Dysplastic Leukoplakia Research Articles

Cancer Stem Cell Marker CD147 Expression in Erosive Oral Lichen Planus Compared to Moderately and Severely Dysplastic Leukoplakia

Oral lichen planus is a frequent, chronic autoimmune disease that affects the oral mucosa and is characterized as an oral potentially malignant disorder. The aim of our study is to examine the presence of CSCs bearing CD147 (a marker related to local inflammation and associated with various cancers) through immunohistochemistry in oral lichen planus (OLP) compared to oral leukoplakia (OL) and healthy tissues. These findings could contribute to clinical practice by providing a marker for the prognostic assessment of OLP lesions with regards to their potentially malignant nature. The study sample consisted of paraffin-embedded oral mucosa specimens from the archives of the Department of Oral Medicine/Pathology, School of Dentistry, Aristotle University of Thessaloniki, Greece during the period 2009–2019. The study sample contained 24 cases of OLP (14 erosive and 10 reticular) and 30 cases of oral leukoplakia, which were compared to 5 normal oral epithelium samples derived from healthy epithelium adjacent to fibromas from other cases. Cell membrane staining of CD147 was observed mostly in the basal and parabasal cell layer. The statistically significantly higher expression of CD147 in the erosive lichen planus subgroup than in the moderately and severely dysplastic leukoplakia subgroup (p = 0.01) constituted the most important finding of this study. The characteristic expression of CD147 in erosive OLP suggests the presence of epithelial cells with CSC characteristics, but its lower expression in oral leukoplakias suggests a more intense relation of the CD147 marker with inflammation rather than with oral dysplastic progression.

Detection of oral cancer and oral potentially malignant disorders using artificial intelligence-based image analysis.

We aimed to construct an artificial intelligence-based model for detecting oral cancer and dysplastic leukoplakia using oral cavity images captured with a single-lens reflex camera. We used 1043 images of lesions from 424 patients with oral squamous cell carcinoma (OSCC), leukoplakia, and other oral mucosal diseases. An object detection model was constructed using a Single Shot Multibox Detector to detect oral diseases and their locations using images. The model was trained using 523 images of oral cancer, and its performance was evaluated using images of oral cancer (n = 66), leukoplakia (n = 49), and other oral diseases (n = 405). For the detection of only OSCC versus OSCC and leukoplakia, the model demonstrated a sensitivity of 93.9% versus 83.7%, a negative predictive value of 98.8% versus 94.5%, and a specificity of 81.2% versus 81.2%. Our proposed model is a potential diagnostic tool for oral diseases.

Malignant Transformation and Long-Term Outcome of Oral and Laryngeal Leukoplakia.

Oral or laryngeal leukoplakia has an increased risk for malignant transformation but the risk of the two anatomical sites has not been compared to each other yet. Clinical data of 253 patients with leukoplakia (oral = 221 or laryngeal = 32) enrolled from January 1996 to January 2022 were analyzed. One hundred and seventy underwent biopsy and 83 did not. The mean follow-up time was 148.8 months. Risk factors for the malignant transformation of leukoplakia were identified using Cox proportional hazard models. In the oral or laryngeal group, the rate of cancer was 21.7% and 50% (p = 0.002), respectively. The 10-year estimated malignant transformation was 15.1% and 42% (p < 0.0001), respectively. The laryngeal group had an increased risk of malignant transformation (p < 0.0001). The 5-year estimated survival with leukoplakia-associated cancer for the oral or laryngeal group was 40.9% and 61.1% (p = 0.337), respectively. Independent predictors of malignant transformation in the oral group were dysplasia and the grade of dysplasia of the leukoplakia, and in the laryngeal group, dysplasia had a significant impact. The malignant transformation rate was low for oral patients without biopsy or with no dysplasia, 3.9% and 5.1%, respectively. The malignant transformation occurred over 10 years. Patients with dysplastic leukoplakia have an increased risk of malignant transformation, but the risk is higher with laryngeal than with oral leukoplakia. There is no significant difference between the groups regarding survival with leukoplakia-associated cancer. Oral patients with no dysplastic lesions have a low risk of malignant transformation. A complete excision and long-term follow up are suggested for high-risk patients to diagnose cancer in an early stage and to control late (over 10 years) malignant events.

