Differential expression of stem cell-like proteins in normal,hyperplastic and dysplastic oral epithelium

Objective

The identification of stem cells (SC) remains challenging. In the human oral mucosal epithelium, these cells are believed to be in the basal layer (stem cell niche), but their exact location is unclear. The aim of this study was to examine the dysplastic oral epithelium for these SC-like proteins in order to assess their diagnostic value as biomarkers complementing the histological grading of dysplasia.

Material and Methods

Thirty oral epithelial dysplasia (OED), 25 oral lichen planus (OLP), 10 oral hyperkeratosis and 5 normal oral epithelium (OE) were immunohistochemically examined for four SC markers [integrin β1, neuron-glial-2 (NG2), notch 1 (N1) and keratin 15 (K15)].

Results

Three of four SC markers were heterogeneously detected in all samples. K15 overexpression in the lower two-thirds of severe OED suggests an expanded SC niche. Integrin β1 distribution pattern was not measurably different between OEDs and control. NG2 was almost negative to absent in all samples examined. N1 expression was weak and highly variable in normal and dysplastic epithelium, making it an unreliable epithelial stem cell marker.

Conclusions

Present findings suggest that these markers were unable to identify individual epithelial stem cells. Instead, subpopulations of cells, most probably stem cells and transit amplifying cells with stem cell-like properties were identified in the dysplastic oral epithelium. The characteristic expressions of K15 might be of diagnostic value for oral dysplasia and should be investigated further.     

Th1/Th2细胞在口腔扁平苔藓发病中的研究

###############################################################################################################################################################################################################################################################     

Expression of TGF-beta1, Smad7 and cell apoptosis in epithelium of oral lichen planus

OBJECTIVE: To examine the expression of TGF-beta1, Smad7 and cell apoptosis in oral lichen planus (OLP) and to evaluate the possible pathogenesis of oral lichen planus. METHODS: Immunohistochemical technique was used to study the expression of TGF-beta1 and Smad7 in the epithelia cells of 17 OLP cases and 7 normal oral mucosa (NOM). TUNEL was used for detecting the cell apoptosis in 17 OLP cases and 7 NOM. RESULTS: TGF-beta1 was moderately positive in the epithelia cells of OLP. All the epithelia cells in OLP showed strong cytoplasmic staining. The expression of TGF-beta1 and Smad7 were significantly increased in OLP compared with that in NOM (P < 0.05). Cell apoptotic index (AI) was remarkably increased in epithelia cells in OLP cases, and the cell apoptosis was localized in basal and suprabasal epithelial layers. There was a positive correlation between TGF-beta1 expression and cell apoptosis in the epithelia of OLP (r = 0.69, P <0.05). CONCLUSIONS: High expression of TGF-beta1 and Smad7 in the epithelia of OLP suggests that TGF-beta1-Smad7 signal pathway was disturbed in oral lichen planus. The imbalance of TGF-beta1-Smad7 pathway may contribute to the mechanisms of cell apoptosis of epithelial cells in OLP.     

VCAM-1 and ICAM-1 are expressed by Langerhans cells, macrophages and endothelial cells in oral lichen planus

Expression of intercellular adhesion molecule-1 (ICAAM-1, CD54) and vascular cell adhesion molecule-1 (VCAM-l, CD106) was examined in oral lichen planus (OLP) and normal oral mucosa (NOM). Immunoperoxidase staining showed ICAM-1 expression by vascular endothelium in all biopsies of OLP and NOM whereas endothelial VCAM-l staining was found in 2/7 NOM and 8/9 OLP. In the lamina propria of NOM occasional cells were ICAM-1 or VCAM-l positive, and virtually no staining of intraepilhelial dendritic cells was seen for either marker. Intraepithelial dendritic cells stained for ICAM-1 in 7/9 and VCAM-1 in 4/9 OLP biopsies. Double immunofluorescence showed dual labelling of Langerhans cells (LC) with CD1a and VCAM-l in a further 5/12 cases of OLP, but there was no such staining in four NOM. This is the first report of LC staining with VCAAM-l. Induction of ICAM-1 and VCAM-l on LC and macrophages in OLP suggests these cells are activated and may contribute to the pathogenesis of OLP by presenting antigen to infiltrating lymphocytes.     

