EGFR and Ki‐67 expression in oral squamous cell carcinoma using tissue microarray technology

J Oral Pathol Med (2010) 39 : 571–578 Objective: Our aim was to validate the use of tissue microarrays (TMA) in oral squamous cell carcinomas (OSCC) to analyse epidermal growth factor receptor (EGFR) and Ki‐67 expression. We also analysed the relationship that the expression of these markers may have with clinical, pathological and survival variables. Patients and methods: The study sample comprised 39 unselected patients diagnosed and treated for OSCC. We analysed Ki‐67 and EGFR expression by immunohistochemistry on formalin‐fixed, paraffin‐embedded surgical specimens. Whole sections (WS) were compared with double 1.5 mm core‐tissue microarrays. Results: High EGFR expression was observed both on TMA (in 98% of the cases) and WS (in 100% of the cases) with substantial agreement kappa value (0.720). EGFR expression was not significantly associated with clinical, pathological and survival variables on TMA and WS. Ki‐67 analysis showed a Spearman correlation of 0.741 with a Ki‐67 mean labelling index of 45% in TMA and 56.8% in WS. We found a significant relationship between gender and Ki‐67 labelling index on WS (P = 0.022) and TMA (P = 0.002). Clinical stage was the only parameter in multivariate analysis that had a significant predictive value. Conclusion: We demonstrate that dual 1.5 mm core TMA is a valid, rapid, economical and tissue‐saving way to study OSCC biopsies and that it presents strong correlation with the WS. EGFR overexpression in OSCC suggests that these tumours may be a candidate for therapy investigation directed to EGFR.     

Cyclooxygenase‐2 expression in oral tongue squamous cell carcinoma

J Oral Pathol Med (2010) 40 : 385–389 Background: Cyclooxygenase‐2 expression is associated with unfavorable outcome in various cancers, and evidence is accumulating that carcinogenesis possibly evolves from intracellular changes in response to induction of this enzyme. Today selective cyclooxygenase‐2 inhibitors are being studied and used as complement in cancer treatment. This study examined the prognostic value of cyclooxygenase‐2 expression in oral tongue squamous cell carcinoma (OTSCC). Methods: Expression of cyclooxygenase‐2 was determined in biopsies from 76 stage matched patients with OTSCC by immunohistochemistry between January 2000 and December 2004 in Stockholm, Sweden. Additionally, twelve samples taken after pre‐operative radiotherapy were investigated. Results: All OTSCC specimen expressed cyclooxygenase‐2 by immunostaining. The cyclooxygenase‐2 staining intensity increased significantly with more advanced stage (P = 0.020). Fifty percent of the surgical specimen showed a decrease in immunostaining post‐radiation (P = 0.031). No association was found with survival. Conclusion: Cyclooxygenase‐2 expression has limited prognostic value in OTSCC.     

口腔鳞状细胞癌组织中P16蛋白的免疫组化定量分析

目的　探讨P16蛋白在口腔鳞状细胞癌(鳞癌)组织中的表达及与口腔鳞癌发生发展的关系。方法　采用免疫组化S -P法,对5 6例口腔鳞癌组织、30例癌旁组织及10例正常口腔黏膜中P16蛋白的表达进行检测,应用全自动图像分析仪对染色结果进行定量测定。结果　口腔鳞癌组织中P16蛋白的表达量低于癌旁组织及正常黏膜(P <0 .0 5 ) ,口腔鳞癌Ⅲ级低于Ⅰ、Ⅱ级(P <0 .0 5 )。颈淋巴结转移组低于无颈淋巴结转移组(P <0 .0 5 )。结论　P16蛋白在口腔鳞癌组织中的表达常为缺失,说明其作为抑癌基因在口腔鳞癌的发生中起作用。P16蛋白的表达程度越低,分化越差,肿瘤越易于浸润和转移。     

Fos‐related activator‐1 is overexpressed in oral squamous cell carcinoma and associated with tumor lymph node metastasis

J Oral Pathol Med (2010) 39 : 470–476 Background: Previously, we established an in vitro cellular carcinogenesis model of oral squamous cell carcinoma (OSCC), including the human immortalized oral epithelia cells (HIOECs) and its derived cancerous HB cells. Then, expression microarray analysis showed that the gene encoding fos‐related activator‐1 (Fra‐1) was significantly upregulated in the cancerous HB cells compared with HIOECs. Methods: To confirm the expression of Fra‐1 at mRNA and protein levels by real‐time PCR and western blotting analysis in the carcinogenesis model of OSCC and CAL27 cells. To investigate Fra‐1 expression in clinical samples from 30 primary OSCC patients by immunohistochemistry. Results: Fra‐1 expression was increased both at mRNA and protein levels in this carcinogenesis model of OSCC and CAL27 cells. Nuclear and cytoplasmic Fra‐1 protein expressions both increased in the cancerous tissues compared with those in the paired adjacent non‐malignant epithelia (nuclear: P < 0.001, cytoplasmic: P = 0.003). A higher level of nuclear Fra‐1 expression was seen in the tumor samples of patients with lymph node metastasis than those without lymph node metastasis (5.07 ± 1.33 vs 3.81 ± 1.33, P = 0.023). Higher level of Fra‐1 expression was also found in the tumor invasive margin than tumor center. Conclusions: Fra‐1 is a positive gene of OSCC development and progression, Fra‐1 can be used as a potential therapeutic target gene and an additional marker for evaluation of lymph node metastasis.     