Effect of clinical and histologic features on time to malignancy in 224 cases of oral leukoplakia treated by surgery

ObjectivesOur main purpose and research question were to analyze and quantify whether there were significant differences in the time to develop cancer among patients with oral leukoplakia (OL), comparing the more susceptible cases to those with the least susceptibility to malignancy.Materials and methodsWe followed 224 cases of OL after surgical or CO2 laser treatment for a mean time of 6.4 years. A Bayesian mixture cure model based on the Weibull distribution was used to model the relationship between our variables and cancer risk. In this model type, the population is considered a mixture of individuals who are susceptible or non-susceptible to developing cancer. The statistical model estimates the probability of cure (incidence model) and then infers the time to malignancy. The model was adjusted using the R-package INLA using default priors.ResultsHistology type (moderate or severe dysplasia) and tongue location showed hazard ratios (HR) of 3.19 (95% CI [1.05–8.59]) and 4.78 (95% CI [1.6–16.61]), respectively. Both variables increased the risk of malignant transformation, thus identifying a susceptible subpopulation with reduced time required to develop cancer, as with non-homogeneous leukoplakias. The median time for cancer development was 4 years and 5 months, with a minimum of 9 months after the diagnosis of OL and a maximum of 15 years and 2 months.ConclusionsSusceptible patients with non-homogeneous leukoplakia, dysplasia, or leukoplakia in the tongue develop cancer earlier than those with homogeneous OL and those without dysplasia.Clinical relevanceThe novel contribution of this research is that, until now, the time it took for oral leukoplakias to develop cancer based on whether they were homogeneous or non-homogeneous, and if they have or not epithelial dysplasia, had not been comparatively described and quantified. As a final result, the time to malignant transformation in non-homogeneous and dysplastic leukoplakias is significantly shorter.

Oral leukoplakia: A clinicopathological study and malignant transformation.

To assess the clinicopathological characteristics of a cohort of patients with oral leukoplakia (OL) managed in a Romanian dental hospital and to evaluate the risk of malignant transformation (MT). We conducted a hospital-based retrospective study using the clinical charts of patients diagnosed with OL that had complete clinical and histopathological evaluation followed up for 1-16years. From 120 included patients, 68 (56.7%) were females, and 71 (59.2%) were current smokers. The homogeneous form was present in 60% of cases; the buccal mucosa was the most frequently involved site. MT was observed in 9 cases, which was more common in females and in those with dysplastic leukoplakia. A significant statistical association was found between MT and dysplasia grade (χ2 test: p=0.007). MT occurred during a mean interval of time 75months in both treated and non-treated patients. In this leukoplakia cohort, most of the lesions encountered were in smokers, clinically homogeneous and 62.5% proved histologically benign. But despite the clinically benign appearance of leukoplakia, tissue diagnosis of some cases was carcinoma. The results of the current study advocate the necessity for biopsy even in apparently homogeneous, clinically benign lesions. The malignization rate was 7.5%; two-thirds were nonhomogeneous lesions.

Matrix Metalloproteinases (MMPs) in skin and oral pathology - Premalignant Diseases, Spinocellular Carcinoma and Basocellular Carcinoma (markers).

Matrix Metalloproteinases (MMPs) in skin and oral pathology - Premalignant Diseases, Spinocellular Carcinoma and Basocellular Carcinoma (markers).

SIRT-1 and CD44 in the saliva of oral squamous cell carcinoma, dysplastic and non-dysplastic oral leukoplakia

Objective: The objective of this study was to investigate the levels of salivary SIRT-1 and CD44 in patients suffering from oral squamous cell carcinoma (OSCC), dysplastic and non-dysplastic oral leukoplakia to assess their potential role as early diagnostic biomarkers for patients with oral leukoplakia and OSCC. Subjects and Methods: sixty patients were recruited from the outpatient clinic of the Department of Oral Medicine and Periodontology, Faculty of Dentistry, Cairo University. Participants were divided into four groups:15 healthy individuals, 15 patients with non-dysplastic leukoplakia, 15 patients with dysplastic leukoplakia, and 15 patients with oral squamous cell carcinoma. Whole unstimulated saliva (WUS) was collected from all participants. Soluble CD44 (solCD44) and SIRT1 were measured using ELISA. Results: CD44 was expressed in all the four study groups with different levels. OSCC showed a statistically significant highest CD44 level. While the control group showed the lowest CD44 level. There was no statistically significant difference between dysplastic leukoplakia and non-dysplastic leukoplakia groups. Similarly, there was no statistically significant difference between non-dysplastic leukoplakia and control groups. OSCC showed the statistically significantly highest SIRT-1 level. There was no statistically significant difference between dysplastic leukoplakia, non-dysplastic leukoplakia, and control groups; all showed statistically significantly lower SIRT-1 levels than the OSCC group. Conclusion: salivary SIRT-1 and CD44 could be used as a screening tool to improve early detection and diagnostic precision of oral leukoplakia and OSCC.

