Oral Mucosa Tissue Samples Research Articles

Investigating the Association Between Fusobacterium nucleatum and Oral Squamous Cell Carcinoma: A Pilot Case-Control Study on Tissue Samples.

Background Fusobacterium nucleatum (F. nucleatum)has been increasingly linked to oral squamous cell carcinoma (OSCC), prompting this study to explore its presence using polymerase chain reaction (PCR) and evaluate its clinical significance. Methods In this pilot case-control study, 12 OSCC tissue samples and 12 non-cancerous oral mucosal tissue samples were analyzed. Total RNA extraction and complementary DNA (cDNA) synthesis were performed using Trizol-based methods, followed by PCR amplification and gel electrophoresis. The clinical characteristics of participants and PCR results were recorded. Results Among the OSCC tissue samples, three out of 12 tested positive forF. nucleatum,while none of the control samples showed its presence. The detection rate ofF. nucleatumin OSCC was 25%. Gel analysis confirmed specific amplicon amplification, and ImageJ software enabled copy number quantification. Discussion Our findings support previous research indicating a potential association betweenF. nucleatumand OSCC. Understanding the etiological significance ofF. nucleatumin OSCC has clinical implications, including early detection, risk stratification, and prognostication. However, the limited sample size and the need for further research to elucidate underlying mechanisms are acknowledged. Conclusion This pilot study provides initial evidence ofF. nucleatum'spresence in a subset of OSCC samples, supporting its potential association with oral cancer. DetectingF. nucleatumin OSCC tissues holds promise for future research and clinical applications as a diagnostic and prognostic biomarker. Understanding its role in oral carcinogenesis will facilitate the development of targeted therapeutic strategies. Larger studies are warranted to validate these findings and investigate the precise mechanisms involved.

DJ-1 Oncogene as a Potential Diagnostic and Prognostic Biomarker for Head and Neck Cancer.

Background Current methods used to diagnose and prognosticate oropharyngeal cancer have contributed to unfavorable patientsurvival rates that have not significantly improved for the last several decades. Precision medicine oncology relies on molecular diagnostics and biomarkers to supplement existing methods of detecting and prognosticating cancers. This study evaluated the expression of DJ-1, an oncogene that is implicated in the pathogenesis of oral squamous cell carcinoma (OSCC), the most common type of head and neck cancer, to determine its utility as a diagnostic and prognostic biomarker. Methodology Immunohistochemistry (IHC) was performed on 13 normal oral mucosa tissue samples and 143 OSCC tissue samples of varying histopathological grades. Computer-assisted image analysis was performed using the Aperio ImageScope software from Leica Biosystems (Buffalo Grove, IL), which utilizes an algorithm of positive pixel counting for the quantification of immunoreactivity and the percentage of positive cell staining, generating a histo-score (H-score). The comparisons of the average H-scores of the different groups were made using a two-tailed T-test with P ≤ 0.05 set as the level of significance. Results The study found a significant increase in DJ-1 expression in oral squamous cell carcinoma tissue samples in comparison to the normal oral mucosa tissue samples. Additionally, the study documented a significant upregulation in DJ-1 expression in the OSCC tissue samples with high histopathological grades compared to the OSCC tissue samples with low histopathological grades. Conclusions DJ-1 expression patterns were able to reliably differentiate between oral squamous cell carcinoma and the normal counterpart tissues of the oral mucosa, thereby highlighting its role as a potential diagnostic biomarker. Moreover, DJ-1 expression significantly correlates with the OSCC histological grade, which serves as an indicator of the differentiation status and a predictor of the biological behavior of malignant neoplasms, adding to DJ-1's potential utility as a prognostic biomarker for this common type of head and neck cancer.

The Role of the Oral Microbiota in the Etiopathogenesis of Oral Squamous Cell Carcinoma.

