell carcinoma, p53, Bcl-2, carcinogenesis, malignant transformation.     

Human papillomavirus infection in oral potentially malignant disorders and cancer

Human papillomavirus (HPV) infects keratinocytes in the mucosa or skin, and persistent infection with HPV may lead to premalignant lesions and invasive cancer, especially cervical cancer. It has also been hypothesized that HPV infection is an etiological factor of oral squamous cell carcinoma and oral precancerous disorders such as lichen planus, leukoplakia, and erythroplakia. A high percentage of HPV in oral lesions supports the possible viral contribution, but an association of HPV infection with these lesions remains to be established. The current paper will update the latest progress of HPV infection in several oral potentially malignant disorders and oral squamous cell carcinoma and discuss the impact of HPV infection on the progression of oral potentially malignant disorders.     

ALDH1在OLP和LK组织中的表达及其意义

目的：通过检测干细胞标记物ALDH1在口腔扁平苔藓（OLP）和白斑（LK）中的表达水平,评价其与癌变的关系。方法：采用免疫组织化学SP法检测ALDH1在10例正常口腔黏膜,30例OLP,60例LK,10例口腔鳞癌（OS－CC）中表达水平;再检测ALDH1在30例癌变与30例未癌变LK中的表达差异。结果：ALDH1在正常口腔黏膜中不表达,在OLP、LK和OSCC中的表达率分别为26．7％,63．3％和90．0％（P＜0．05）;ALDH1在未癌变和癌变LK中的表达率分别是43．3％和83．3％（P＜0．01）。结论：ALDH1与口腔黏膜恶性潜能程度相关,可能是预测癌变的分子标记物。     

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.     

Study of Biopsies from lesions of the oral mucosa by scanning electron microscopy (S.E.M.).

Surface epithelial cells from 55 biopsies including lichen planus, leukoplakia and squamous cell carcinoma were studied by scanning electron microscopy. The surface structures of the various oral lesions demonstrated morphological differences. After more information has been collected, the method of SEM may be of value for the recognition of pre- and early neoplastic lesions in the oral cavity.     

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.     

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。     

P53 and bcl-2 immunoexpression in patients with oral lichen planus and oral squamous cell carcinoma

Objective: The aim of this study was to determine by immunohistochemistry the presence and significance of p53 and bcl-2 proteins in oral lichen planus (OLP) and oral squamous cell carcinoma (OSCC). Study Design: We used 21 cases diagnosed as OLP 16 diagnosed as OSCC and four normal gingival biopsies taken from healthy patients were used as controls. Slides were processed for immunohistochemistry using anti-p53 and anti-bcl-2 monoclonal antibodies. Results: We found p53 immunoexpression in 71.4% OLP cases and 68.7% OSCC cases, with no immunoexpression in control cases. Bcl-2 was negative for all OLP and OSCC cases, and mild positivity was observed in normal tissue. We found significant correlation among p53 expression and OSCC malignancy. Conclusions: Our results suggest that TP53 system mainly promotes a hyperproliferative state by cell cycle arrest of the OLP epithelial cells for repairing damaged DNA nor apoptosis and that anti-apoptotic action of bcl-2 is not important in this disease.     

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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。     

Bcl-2、MDM2和p21在口腔白斑和口腔鳞状细胞癌中的表达水平研究

目的：了解Bcl-2、MDM2、p21在人口腔正常黏膜、白斑、鳞状癌细胞中的表达情况.方法：采用免疫组化二步法检测15例人正常口腔黏膜、15例单纯增生性白斑、10例异常增生性白斑,25例口腔鳞癌中Bcl-2、MDM2和p21蛋白的表达情况.结果：Bcl-2、MDM2蛋白在异常增生性白斑和口腔鳞癌中的表达明显高于正常口腔黏膜上皮细胞;p21蛋白在异常增生性白斑和口腔鳞癌中的表达明显低于正常口腔黏膜上皮细胞.结论：Bcl-2、MDM2和p21三者与口腔黏膜癌变具有相关性.     