Clinicopathologic factors associated with malignant transformation of oral leukoplakias: a retrospective cohort study

It is clinically challenging to identify oral leukoplakias that have a high risk of undergoing malignant transformation. The aim of this retrospective study was to elucidate the associations between malignant transformation of oral leukoplakias and various clinicopathologic factors. Patients with a diagnosis of clinical oral leukoplakia, verified through histopathologic examination and with access to digital images of the lesion, were retrospectively included for the period 2003–2013. Using the clinical images, all lesions were re-evaluated regarding diagnosis and clinical subtype. Of the 234 included patients, with a median follow-up of 9 years, 27 (11.5%) developed oral squamous cell carcinoma. Among the clinicopathologic factors investigated, non-homogeneous oral leukoplakia (OL), OL with dysplasia, and OL localized to the tongue showed statistically significant increased rates of malignant transformation in the multivariate Cox regression analysis. Non-homogeneous OL showed a 15.2-times higher transformation rate than homogenous OL (P<0.001). Dysplastic leukoplakias developed into carcinomas 2.4-times more often than did non-dysplastic leukoplakias (P=0.048). OL located on the tongue showed a 2.8-times higher malignant transformation rate than OLs at other oral locations (P=0.018), when other locations were combined into one group. Non-homogeneous OL, OL with dysplasia, and OL localized to the tongue have higher transformation rates.

Evaluation of CD44 and TGF-B Expression in Oral Carcinogenesis.

Statement of the Problem: Oral squamous cell carcinoma (OSCC) is the most common malignancy of the oral cavity. Early diagnosis of OSCC by using biomarkers provides preventive treatment approach to suppress the disease in early stages. CD44 as a cancer stem cell (CSC) marker may be cleaved by MT1-MMP and plays an important role in migration of cancer cells. TGF-B promotes formation of invasive cancer cells phenotype through epithelial mesenchymal transition (EMT) and induces MT1-MMP formation. Purpose: The aim of this study was to evaluate the expression of TGF-B and CD44 in leukoplakia (premalignant lesion), squamous cell carcinoma (SCC), and normal oral mucosa to determine the role of these markers in the carcinogenesis process of the oral mucosa.Materials and Method: In this retrospective study, the expression of TGF-B and CD44 were evaluated in 55 paraffin-embedded specimens (10normal mucosa, 15 non-dysplastic leukoplakia, 15 dysplastic leukoplakia, and 15 OSCC) by immunohistochemistry. Statistical analyses were performed using Kruskal-Wallis, Mann-Whitney, and Spearman’s rank correlation tests.Results: Evaluation of CD44 and TGF-B expression in the four studied groups showed statistical significant difference for each marker (p< 0.001). Pairwise comparison of CD44 and TGF-B expression in all groups except normal mucosa and non-dysplastic leukoplakia demonstrated statistical significant difference. In addition, there was positive significant correlation between two markers (r= 0.914, p< 0.001). Diagnostic test’s accuracy for identification of OSCC and dysplastic leukoplakia from non-dysplastic leukoplakia and normal tissues and recognition of OSCC from dysplastic leukoplakia showed optimum sensitivity and specificity.Conclusion: Increased expression of CD44 as a cancer stem cell marker and TGF-B as an EMT marker from normal mucosa to non-dysplastic leukoplakia, dysplastic leukoplakia, and OSCC and also the significant correlation between these two markers indicated their role in carcinogenesis of oral mucosa.

BMI-1 expression increases in oral leukoplakias and correlates with cell proliferation.