Dysbiosis in the oral environment may play a role in the etiopathogenesis of oral squamous cell carcinoma (OSCC). This review aims to summarize the current knowledge about the association of oral microbiota with OSCC and to describe possible etiopathogenetic mechanisms involved in processes of OSCC development and progression. Association studies included in this review were designed as case–control/case studies, analyzing the bacteriome, mycobiome, and virome from saliva, oral rinses, oral mucosal swabs, or oral mucosal tissue samples (deep and superficial) and comparing the results in healthy individuals to those with OSCC and/or with premalignant lesions. Changes in relative abundances of specific bacteria (e.g., Porphyromonas gingivalis, Fusobacterium nucleatum, Streptococcus sp.) and fungi (especially Candida sp.) were associated with OSCC. Viruses can also play a role; while the results of studies investigating the role of human papillomavirus in OSCC development are controversial, Epstein–Barr virus was positively correlated with OSCC. The oral microbiota has been linked to tumorigenesis through a variety of mechanisms, including the stimulation of cell proliferation, tumor invasiveness, angiogenesis, inhibition of cell apoptosis, induction of chronic inflammation, or production of oncometabolites. We also advocate for the necessity of performing a complex analysis of the microbiome in further studies and of standardizing the sampling procedures by establishing guidelines to support future meta-analyses.

Analysis of HRAS mutations in Japanese patients with oral squamous cell carcinoma

ObjectivesThe incidence of oral squamous cell carcinoma (OSCC) is associated with chewing tobacco and heavy consumption of alcohol. OSCC is also associated with Harvey rat sarcoma viral proto-oncogene homolog (HRAS) gene mutations, which are known to be high-risk factors for poor survival outcomes in patients with OSCC. In this study, we analysed HRAS mutations in Japanese patients with OSCC. MethodsTumours and oral mucosal tissue samples were collected from 84 treatment-naïve patients with OSCC, between April 2010 and November 2012 in the Department of Oral and Maxillofacial Surgery at a university hospital in Japan. ResultsAn HRAS mutation was identified in one of the 84 tumours (1.2%) using standard Sanger sequencing. This mutation occurred in codon 13 (c.38G > T) of exon 2, and it corresponded to amino acid substitution p.Gly13Val. Statistical analysis demonstrated that HRAS mutations were associated with chewing tobacco (p < 0.05). Consistent with the fact that Japanese people generally do not chew tobacco, the frequency of HRAS mutations among Japanese patients with OSCC was low. ConclusionEven though HRAS mutations are associated with OSCC, the HRAS gene may be a poor biomarker for investigating OSCC among the Japanese population due to the low frequency of the mutation. This phenomenon may be explained by a relatively low rate of chewing tobacco consumption in this population.

Evaluation of TGF-β1 and EGFR in Cleft Affected Lip Mucosa.

Background.The morphopathogenesis of orofacial cleft development is only partly understood; therefore, it is important to identify factors, which possibly could be involved in it. The aim of the study was to evaluate the distribution of TGF-β1 and EGFR-containing cells in cleft affected lip mucosa.Materials and Methods.The study group included lip mucosa tissue samples from 14 patients with orofacial cleft. The control group contained 11 healthy oral mucosa tissue samples. The tissue sections were stained by immunohistochemistry for TGF-β1 and EGFR. The expression of positive structures was graded semiquantitatively. IBM SPSS 26.0 was used for statistical analysis, Spearman`s rank correlation and Mann-Whitney U tests were performed.Results.Mostly few to moderate number (+/++) of TGF-β1-containing cells was found in epithelium, also the same number of fibroblasts and macrophages was seen in the lamina propria of cleft affected lip mucosa. Meanwhile, healthy oral mucosa on average demonstrated a moderate number (++) of TGF-β1-containing epithelial cells, fibroblasts, and macrophages. A variable, mostly indistinct number of EGFR-containing cells was seen in the epithelium of cleft affected lip mucosa, meanwhile, mostly no (0) EGFR positive cells were found in the epithelium of healthy mucosa. Statistically significantly less TGF-β1-containing cells were found in the epithelium of cleft affected lip mucosa than in the healthy mucosa (U=33.000; p=0.015). Also, the lamina propria of cleft affected lip mucosa showed statistically significantly less TGF-β1 immunoreactive fibroblasts and macrophages than the healthy mucosa (U=28.500; p=0.006).Conclusions.The decreased number of TGF-β1-containing epithelial cells, fibroblasts and macrophages in cleft affected lip mucosa proves the role of problematic tissue remodelation in the cleft pathogenesis. The distribution of EGFR in cleft affected and healthy mucosa is similar and possibly does not play a role in the cleft development of humans.