Expression of caspase-3 and structural changes associated with apoptotic cell death of keratinocytes in oral lichen planus

OBJECTIVE: Apoptosis appears to be the mode of cell death by which damaged cells are removed from the lesional tissue. The aim of this study was to examine keratinocyte apoptosis and caspase-3 (CPP32) expression in oral lichen planus (OLP). MATERIALS AND METHODS: Paraffin-embedded samples of OLP (n = 30) and normal oral mucosa (NOM; n = 5) were prepared for haematoxylin-eosin (H & E), immunohistochemistry and electron microscopy. The number of apoptotic cells and the proportion of total cells that were either apoptotic (apoptotic index; AI) or mitotic (mitotic index; MI) were assessed in H & E stained sections. An immunostaining-intensity-distribution index (IIDI; proportion of stained cells x staining intensity) was used to assess CPP32 immunoreactivity. RESULTS: Results showed a significant increase in the number of apoptotic cells in OLP (P < 0.001). In OLP, all apoptotic bodies were found in the basal and prickle epithelial layers. Compared with NOM, the AI was significantly greater in atrophic (P < 0.05), reticular (P < 0.001) and plaque-like (P < 0.01) OLP. The MI was significantly greater in plaque-like OLP (P < 0.01). The proportion of CPP32-positive cells and the IIDI were significantly greater in all forms of OLP compared with NOM (P < 0.05). No difference in CPP32 expression was evident between clinical forms of OLP. Electron microscopy confirmed the light microscopic finding of apoptosis. CONCLUSION: Keratinocyte apoptosis and caspase-3 expression co-localized to the basal and parabasal epithelial layers, suggesting that proliferating epithelial cells may be targeted for destruction in OLP. Differences in epithelial AI and MI may underlie the various clinical and histological appearances of OLP.     

GP—PCR检测口腔粘膜中HPV的研究

为了解口腔粘膜中人乳头瘤病毒的感染情况，诈者采用通用引物介导的聚合酶链反应（GP－PCR）技术，对临床正常口腔粘膜（NOM），口腔扁平苦藓（OLP），口腔鳞状细胞癌（OSCC）中的人乳头瘤病毒（HPV）进行检测。结果：30例口腔粘膜中，HPVDNA阳性率43．3％，其中NOM28％，OLP46．7％、（JSCC57．1％。其结果表明：临床正常和病变口腔粘膜中存在多种型别的HPVDNA．此外，GP可作为普查口腔粘膜HPV的较好方法。     

FOXP3和GITR在口腔扁平苔藓及口腔鳞状细胞癌中的表达及意义

目的研究FOXP3、GITR在口腔扁平苔藓（OLP）和口腔鳞状细胞癌（OSCC）组织中的表达及其意义。方法采用免疫组化方法检测30例OLP组织、30例OSCC组织和15例口腔正常黏膜（NOM）组织中FOXP3、GITR的表达。结果NOM组FOXP3和GITR阴性表达;OLP组和OSCC组FOXP3和GITR表达分别较NOM组显著增加（P〈0.01）;OLP组和OSCC组之间FOXP3和GITR表达无显著性差异（P〉0.05）;FOXP3和GITR在OLP组和OSCC组中的表达存在正相关关系。结论在OLP的发生及发展成OSCC过程中,FOXP3和GITR均可能起着一定的作用。     