Oral leukoplakia (OL) is a white lesion of an indeterminate risk not related to any excluded (other) known diseases or disorders that carry no increased risk for cancer. Many biological markers have been used in an attempt to predict malignant transformation; however, no reliable markers have been established so far. Objective To evaluate cell proliferation and immortalization in OL, comparing non-dysplastic (Non-dys OL) and dysplastic OL (Dys OL).Methodology This is a cross-sectional observational study. Paraffin-embedded tissue blocks of 28 specimens of Non-dys OL, 33 of Dys OL, 9 of normal oral mucosa (NOM), 17 of inflammatory hyperplasia (IH), and 19 of oral squamous cell carcinomas (OSCC) were stained for Ki-67 and BMI-1 using immunohistochemistry.Results A gradual increase in BMI-1 and K-i67 expression was found in oral carcinogenesis. The immunolabeling for those markers was higher in OSCC when compared with the other groups (Kruskal-Wallis, p<0.05). Ki-67 expression percentage was higher in OL and in IH when compared with NOM (Kruskal-Wallis/Dunn, p<0.05). Increased expression of BMI-1 was also observed in OL when compared with NOM (Kruskal-Wallis/Dunn, p<0.05). No differences were observed in expression of both markers when non-dysplastic and dysplastic leukoplakias were compared. A significant positive correlation between Ki-67 and BMI-1 was found (Spearman correlation coefficient, R=0.26, p=0.01). High-grade epithelial dysplasia was associated with malignant transformation (Chi-squared, p=0.03).Conclusions These findings indicate that BMI-1 expression increases in early oral carcinogenesis and is possibly associated with the occurrence of dysplastic changes. Furthermore, our findings indicate that both Ki-67 and BMI-1 are directly correlated and play a role in initiation and progression of OSCC.

Salivary proteins from dysplastic leukoplakia and oral squamous cell carcinoma and their potential for early detection

Dysplastic leukoplakia (LP) of the oral cavity is a potentially malignant condition for oral squamous cell carcinoma (OSCC), early detection of which remains an unmet clinical need. In an effort to develop non-invasive biomarker based method for early detection of the disease, differential proteomic profiling was carried out with the saliva from patients with risk habits and diagnosed with LP and those with lymph node negative and positive OSCC in comparison to healthy controls with risk habits. Ninety three proteins were observed at elevated level (≥1.5 fold), and 30 were prioritized based on a scoring system comprising of confidence of identification, presence in the various specimen groups, functional relevance, and their secretory potential. Verification was carried out in independent patient cohorts for 8 selected, representative, upregulated proteins using ELISA. Three of them CD44, S100A7, and S100P were significantly altered in patients with LP as well as OSCC and can be regarded as a panel of biomarker candidates for early detection of the malignancy. Other members may also be investigated in a targeted manner to expand the portfolio of biomarkers for early detection. The mass spectrometry data are available via ProteomeXchange with identifier PXD015722. SignificanceThere is an unmet clinical need for non-invasive, biomarker based methods for the improved early detection and the subsequent management of oral cancer. The study represents differential proteome profiling of the saliva of patients with oral dysplastic leukoplakia (LP) – a potentially malignant lesion, patients diagnosed with oral squamous cell carcinoma (OSCC), and healthy controls to identify potential markers for the purpose of early detection of malignancy. From among the matched and prioritized proteins with elevated levels in the saliva of patients with LP and those with OSCC, eight were verified. Three of them – CD44, S100A7 and S100P appeared promising candidates as biomarkers for early detection of the neoplastic predisposition and may form the basis of clinical assays for this purpose.

Dysplastic oral leukoplakia is molecularly distinct from leukoplakia without dysplasia.