Cytokines Levels and Salivary Microbiome Play A Potential Role in Oral Lichen Planus Diagnosis

Oral lichen planus (OLP) is a chronic Th1-mediated inflammatory mucocutaneous disease of the skin and oral mucosa that can have various clinical presentations. Lesions are usually bilateral and often painful. While cutaneous Lichen Planus (LP) lesions are self-limiting, the oral lesions are chronic and rarely remissive. The diagnosis of oral lichen planus (OLP) is often challenging, and confirmation by histopathological criterion is generally advised. The aim of our study was to identify the cytokines present in OLP-suggestive lesions and in non-specific inflammatory lesions (NSIL) used as controls. Moreover, assess cytokines protein levels and oral microbiota composition in whole saliva samples. Histopathological analysis, immunohistochemistry and gene expression were used as techniques to analyze the oral mucosal tissue samples. ELISA was conducted to analyze salivary cytokine levels and 16S rRNA sequencing was used to determine the salivary microbiome. As a result we observed larger number of infiltrated lymphocytes (p = 0.025), as well, more T CD4 lymphocytes in the epithelial tissue (p = 0.006) in OLP samples compared to NSIL. In addition, the OLP samples displayed more apoptotic cells compared to NSIL (p = 0.047). Regarding the cytokine analysis, IFN-γ and IL-33 were more expressed in OLP lesions than in NSIL samples (p < 0.001; p = 0.026). Furthermore, our results demonstrated higher levels of IFN-γ protein expression in the saliva of OLP group compared to controls (p = 0.0156). We also observed noted differences in the oral microbiota composition between OLP and NSIL saliva samples. In conclusion, OLP lesions presented larger numbers of apoptotic and inflammatory cells, higher levels of IFN-γ and IL-33 compared to NSIL, and these lesions also differ regarding oral microbiota composition. These results are consistent with the Th-1-mediated chronic inflammation nature of oral lichen planus investigated lesions and displayed unique features that could be used as a diagnostic tool.

IL-25 promotes Th2-type reactions and correlates with disease severity in the pathogenesis of oral lichen planus

ObjectiveThe aim of the present study was to investigate the correlation between IL-25 expression and disease severity, and the potential immunoregulatory role of IL-25 expression in oral lichen planus (OLP). Materials and MethodsThe oral mucosal tissue samples obtained from OLP patients and healthy controls (HCs) were analyzed for IL-25 expression by real-time quantitative PCR (qPCR) and immunohistochemistry. Recombinant IL-25 was used to stimulate OLP patient-derived CD4 + T cells, and then IL-4 secretion and mRNA expression were evaluated by ELISA and qPCR, respectively. The efficiency of the siRNA-mediated knockdown of IL-25R expression in oral keratinocytes was determined by qPCR and Western blotting. Human oral keratinocyte cells were cultured with the recombinant human cytokines IL-25, IL-17 A and IL−17 F. The production of associated cytokines by keratinocytes was determined by qPCR. Statistical analyses of quantitative data were performed using SPSS software. ResultsThe IL-25 and IL-4 mRNA levels were elevated and correlated significantly with each other in specific OLP subtype lesions compared to HCs, while the numbers of IL-25 positive cells were also increased in local OLP lesions as compared to HCs. In vitro culture with recombinant IL-25 could significantly promote CD4 + T cells from both subtypes of OLP to produce IL-4 mRNA and remarkably elevate supernatant IL-4 levels in reticular OLP CD4 + T cell cultures, which may be attributed to the elevated expression of IL-25R in local OLP lesions. Statistical analyses demonstrated that the simultaneously increased levels of IL-4, CXCL8 and CCL20 in keratinocytes were induced by IL-25 but not IL-17 A or IL−17 F. Decreasing IL-25R subunit expression by siRNA-mediated knockdown significantly blocked the expression of all cytokine-produced inflammatory mediators in oral keratinocytes. ConclusionsIn OLP lesions, IL-25 can function to mediate the Th2 response in specific disease subtypes, which may be an important cause of OLP disease chronicity and persistent inflammation.

Transcriptome analysis of sheep oral mucosa response to Orf virus infection.