Immunohistochemical study of the expression of fatty acid synthase and Ki‐67 in salivary gland tumors

J Oral Pathol Med (2010) 40 : 467–475 Background: The enzyme fatty acid synthase plays a fundamental role in the biosynthesis of fatty acids. Several recent studies have demonstrated a high fatty acid synthase expression in malignant tumors. Few studies have been conducted in oral and salivary gland tumors describing the fatty acid synthase expression. Methods: This study evaluated and compared, by immunohistochemical reaction, the expression of fatty acid synthase and Ki‐67 in salivary gland tumors. The immunohistochemical study used the streptavidin–biotin–peroxidase technique, with antibodies anti‐fatty acid synthase and anti‐Ki‐67. The fatty acid synthase was analyzed by scores, considering the intensity of labeling, quantity of labeled cells and histological component. The Ki‐67 was analyzed by counting of one thousand cells, calculating the quantity of positive cells in regions with higher density of labeling. Statistical analysis was performed using the Pearson and Mann–Whitney correlation tests. Results: There was greater fatty acid synthase expression in pleomorphic adenoma compared to other tumors, and predominance of Ki‐67 in malignant tumors. Among these, the mucoepidermoid carcinoma presented the highest proliferation rate. The expression of fatty acid synthase and Ki‐67 did not present correlation between the tumors analyzed, except in pleomorphic adenomas, with statistically significant relationship between them. Conclusion: It is suggested that the fat metabolism in salivary gland tumors is related to maintenance of cell differentiation. Its expression prevailed in benign tumors, while Ki‐67 prevailed in the mucoepidermoid carcinoma, demonstrating its high proliferation rate, followed by the cystic adenoid carcinoma and polymorphous low grade adenocarcinoma.     

Association of macrophages with angiogenesis in oral verrucous and squamous cell carcinomas

J Oral Pathol Med (2010) 39 : 559–564 Background: Tumor‐associated macrophages (TAMs) are a major cellular component of human cancers, yet there is still no consensus as to their role in cancer growth and angiogenesis. Methods: The association between TAMs and angiogenesis was investigated in formalin‐fixed, paraffin‐embedded archival material of oral squamous cell carcinoma (OSCC) and oral verrucous carcinoma (OVC). TAMs shown by immunohistochemistry for CD68 and microvessels demonstrated by immunohistochemistry for CD31 were quantified using an image analyzer computer system. Results: TAMs were observed in all studied specimens. The area percentage of CD68 immunoreactivity and microvessel density (MVD) were significantly lower in OVC compared with the different grades of OSCC (P = 0.0009), (P = 0.0045). Both parameters increased in high‐grade malignancy of OSCC. Linear regression analysis revealed a positive correlation between the area percentage of CD68 immunoreactivity and the MVD in the studied tumors. Conclusions: Increased TAMs is associated with angiogenesis and higher histopathological grades in oral cancer.     

Id－1和Ki67在TSCC中的表达及其临床意义

目的：探讨舌鳞状细胞癌（TSCC）组织中分化抑制因子－1（Inhibitor of differentiation－1,Id－1）和Ki67的表达,及其与临床病理参数的关系。方法：应用免疫组织化学SP法检测65例TSCC中Id－1,Ki67的表达。结果：65例TSCC中Id－1有43例（66．2％）阳性表达,36例Ki67（55．4％）阳性表达。 Id－1和Ki67阳性表达与TSCC病理分级,淋巴结转移及TNM分期显著相关（P＜0．05,x2检验）,与性别、年龄无关（P＞0．05,x2检验）,Ki67阳性表达还与肿瘤大小有关（P＜0．05,x2检验）。Id－1和HIF－1α阳性表达呈正相关（P＜0．05,x2检验）。结论：Id－1和Ki67与TSCC的分化,增殖和转移密切相关,在其发生发展过程中扮演重要角色。联合检测Id－1和Ki67的表达有利于TSCC病程分级,对其临床治疗及预后评估能够提供有效帮助。     

PCNA/AgNOR and Ki-67/ AgNOR double staining in oral squamous cell carcinoma

This study was performed on oral squamous cell carcinomas (OSCC) in order to investigate the relation between the number of interphase silver-stained nucleolar organizer regions (AgNORs) and the immunolabeling of proliferation-associated markers, using antibodies to Ki-67 and proliferating cell nuclear antigen (PCNA). Fifteen consecutive cases of oral squamous cell carcinoma were used and a double staining technique was performed in order to quantify the number of NORs in PCNA-positive and -negative cells as well as in Ki-67-positive and -negative cells. Our results showed a higher mean number of AgNORs in PCNA- and Ki-67-positive cells than in PCNA- and Ki-67-negative cells. We concluded that there is an association between cell proliferation and AgNOR score in OSCC.     