HERG1在口腔扁平苔藓和口腔鳞状细胞癌中的表达

目的：研究钾离子通道蛋白HERG1在正常口腔黏膜（normal oral mucosa,NOM）、口腔扁平苔藓（oral lichen planus,OLP）、口腔鳞状细胞癌（squamous cell carcinoma,OSCC）中的表达及意义。方法：免疫组织化学技术检测16例NOM、20例OLP、30例OSCC组织中HERG1的表达。采用χ2检验,P〈0.05为差异具有统计学意义。结果：HERG1在OSCC中的表达强度高于OLP（P〈0.05）,HERG1在OLP组织的表达高于正常组织（P〈0.05）,且糜烂型OLP中的表达高于非糜烂型OLP（P〈0.05）,差异均具有统计学意义。结论：HERG1可能与OLP及OSCC的发生发展有一定的关系。     

T cell antigen receptor expression by intra-epithelial lymphocytes in oral lichen planus

The distribution of T lymphocytes expressing the αβ or γδ heterodimer of the T cell receptor (TCR) was examined in normal oral mucosa (NOM) and reticular oral lichen planus (OLP) using a panel of antibodies specific for CD3, the αβ TCR and the γδ TCR. Intra-epithelial lymphocytes were counted and epithelial surface length was measured by image analysis. T cells in the lamina propria were not quantified. Total intra-epithelial lymphocytes were increased in OLP compared with NOM (p=0.0004). The proportions of cells expressing the γδ TCR in NOM and OLP were 10% and 9.3%, respectively, suggesting there is no selective recruitment from the circulation of either αβ or γδ TCR-bearing cells into normal oral epithelium or that affected by OLP The role. if any, of γδ T cells in the pathogenesis of OLP remains to be determined.     

Activation of nuclear factor-kappa B correlates with tumor necrosis factor-alpha in oral lichen planus: a clinicopathologic study in atrophic-erosive and reticular form

Backgroud:  Nuclear factor-kappa B (NF-κB) is believed to be involved in the pathogenesis of various inflammatory diseases, including oral lichen planus (OLP). The objective of the present study was to investigate the possible relationship between NF-κB activation and expression of tumor necrosis factor-alpha (TNF-α) in OLP and their expression pattern in relation to several clinical features.
Methods:  Thirty OLP cases were divided into atrophic-erosive form (14 cases) and reticular form (16 cases) according to their clinical manifestations. The expression of NF-κB p65 and TNF-α of both two groups were investigated by immunohistochemical staining, and the percentage of positive cells was calculated in each case. Biopsies of 10 normal oral mucosa (NOM) also underwent the same procedure as controls.
Results:  Nuclear factor-kappa B p65 nuclear staining was found in nuclei of basal and suprabasal epithelial keratinocytes in OLP, however, no positive staining was found in NOM. Positive TNF-α staining was detected in cytoplasm of basal epithelial keratinocytes in OLP, and only scattered staining was detected in NOM. Expression of NF-κB p65 and TNF-α were significantly different with respect to clinical forms and lesion sites ( P  < 0.05), except for genders ( P  > 0.05) in 30 OLP cases. NF-κB nuclear staining positively correlated ( r  = 0.676, P  < 0.01) with TNF-α overexpression in OLP.
Conclusions:  Nuclear factor-kappa B activation and its correlation with overexpression of TNF-α may play an important role in pathogenesis of OLP. There might be a positive regulatory loop between NF-κB and TNF-α, which may contribute to inflammation in OLP; NF-κB may also protect epithelial keratinocytes from excessive apoptosis.     

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...     