Contagious ecthyma is a highly contagious disease with worldwide distribution, which is caused by the Orf virus (ORFV) belonging to the Parapoxvirus. To study the alteration of host gene expression in response to ORFV infection at the transcriptional level, several young small-tailed Han sheep were inoculated with ORFV, and their oral mucosa tissue samples (T0, T3, T7 and T15) were collected on day 0, 3, 7 and 15 after ORFV infection respectively. RNA-seq transcriptome comparisons were performed, showing that 1928, 3219 and 2646 differentially expressed genes (DEGs) were identified among T3 vs. T0, T7 vs. T0, and T15 vs. T0 respectively. Gene Ontology (GO) analyses of the DEGs from these comparisons, revealed that ORFV might provoke vigorous immune response of the host cells during the early stage of infection. Moreover, GO and network analysis showed that positive and negative regulative mechanisms of apoptosis were integrated in the host cells through up or down-regulating the expression level of DEGs involved in apoptotic pathways, in order to reach a homeostasis of oral mucosa tissues during the exposure to ORFV infection. In conclusion, our study for the first time describes the direct effects of ORFV on the global host gene expression of its host using high-throughput RNA sequencing, which provides a resource for future characterizing the interaction mechanism between the mammalian host and ORFV.

Postmortem Histological Changes of Oral Mucosal Tissue in Hypoxic and Histotoxic Death to Differentiate Cause and Manner of Death

Abstract Cell is the basic structural and functional unit of all living organisms. In biological aspects humans though consider to be similar to other living organisms,they are unique as humans can think, imagine, and create so on.It is necessary to know the accurate cause of the death for solving many of the legal and administrative problems. In many cases cause of the death is determined by medical investigator. Aims & Objective: To study microscopic changes in oral mucosal tissue from cases of hypoxic and histotoxic death. Materials & Method: Fifteen oral mucosal tissue samples each form poisonous and hanging death were collected during autopsy. Tissues were routinely fixed, processed and stained using Haematoxylin and Eosin stain. Sections were studied and nuclear and cytoplasmic changes of epithelial cells were observed. Results: Definitive changes like prominent cytoplasmic vacuolation, in case of hanging and nuclear morphological changes in cases of poison death were observed. Conclusion: Biochemical mechanism behind cell death varies with cause of death, which in turn can be observed as morphological changes.

Expression of apoptosis-regulating miRNAs and target mRNAs in oral squamous cell carcinoma

Aberrations in the apoptotic mechanisms that cause excessive or deficient programmed cell death have been linked to a wide array of pathological conditions. In this study, using real-time reverse transcriptase-PCR, we analyzed the expression of apoptosis-regulating miRNAs (miR-15a, miR-16, miR-17-5p, miR-20a, miR-21, miR-29a, and miR-34a) in 20 oral squamous cell carcinoma and 5 normal oral mucosa tissue samples. Bioinformatic algorithms were used to identify the target genes of these miRNAs (BCL2, CASP2, CASP7, CASP8, DIABLO). The expression transcript levels of the target genes were measured in 50 oral squamous cell carcinoma and 10 normal oral mucosa tissue samples. We observed downregulation of miR-15a, miR-29a, and miR-34a in 50, 75, and 70% of samples, respectively. miR-16, miR-17-5p, miR-20a, and miR-21 expression was normal in 80, 75, 90, and 60% of samples, respectively. BCL2 transcripts were downregulated in 60% of samples, and normal-like expression was observed for CASP2, CASP7, CASP8, and DIABLO transcripts in 66, 82, 68, and 60% of samples, respectively. BCL2 expression was negative to weak, and that of proteins CASP2, CASP7, CASP8, and DIABLO was moderate tostrong. Our study provides evidence of alterations in the expression of apoptosis-regulating miRNAs and genes in the apoptotic pathway, demonstrating that regulation of apoptosis is a hallmark of oral squamous cell carcinoma pathogenesis.

Abstract 3160: Expression of apoptosis-regulating miRNAs and target mRNAs in oral squamous cell carcinoma