The molecular mechanisms underlying the development of dysplasia in leukoplakia are unknown. We used RNA sequencing to examine the molecular and biological pathway differences in oral leukoplakia with and without oral epithelial dysplasia. Excisional biopsy specimens (25) were taken from 24 patients with oral leukoplakia diagnosed histopathologically as either oral epithelial dysplasia (13) or epithelial hyperplasia and keratosis without dysplasia (12). Transcriptome analysis used RNA sequencing, differential expression and hierarchical clustering. Biological signalling was examined by gene ontology, pathway and protein-protein interaction analysis. Differential expression analysis showed distinction between the two groups identifying 47 genes as altered in leukoplakia with dysplasia, including SAA1, SAA2, KRT31, KRT37, KRT76, ROBO2, DNAJB5 and DNAJA4. Using hierarchical clustering, dysplastic leukoplakia readily segregated from leukoplakia without dysplasia. Pathway and ontology enrichment analysis provided evidence that downregulation of extracellular matrix (ECM) pathways was a feature of dysplastic lesions. Our results suggest that there are detectable changes in the molecular profile of oral leukoplakia exhibiting dysplasia including downregulated ECM as a distinguishing feature of dysplastic lesions. This suggests that reactive changes in stroma may be an early manifestation of dysplastic development. Our study also demonstrates the feasibility of detecting such molecular changes in oral leukoplakia, providing avenues for further investigation of molecular mechanisms of oral dysplasia.

Efficacy of Rose Bengal in comparison with Toluidine Blue in detection of potentially malignant lesions: a preliminary study

Introduction: Oral cancer is usually diagnosed when it becomes symptomatic and approximately two thirds of the patients present with the advanced disease, regional metastasis and consequently poor prognosis. The risk of development of carcinoma within an area of premalignant lesion is 5 times higher than in those without lesion. Malignant transformation has been reported in 43% of dysplastic leukoplakia cases. It is therefore important to identify these lesions early, for early management.&#x0D; Aim: To compare the diagnostic efficiency of Rose Bengal (RB) with Toluidine blue (TB) in&#x0D; detecting the biopsy sites and thus establish an accurate diagnosis in detecting oral potentially malignant disorders. &#x0D; Materials And Methods: In our study 27 patients with 41 lesions were included. Since one patient had not quit the habit in the two weeks following initial examination and another patient lesion disappeared in the waiting period, 2 patients (3 lesions) were not included in the study. After initial examination they were subjected to Rose Bengal and Toluidine blue stain. Positively stained lesions were subjected to biopsy and assessed for dysplasia. All the data were tabulated and statistically analysed in SPSS software version v.22.IBM, corp. Chi square test was done to compare the difference between two stains.&#x0D; Results: When compared with TB staining RB showed 100% Sensitivity, 90% specificity, 96.6% positive predictive value and 100% negative predictive value.&#x0D; Conclusion: Based on the results and other observations made during the study it could be suggested that RB could be used as an alternative to TB clinically. Further studies with large samples are recommended to support the evidence of this study.

The evaluation of the chemopreventive effect of β-carotene (A-Viton®) on oral premalignant lesions through the estimation of the expression of p53 levels

Background: Oral premalignant lesions (OPL) include a wide range of lesions including leukoplakia, dysplastic leukoplakia, erythroplakia, oral submucous fibrosis, dysplastic lichenoid lesions, and oral lichen planus. Proper diagnosis and management of oral premalignant lesions (OPL) are very important; as 17% of these lesions have been reported to be transformed into malignant lesions within first 7 years of their diagnosis. Vitamin A has always been known for its chemopreventive effect in several diseases through the modulation of cell proliferation and, it is also known for being a potent antioxidant and anti-inflammatory agent. Any alterations in p53 protein results in its accumulation in the cell nuclei; may play a critical role in carcinogenesis, including oral premalignant lesions and oral malignant lesions, i.e., p53 protein is one of the determinants in the progression of oral dysplasia to invasive malignancy.Methods: Fifteen patients having oral premalignant lesions of mild and moderate dysplasia; diagnosed both clinically and histologically, were given a chemopreventive therapy in the form of βeta- carotene (Vit. A) for four months. The levels of p53 protein immunostaining were measured pre- and post-treatment.Results: The mean value of optical staining density of P53 in pre-treatment specimen was (59.78 ± 10.22) compared to (39.74 ± 3.36) in post-treatment cases; this value was higher in pre-treatment cases than post-treatment cases. The p-value was 0.0003.Thus when statistically compared the difference is considered statistically significant.Conclusion: The expression of P53 protein in (OPL) is found to be inversely related to the clinical as well as histopathological response to beta-carotene supplementation.