Immunohistochemical analysis of the p53 tumor suppressor gene product in oral leukoplakia

Squamous Cell Carcinoma (SCC) is the most frequent malignancy in the oral cavity. p53 protein has been reported to be expressed at high levels in malignant lesions, while the level in premalignant lesions has yet to be determined. In this study, oral leukoplakia and oral SCC were examined. Seventy-four incision or excision samples from 43 cases diagnosed as leukoplakia, and 41 samples from 37 SCC cases in the oral cavity, were obtained. All samples (formalin-fixed, paraffin embedded) were examined immunohistochemically for overexpression of p53 protein with monoclonal antibody BP 53-12. As the result, 1. Twenty-two out of 43 leukoplakia cases, and 29 out of 37 oral SCC cases, were positive for p53 protein. 2. p53 protein was overexpressed in premalignant lesions, especially in the cases with moderate and severe epithelial dysplasia. 3. There was a relation between p53 protein expression and pathological features of leukoplakia (epithelial dysplasia), statistically. 4. There was a relation between p53 protein expression and clinical features of leukoplakia, statistically. 5. Malignant transformation during clinical observation was seen in 11 cases. Nine out of 11 cases were positive for p53 even before malignant transformation. Since in cancer-development cases, p53 staining was detected even before malignant transformation of oral leukoplakia to squamous cell carcinoma, it is indicated that p53 accumulation occurred at a early stage of cancer-development. In conclusion, immunohistochemical analysis of p53 protein is suggested to be useful diagnostic procedure for oral leukoplakia, which may develop into oral SCC.     

Molecular genetics of premalignant oral lesions

Oral squamous cell carcinoma (OSCC) is characterized by cellular and subcellular alterations that are associated with a progression towards dedifferentiation and growth. There are several histologically distinct lesions of the oral cavity which have malignant potential. These are leukoplakia, erythroplakia, lichen planus, and submucous fibrosis. These are characterized by a spectrum of chromosomal, genetic, and molecular alterations that they share with each other as well as with the malignant lesions that develop from them. In this review we summarize the investigation of the molecular genetics of each of these lesions and relate them to the alterations, which have been demonstrated in OSCC, to define their location on the continuum of changes, which lead to malignant transformation.     

Oct4在口腔黏膜白斑癌变进程中的作用初探

通过检测干细胞标记物Oct4在口腔黏膜白斑中的表达水平,探讨Oct4在口腔黏膜白斑癌变进程中的作用。方法：采用免疫组织化学sP法检测Oct4在10例121腔正常黏膜,40例扁平苔藓,60例白斑,20例鳞癌中表达水平;同时检测0ct4在30例癌变与30例未癌变白斑中的表达差异。结果：Oct4在正常黏膜中不表达,在扁平苔藓、白斑和鳞癌中的表达率分别为22．5％,56．7％和85．0％（P〈0.05）;Oct4在未癌变和癌变白斑中的表达率均为36.7％沪〈0.01）。结论：Oct4与口腔黏膜恶性潜能程度相关,可能直接参与白斑癌变进程。     

Oral lichen planus has a high rate of TP53 mutations. A study of oral mucosa in icelanD

Oral squamous cell carcinoma (OSCC) is a world-wide health problem. In addition to external exposure (smoking and alcohol), certain oral lesions may increase the risk of oral cancer (e.g. leukoplakia, erythroplakia, and oral lichen planus). TP53 has been implicated in OSCC, but there are limited studies of mutations in premalignant oral lesions. In this study, 55 samples from OSCC, 47 from hyperkeratotic (HK) oral mucosa, clinically diagnosed as white patches, 48 samples from oral lichen planus (OLP), and 12 biopsies from normal oral mucosa were studied immunohistochemically for expression of TP53 protein. From all the carcinoma samples and selected non-malignant samples showing moderate or strong TP53 protein expression, malignant cells or TP53-positive nuclei were microdissected and screened for mutations in exons 5-8 by constant denaturation gel electrophoresis. Moderate to strong TP53 protein staining was seen in 56% of OSCC, 32% of OLP but only in 13% of HK. All OLP samples showed a characteristic pattern of positive nuclei confined to the basal layer, whereas TP53 staining was seen in suprabasal nuclei in HK. Mutation rate was 11 out of 52 for OSCC, three out of 20 tested for HK and, remarkably, nine out 27 tested for OLP. There was no correlation between TP53 protein staining and TP53 mutations. No associations were found with anatomical sites or disease progression. The unexpectedly high mutation rate of OLP might explain the premalignant potential of this lesion.     