Abstract Apoptosis is a genetically programmed form of cell death, which primarily functions to eliminate senescent or altered cells that are useless or harmful for the multicellular organism. In contrast, aberrations of the apoptotic mechanisms that cause excessive or deficient programmed cell death have been linked to a wide array of pathologic conditions. MicroRNAs (miRNAs) are approximately 22 nucleotide non-coding RNA molecules that regulate gene expression post-transcriptionally and recent studies have indicated the dysregulation of miRNAs involved in apoptosis. In this study, using Real Time RT-PCR we have analyzed the expression of apoptosis-regulating miRNAs (miR-15a, miR-16, miR-29a, miR-34a, miR-17-5p, miR20a, miR21) in a series of 20 oral squamous cell carcinoma samples and a pool of 5 normal oral mucosa tissue samples. Using TargetScan algorithms, target genes of these miRNAs were identified and the expression of mRNA transcripts of these target genes was also evaluated (Bcl-2, Caspase-2, Caspase-7, Bid, SMAC/DIABLO) in a series of 50 oral squamous cell carcinoma samples and a pool of 10 normal oral mucosa tissue samples. Considering up- or down-regulation as a 2-fold difference in expression relative to the normal tissue, we observed down-regulation of miR-15a, miR29a and miR34a in 50%, 75% and 70% of the samples, respectively. Normal-like expression was observed for miR-16, miR-17-5p, miR-20a and miR-21 in 80%, 75%, 90% and 60% of the samples, respectively. Down-regulation of the BCL-2 transcripts was observed in 60% of the samples and normal-like expression was observed for BID, CASPASE-2, CASPASE-7 and SMAC/DIABLO transcripts in 66%, 82%, 68% and 60% of the samples, respectively. Expression of BID, CASPASE-2 and CASPASE-7 might be regulated by miR-17-5p and miR-20a and we found no alteration in expression of both mRNAs and miRNAs. Expression of BCL-2 might be regulated by miR-15a, miR-16, miR-21 and miR-34. We found down-regulation of BCL-2 transcripts and miRNAs-15a and -34, suggesting that other mechanisms might be involved in the regulation of BCL-2. Expression of SMAC/DIABLO might be regulated by miR-29 and we found normal-like expression of SMAC/DIABLO transcripts and down-regulation of miRNA-29 also suggesting that other mechanisms might be involved in the regulation of SMAC/DIABLO. Our study provides evidence of alterations in the expression of apoptosis-regulating miRNAs and alterations in the expression of genes involved in the apoptotic pathway demonstrating that regulation of apoptosis is an important feature in OSCC behavior. FAPESP research grants 98/14335-2 and 07/50608-4. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3160. doi:1538-7445.AM2012-3160

Human Calmodulin‐Like Protein (CLP) Expression in Oral Squamous Mucosa and in Malignant Transformation

The purpose of this study was to test whether calmodulin-like protein (CLP) is expressed in normal human oral mucosal cells and if downregulation of CLP occurs in malignant transformation. Oral mucosal tissue was taken from three individuals in a double-blind manner. The samples were cut, measured, and homogenized. Total RNA was extracted and reverse transcribed. Each cDNA sample was subjected to polymerase chain reaction (PCR). PCR fragments were purified, cloned, and sequenced to verify the presence of CLP. Three oral mucosal tissue samples with biopsy-confirmed squamous cell carcinoma were obtained. These samples demonstrated regions of normal epithelial cells as well as invasive squamous cell carcinoma. One normal breast epithelial sample was also obtained for positive control. Sections were stained with an affinity-purified CLP antibody and counterstained with a diluted hematoxylin. Two observers evaluated the specimens for expression of CLP. Staining patterns and intensity were noted in normal oral mucosa, comparing them to the normal breast epithelium sample. Staining patterns and intensity were then observed in squamous tumor cells, comparing them to the patterns of benign squamous mucosa. CLP coding sequences were positively identified from the normal oral mucosal tissue samples by reverse transcription and polymerase chain reaction (RT-PCR) with 100% identity to the published CLP sequence (accession #M58026). In the three oral mucosa tissue samples with known squamous cell carcinoma, expression of CLP was readily detected in areas of normal oral mucosa, while a notable downregulation of CLP expression occurred in areas of malignant transformation. The staining intensity was equivalent to the staining seen in the benign breast epithelium used as a control. In the areas of squamous cell carcinoma, a decrease in CLP immunoreactivity occurred. There was a sharp contrast in staining quality and clarity between benign and malignant tissue. In the majority of the carcinoma regions, a complete lack of immunoreactivity was noted. The RNA for human CLP is found in normal oral mucosa. CLP expression is seen in normal oral mucosa with a downregulation of CLP expression in malignant transformation.