Actual proliferating index in oral squamous cell carcinoma and leukoplakia

Aim: To examine the possible association between epithelial proliferation and disease progression in the oral mucosa using the actual proliferation index. Methods: The actual proliferation index was measured by the Ki‐67 labeling index and argyrophilic nucleolar organizer region count per nucleus. Immunohistochemistry was carried out for Ki‐67 by using the molecular immunology borstel‐1 clone in 20 leukoplakias, 20 oral squamous cell carcinomas, and 10 normal oral mucosae. Results: The argyrophilic nucleolar organizer region count per nucleus, Ki‐67 labeling index, and actual proliferation index were significantly higher in oral squamous cell carcinoma, followed by leukoplakia and normal oral mucosa. Leukoplakia with dysplasia showed a significantly higher Ki‐67 labeling index and actual proliferation index, compared to leukoplakia without dysphasia. There was a significant correlation of Bryne’s histological malignancy grading with the argyrophilic nucleolar organizer region count and the Ki‐67 labeling index. There was a significant positive correlation between the argyrophilic nucleolar organizer region count and the Ki‐67 labeling index among all groups. Conclusions: Leukoplakia or suspected epithelial dysplasia should be stained for argyrophilic nucleolar organizer regions and Ki‐67. The actual proliferation index is not only useful as a prognostic factor, but could also be a promising treatment determining modality for patients with premalignant and malignant lesions.     

Survivin在口腔鳞癌组织中的表达及其与COX-2相关性研究

目的:探讨凋亡抑制基因Survivin在口腔鳞癌组织中的表达及其与COX-2表达的相关性。方法:运用S-P免疫组化技术,检测Survivin和COX-2蛋白在50例口腔鳞癌组织、10例正常口腔黏膜组织中的阳性率。结果:Survivin蛋白在10例正常口腔黏膜组织中呈阴性,而在50例口腔鳞癌组织中有42例阳性,占84%,差异有极显著性差异(P<0.01),COX-2蛋白在口腔鳞癌组织中的阳性率为86%(43/50)。Survivin蛋白阳性率与年龄和性别不相关,而与COX-2蛋白阳性呈正相关(P<0.05)。Survivin蛋白在低分化癌组织中的表达比在高分化癌中高,但在统计学上无明显差异性。结论:肿瘤组织中Survivin蛋白的高阳性表达对口腔鳞癌的发生发展起重要作用;COX-2蛋白在口腔鳞癌组织中也有高阳性表达,并与Survivin有相关性。两者可能存在共同的激活机制,从而抑制口腔鳞癌细胞的凋亡。     

口腔鳞癌脉管侵袭的研究

对４４例口腔鳞癌组织切片用抗血型抗原ＡＢＨ抗体作免疫组化染色，以研究癌组织对脉管的侵袭。结果表明，该方法为脉管内皮细胞提供了清晰的染色，显示脉管侵袭的阳性率为４０．９％，显著高于以Ｈ·Ｅ染色显示的阳性率（２０．５％）。口腔鳞癌组织侵袭脉管与其分化程度、生长方式及淋巴结转移有密切关系。作者提示口腔鳞癌脉管侵袭的研究对判断预后有重要意义     

口腔鳞癌组织中Ki-67和p53蛋白的表达

目的 探讨口腔鳞癌组织中Ki-67和p53蛋白的表达及其与临床病理特征的关系。方法采用免疫组织化学S-P法对10例正常口腔黏膜组织、16例口腔白斑（OLK）组织、48例口腔鳞癌（OSCC）组织中的Ki-67和p53蛋白表达进行检测,结合患者临床病理资料进行分析,使用SPSS17.0 软件包对数据进行统计学处理。结果Ki-67蛋白在正常口腔黏膜组织、口腔白斑和口腔鳞癌组织中的阳性表达率分别为30.0%、56.3%和79.2%;p53的阳性表达率分别为0.0%、43.8%和70.8%,Ki-67和p53在正常黏膜组与口腔白斑和口腔鳞癌组差异均具有显著性（P<0.05）;Ki67蛋白在口腔鳞癌组织中的表达与肿瘤的临床分期、分化程度、有无淋巴结转移有关（P<0.05）,p53蛋白的表达与肿瘤的分化程度有关(P<0.05);Ki-67和p53蛋白在口腔鳞癌组织中的表达呈正相关（P<0.05）。结论Ki-67和p53蛋白在口腔鳞癌组织中高表达,可能在口腔鳞癌的发生、发展过程中起着重要作用。