Expression of keratins 8, 18, and 19 in epithelia of atrophic oral lichen planus

Keratins form intermediate filaments of the cytoskeleton in keratinocytes and have roles in cell structure, signaling, intracellular transport, and cell death. Oral lichen planus (OLP) is an oral inflammatory disease with derangements in basal keratinocytes and disruption of the basal membrane. Here, we focused on epithelial expression of keratins 8, 18, and 19 because these proteins are known to modulate cell death. Biopsies were taken from buccal oral mucosa of persons with normal oral mucosa (n = 10) or atrophic OLP (n = 10). Cultured normal oral keratinocytes (n = 4) showed expression of mRNA and protein for keratins 8, 18, and 19. Immunohistochemistry showed consistent staining for keratins 8 and 18 in basal keratinocytes of normal oral mucosa. In OLP, staining for keratin (K)8 was mostly negative and staining for K18 was weak. Keratin 19 was expressed irregularly in most biopsies of normal oral mucosa and not at all in OLP. Several mononuclear leukocytes in the cellular infiltrate showed membrane staining for K8 and K18. Positive staining for K16 confirmed partial collapse of the basal cell layer in OLP. The basal cell niche in OLP therefore appeared to be partly populated with keratinocytes demonstrating a higher degree of differentiation (K8⁻ K18⁻ K19⁻ K16⁺); consequently, such areas may be more susceptible to the action of cell death factors released from the cell infiltrate as a result of lacking the protective, normal keratin present in the basal epithelial cell layer of normal oral mucosa.     

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.     

口腔扁平苔藓中人乳头瘤病毒的检测

为研究口腔扁平苔藓（OLP）与人乳头瘤病毒（HPV）的关系．作者采用聚合酶链（PCR）技术，对22例OLP中的HPV低危型──11型以及高危型—16型DNA分别进行了检测，并以10例正常口腔粘膜（NOM）作为对照。结果表明：PCR是检测OLP中HPV较为理想的方法。OLP中的HPV与NOM相比较，11型相对升高。OLP各临床分型中，网纹型的HPV阳性百分比最高，该型的病程迁延．治疗后易复发，可能与HPV的潜伏和感染有关，4例糜烂型，即有2例含有HPV16，提示糜烂型与癌变的关系。HPV在NOM中的存在。表明HPV可以无症状或以潜伏的形式存在于口腔粘膜。     

K_v3.4在口腔扁平苔藓和口腔鳞状细胞癌中的表达

目的:研究钾离子通道蛋白kv3.4在正常口腔黏膜(normal oral mucosa,NOM)、口腔扁平苔藓(oral lichen planus,OLP)、口腔鳞状细胞癌(squamous cell carcinoma,OSCC)中的表达及意义。方法:免疫组织化学技术检测16例NOM、20例OLP和30例OSCC组织中kv3.4的表达,采用Wilcoxon秩和检验,α=0.05。结果:kv3.4在OSCC中的表达强度高于OLP(P<0.05),kv3.4在OLP组织的表达高于正常组织(P<0.05),且糜烂型OLP中的表达高于非糜烂型OLP(P<0.05),差异均具有统计学意义。结论:kv3.4可能与OLP及OSCC的发生、发展有一定的关系。     

p63在口腔扁平苔藓中的表达

目的探讨p53家族新成员p63在口腔扁平苔藓（OLP）中的表达，其与OLP发生、发展的关系以及是否可作为一种标志物预测OLP的转归。方法采用免疫组织化学方法检测30例012中p63蛋白的表达，同时采用RT—PCR方法分析p63编码的两大类转录产物TAp63和ANp63的mRNA水平，并且与口腔鳞状细胞癌（OSCC）及正常口腔黏膜（NOM）进行比较。结果p63蛋白在NOM、OLP和OSCC组中均有表达，与NOM比较，其累积光密度（IOD）值在OLP中下调，在OSCC中明显上调。在糜烂萎缩型OLP中，p63蛋白表达高于非糜烂型（P〈0．001）。RT-PCR检测结果显示，TAp63的mRNA水平在OLP中表现为中等。略低于NOM．明显高于OSCC；ANp63的mRNA水平在NOM和OLP中极低．其转录扩增带大多数检测不到．但在OSCC组都能检测到．而且呈高水平表达。TAp63的mRNA水平在糜烂萎缩型OLP中低于非糜烂型（P=0．018），ANp63则高于非糜烂型（P=0．049）。结论p63与OLP的发病机制有关，其中TAp63和ANp63分别起不同的作用。p63表达的高低可能可以作为预测OLP是否有高风险癌变的检测指标。     