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     

Detection of human papillomavirus DNA in oral mucosal lesions using in situ DNA-hybridization applied on paraffin sections

The in situ DNA hybridization technique, carried out under stringent conditions, was used to detect human papillomavirus (HPV) DNA of types 6, 11, and 16 in paraffin sections of 32 surgically treated oral mucosal lesions. Expression of HPV structural proteins was analyzed by means of the immunoperoxidase (IP-PAP) method. A total of 10 (31.3%) of the 32 lesions proved to express HPV antigens, which were found in 4 of 7 squamous cell papillomas, in 2 of 2 classic condylomas, in 2 of 10 papillary hyperplasias, and in 2 of 3 leukoplakia lesions. Two of the squamous cell papillomas contained HPV 6 DNA, and 4 additional lesions were positive for HPV 11 DNA. In one of the condylomas, a double infection by HPV 6 and 11 was found, while the second was positive for HPV 11 alone. Both the HPV antigen-positive papillary hyperplasias contained HPV 6 DNA, as did the HPV antigen-positive leukoplakia lesions. Of the latter, one was infected by HPV 6 and 11. DNA of the "high-risk" HPV 16 was contained in two lesions: one lichen planus lesion and one of the two squamous cell carcinomas. The results confirm the previously reported evidence of HPV involvement in oral mucosal lesions. The implications of these findings are discussed in terms of the well-established premalignant character of oral leukoplakia and oral lichen planus, although far less commonly versus leukoplakia, with special emphasis on the discovery of the "high-risk" HPV 16 in the latter as well as in oral cancer.(ABSTRACT TRUNCATED AT 250 WORDS)     

凋亡蛋白Bcl-2、Bax在白斑、口腔扁平苔藓中的表达

目的:观察凋亡蛋白Bcl-2、Bax在白斑、口腔扁平苔藓上皮细胞中的表达,探讨其在口腔白斑、口腔扁平苔藓癌变过程中的作用机制。方法:采用免疫组化法检测10例正常口腔黏膜上皮、18例口腔扁平苔藓、23例白斑、22例口腔鳞癌上皮组织中凋亡相关蛋白Bcl-2、Bax的表达水平。结果:Bcl-2在白斑、口腔扁平苔藓上皮细胞层无异常表达,但在口腔扁平苔藓淋巴细胞浸润带过度表达。Bcl-2在鳞癌组织中呈高表达,与正常黏膜相比有显著性差异(P<0.05)。Bax在上皮单纯增生、轻度、中度不典型增生和低分化鳞癌及糜烂型口腔扁平苔藓组织中呈过度表达,与正常黏膜相比有显著性差异(P<0.05)。结论:Bax参与了口腔白斑癌变的早期事件,而Bcl-2在不典型增生转化为鳞癌的阶段并未发生作用。口腔扁平苔藓的发病机制可能与Bcl-2抑制淋巴细胞凋亡,使细胞免疫亢进,从而刺激上皮细胞Bax过度表达,诱导角朊细胞凋亡有关。     

PCNA,p53在口腔粘膜癌前病变及鳞癌中表达的免疫组化研究

应用免疫组化方法对３９例口腔粘膜癌前病变和癌中的增殖细胞核抗原，突变型ｐ５３的表达进行检测。结果表明：从ＬＰ，ＬＫ到ＩＳＣ，其ＰＣＮＡ和Ｐ５３的阳性表达均呈递增趋势。提示ＰＣＮＡ，Ｐ５３表达程度与细胞增殖程度和分化程度关系密切。     

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。