Lymphatic Microvessel Density and the Expression of Lymphangiogenic Factors in Oral Squamous Cell Carcinoma

Objective: In this study, the expression of several lymphangiogenic growth factors was investigated to determine the mechanism underlying lymphangiogenesis in oral squamous cell carcinoma (OSCCa). Materials and Methods: An immunohistochemical study was performed to determine the lymphatic microvessel density (LMVD) and the expression of vascular endothelial growth factor (VEGF), platelet-derived growth factor (PDGF), basic fibroblast growth factor (bFGF), insulin-like growth factors (IGF-I/II), and hepatocyte growth factor (HGF) in 25 OSCCa and 20 normal oral mucosal tissue samples. Results: Significantly higher levels of expression of VEGF, PDGF, bFGF, HGF, IGF-I, and IGF-II existed in OSCCa tissue compared to normal oral mucosal tissue (p < 0.001, p < 0.001, p < 0.001, p < 0.001, p < 0.001, and p < 0.01, respectively; Mann-Whitney test). In OSCCa samples, significant positive correlations, based on the Spearman correlation test, existed between the LMVD and VEGF (r = 0.573; p < 0.01), PDGF (r = 0.559; p < 0.01), bFGF (r = 0.516; p < 0.01), HGF (r = 0.448; p < 0.05), and IGF-I (r = 0.6; p < 0.01), but no correlation existed between LMVD and IGF-II (r = 0.380; p > 0.05). Conclusion: These findings indicate that several growth factors may promote lymphangiogenesis in OSCCa. Future treatment for OSCCa utilizing antilymphangiogenic agents should target these different growth factor signaling pathways that are involved in lymphangiogenesis in OSCCa.

Significance of liquefaction degeneration in oral lichen planus: a study of its relationship with apoptosis and cell cycle arrest markers

To assess the utility of liquefaction degeneration as a marker of apoptosis in oral lichen planus (OLP). TdT-mediated dUTP-biotin nick-end labelling (TUNEL) assay and immunohistochemical methods were used to detect p21 proteins and the active form of caspase 3 in 32 tissue samples of oral mucosa with OLP and 20 samples of normal oral mucosa. Liquefaction degeneration was moderate or intense in 27.5% (n=8) and slight in 72.4% (n=21) of OLP samples. There was low expression of apoptosis markers (TUNEL, active caspase 3 form), which was not significantly associated with liquefaction degeneration of the basal cell layer. Basal and suprabasal expression of p21 was significantly more frequent in samples with more intense liquefaction degeneration of basal cells (P<0.01). Our results demonstrate that liquefaction degeneration, as a morphological expression of T lymphocyte attack, does not unequivocally indicate apoptosis. Attacked basal cells more frequently respond with cell-cycle arrest or senescence than with apoptosis.

In vivo experimental model of human gingival mucosa using immunodeficient mice

To establish an in vivo experimental model for examining human periodontal tissue, the present study examined several transplant techniques that maintain the structure and characteristics of human gingival mucosa. Human oral mucosal tissue samples were collected from the gingiva (n = 11), palate (n = 1), and tongue (n = 3). These mucosal grafts were transplanted onto BALB/c nu/scid mice with double-mutant immunodeficiency. Murine skin, twice the size of the graft, was cut open in an ' square superset'-shape. Next, the connective tissue side of the graft was placed onto the murine connective tissue. Immunohistochemical analysis was also performed, using polyclonal rabbit antibody to involucrin, monoclonal antibody to vimentin, monoclonal antibody to CD34, and monoclonal antibody to Ki-67, to determine whether the characteristics of human oral mucosa were maintained. When the connective tissue side of the graft was placed on the murine fascial membrane, the histological structure of the graft was maintained for 60 d. These grafts were examined for human characteristics using human-specific antibodies. Immunohistochemically, the expression patterns of involucrin, vimentin, and Ki-67 indicated that transplanted mucosa revealed normal human characteristics, including differentiation and proliferation up to 80 d. CD34 was not detected in the graft endothelial cells. The present study revealed that the novel technique of transplantation of human gingival mucosa in nu/scid mice may serve as an in vivo experimental model of periodontal disease.

Apoptosis and cell cycle arrest in oral lichen planus: Hypothesis on their possible influence on its malignant transformation

The quantitative importance of cell cycle arrest and apoptosis mechanisms in oral lichen planus (OLP) was analysed in order to assess the cell response to T lymphocyte aggression and establish a hypothesis on the influence of these phenomena in the malignant transformation process. The TUNEL assay and immunohistochemical methods were used to detect caspase-3, bax, and p21 in 32 tissue samples of oral mucosa with OLP and in 20 samples of normal oral mucosa. Positivity for TUNEL, caspase-3 and p21 was significantly more frequent in cases than in controls (p<0.001). Both TUNEL and caspase-3 positivity was significantly greater in the basal versus suprabasal layer (p=0.004 and 0.052, respectively). The basal and suprabasal expression of p21 was significantly higher in cases with a more intense liquefaction degeneration (p<0.01). There was no significant difference in basal expression of bax between cases and controls. The quantitative importance of apoptosis was small in OLP. Epithelial cells attacked in OLP have a very low response to apoptosis and cell cycle arrest mechanisms, which may produce an epithelial substrate that favours malignant transformation.