IL-2、IL-1O在口腔扁平苔藓发病中的作用

目的：通过对口腔扁平苔藓（OLP）局部组织及外周血中IL-2、IL-10表达进行研究,进-步探讨OLP发病机制。方法：应用免疫组织化学SABC法检测30例OLP中IL-2、IL-10的表达数量及分布,同时用ELISA法检测外周血中IL-2、IL-10的含量。结果：①在病损组织中,OLP组中IL-2、IL-10表达显著高于对照组（P〈0．05）。IL-2主要分布在淋巴细胞浸润带,在基底膜附近较集中;IL-10主要分布在固有层,远离基底膜。②在外周血中,糜烂型OLP组IL-2、IL-10含量明显高于对照组和非糜烂型OLP组（P〈O．05）。结论：①根据IL-2、IL-10在OLP中表达和分布情况,提示IL-2、IL-10在OLP发病机制中参与了机体免疫的调节和介导作用。②外周血中IL-10高表达,提示糜烂型OLP患者全身的免疫反应参与了本病的形成或可能伴有全身免疫系统功能紊乱。     

Overexpression of cdk4 and p16 in oral lichen planus supports the concept of premalignancy

J Oral Pathol Med (2011) 40 : 294–299 Background: Cell cycle arrest and increased cell proliferation have been demonstrated in oral lichen planus (OLP). This study evaluated the expression of cdk4, cdk6 and p16, important proteins in the G1 phase, in OLP and compared the expression of these proteins of OLP with those of normal mucosa. Methods: Expression of cdk4, cdk6 and p16 were investigated in 23 OLP and 10 normal mucosae using immunohistochemistry technique. Positive cells were counted and presented as a percentage of positive cells. Results: Expression of cdk4, cdk6 and p16 was observed in 3/10 (30%), 1/10 (10%) and none of normal mucosa, respectively. Expression of cdk4, cdk6 and p16 was detected in 18/23 (78.3%), 8/23 (34.8%) and 15/23 (65.2%), of OLP, respectively. The numbers of cdk4 and p16 positive cases of OLP were significantly higher than normal mucosa. In normal mucosa, the averages of the percentage of positive cells for cdk4 and cdk6 staining were 1.48 and 0.18, respectively. In OLP, the averages of the percentage of positive cells for cdk4, cdk6 and p16 staining were 2.73, 1.06 and 2.24, respectively. The percentage of cdk4‐positive cells of OLP was significantly higher than those of normal mucosa group. Conclusion: Oral lichen planus demonstrated overexpression of cdk4 and p16, but not cdk6, suggesting that epithelial cells in OLP are in the hyperproliferative state and in cell arrest. Altered expression of cdk4 and p16 provides evidence of the malignant potential in OLP.     

TL1A和IDO在口腔扁平苔藓患者外周血中的表达研究

目的 研究肿瘤坏死因子样配体1A（tumor necrosis factor-like ligand 1A,TL1A）和吲哚胺2,3双加氧酶（indoleamine 2,3-dioxygenase,IDO）在口腔扁平苔藓（OLP）患者外周血中的表达及其是否存在相关性。方法 采用实时荧光定量聚合酶链反应（FQ-PCR）法检测30例OLP患者和30例健康人外周血单个核细胞中TL1A、IDO的基因表达水平。结果 在OLP患者中,TL1A、IDO的基因表达水平均高于对照组（P＜0.05）,结果经2-ΔΔCT转换后,OLP组TL1A、IDO mRNA表达水平分别是健康对照组的1.7872、2.0763倍。TL1A、IDO mRNA的表达水平无明显相关性（P>0.05）。结论 TL1A、IDO mRNA在OLP的发病中起了一定作用,两者的作用机制有待于进一步研究。