A genomic predictor of oral squamous cell carcinoma.

The objective was to identify a genomic profile that predicts the likelihood of oral squamous cell carcinoma compared with normal oral mucosa in unknown tissue samples. Using a training set of tissue samples that were histologically classified as oral squamous cell carcinoma or normal mucosa, the authors used principal component analysis to develop a genomic predictor for oral squamous cell carcinoma. On a separate test set of unclassified samples, the authors used the predictor to classify the samples, then evaluated the performance of the predictor using histological diagnosis. The authors used a data set consisting of messenger RNA extracted from 29 oral squamous cell carcinoma and 19 normal oral mucosa tissue samples and hybridized to Affymetrix oligonucleotide microarrays containing probe sets for 7070 genes and expressed sequence tags. The samples were divided into a training set of 15 oral squamous cell carcinoma and 10 normal samples and a test set consisting of the remaining samples. Using principal component analysis on the training set, the authors found a composite gene expression vector (principal component vector), which they used to compute likelihood ratios for oral squamous cell carcinoma on the test set. By calculating the contribution of each gene to the principal component vector, the authors identified genes with the greatest predictive value. Using the likelihood ratio, the authors correctly classified all 23 samples in the test set as either oral squamous cell carcinoma or normal. The authors found that many of the most predictive genes are known to be markers of squamous cell carcinoma or normal mucosa. Principal component analysis can be used with genomic microarray data to correctly predict the presence of oral squamous cell carcinoma in unknown tissue samples.

Studies of antigens in mucous membrane pemphigoid using laser scanning confocal microscopy

This study employed laser scanning confocal microscopy to characterize the distribution and colocalization of hemidesmosomal antigens in oral mucosa in normal individuals and patients with mucous membrane pemphigoid. Mucous membrane pemphigoid (MMP) is one of a group of conditions known as autoimmune vesiculobullous diseases. Individuals with these diseases develop autoantibodies to cell-cell or cell–basement membrane adhesion complexes that result in blister formation. Potentially severe sequelae can include limited opening of the oral cavity, blindness, and even death. Many studies in recent years have begun to elucidate that the antigens involved in MMP are located in hemidesmosomes, protein complexes that provide tight adhesion between basal cells and the underlying lamina propria. Although these experiments have greatly expanded the body of knowledge regarding the characterization of these proteins, little or no structural information is available regarding hemidesmosomal antigens and their interaction in the normal and diseased states. Laser scanning confocal microscopy (LSCM) is a powerful tool that allows precise visualization of multiple antigens simultaneously in tissue samples, as well as mathematical analysis for quantification and colocalization of these proteins. We obtained tissue samples of oral mucosa from normal individuals and patients with MMP. Using LSCM to analyze these tissue samples we demonstrated for the first time the localization and distribution of these antigens in the normal and diseased states. We also performed computer analysis of the image data, proving colocalization of specific antigens in the normal state and disruption of these associations in diseased tissue. These studies serve as an initial step in shedding light on the cryptogenic mechanism by which autoantibodies to hemidesmosomal antigens in MMP elicit blister formation.

Identification of Candida species in formalin fixed, paraffin wax embedded oral mucosa by sequencing of ribosomal DNA

Aim-To identify Candida species in formalin fixed, paraffin wax embedded tissue by sequencing candidal rDNA.Methods-Target rDNA sequences were amplified by polymerase chain reaction (PCR) from fresh isolates of Candida and from 18 preserved oral mucosal tissue samples (16 cases of chronic hyperplastic candidiasis and two fibroepithelial polyps), shown histologically to contain Candida. Identification of Candida species within tissue was based on a comparison of the rDNA sequences obtained with those from the fresh isolates of Candida and those present in the GenBank database.Results-The PCR products obtained from 12 of the 18 tissue specimens studied were characteristic of Candida albicans. In two of these cases a second, larger PCR product was obtained and these sequences were characteristic of Candida glabrata.Conclusions-Candidal DNA was amplified successfully from formalin fixed, paraffin wax embedded tissue. Sequencing of the PCR product enabled identification of the Candida species present.