Autophagy analysis in oral carcinogenesis

ObjectiveThe aim of this study was to evaluate the levels of autophagy in oral leukoplakia and squamous cell carcinoma and to correlate with clinical pathological features, as well as, the evolution of these lesions. Methodology7 Normal oral mucosa, 51 oral leukoplakias, and 120 oral squamous cell carcinomas (OSCC) were included in the study. Histological sections of the mucosa and leukoplakias were evaluated throughout their length, while the carcinomas were evaluated using Tissue Microarray. After the immunohistochemical technique, LC3-II positive cells were quantified in the different epithelial layers of the mucosa and leukoplakias and in the microarrays of the squamous cell carcinomas. The correlation between positive cells with the different clinical-pathological variables and with the evolution of the lesions was tested using the t test, ANOVA, and Kaplan-Meier survival analysis. ResultsWe observed increased levels of autophagy in the oral squamous cell carcinomas (p<0.001) in relation to the other groups, but without any association with poorer evolution or survival of these patients. Among the leukoplakias, we observed a higher percentage of positive cells in the intermediate layer of the dysplastic leukoplakias (p=0.0319) and in the basal layer of lesions with poorer evolution (p=0.0133). ConclusionThe levels of autophagy increased during the process of oral carcinogenesis and are correlated with poorer behavior of the leukoplakias.

Efficacy of argyrophilic nucleolar organizing region analysis using computer-assisted and manual in oral leukoplakia: A comparative study.

Nucleolar organizer regions are loops of DNA containing ribosomal RNA genes and presumably are associated with ribosomal RNA activity, protein synthesis, and cell proliferation. Argyrophilic nucleolar organizer region (AgNOR) count has been suggested as an objective method in differentiating dysplastic lesions from non-dysplastic lesions. This descriptive study was done on archival paraffin blocks (n = 60), consisting of 10 normal human oral epithelium, 22 cases of non-dysplastic leukoplakia (NDLK), and 28 cases of dysplastic leukoplakia (DLK). The AgNORs were counted with the aid of a manual using conventional light microscopy and photographs of the same were taken and analyzed using Image Pro Express 6.0 (Media Cybernetic Inc., USA) for windows. The mean AgNOR count per nucleus was found to be higher in patients with DLK as compared to NDLK and controls using both manual counting and image analysis method and on comparing both the techniques, image analysis provide a more accurate reflection of AgNOR counts than manual counting. To conclude, reliability of computerized image technique of AgNOR count is the most appropriate marker to differentiate between dysplastic and NDLK. Computer-assisted image analysis system was found to be an effective tool in achieving high reproducibility as compare to manual.

Abstract 430: Identification of novel methylation-silenced genes in oral squamous cell carcinomas

Abstract To identify novel methylation-silenced genes in oral squamous cell carcinomas (OSCCs), we took a combined approach of methylated DNA immunoprecipitation (MeDIP)-CpG island (CGI) microarray analysis and expression microarray analysis after treatment with a demethylating agent. MeDIP-CGI microarray revealed that promoter CGIs of 324 genes were methylated in two OSCC cell lines (Ca9-22, HSC-2). By treatment of the cell lines with 5-aza-2′-deoxycytidine, 125 of the 324 genes were up-regulated in at least one of the two cell lines. For 23 of the 125 genes that have not been focused on in the field of oral malignancy, we analyzed their promoter methylation statuses in 16 normal oral tissues, five OSCC cell lines, 26 OSCC tissues and 24 oral leukoplakia tissues. Ten promoter CGIs were methylated not only in OSCC cell lines but also OSCC tissues. The methylation profile indicated the presence of the CpG island methylator phenotype (CIMP) in OSCC tissues. Furthermore, the CIMP was observed in three of the 13 oral dysplastic leukoplakia tissues. The CIMP was considered as one of early molecular events in malignant transformation from oral premalignant lesions to OSCCs. Citation Format: Masanobu Abe, Satoshi Yamashita, Yoshiyuki Mori, Takahiro Abe, Hideto Saijo, Toshikazu Ushijima, Tsuyoshi Takato. Identification of novel methylation-silenced genes in oral squamous cell carcinomas. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 430. doi:10.1158/1538-7445.AM2014-430

Detection and typing of Human Papillomaviruses (HPV) in malignant, dysplastic, nondysplastic and normal oral epithelium by nested Polymerase Chain Reaction, immunohistochemistry and transitional electron microscopy in patients of Northern Greece

To evaluate the role of HPV in oral carcinogenesis, we examined the prevalence of HPV in malignant, potentially malignant and normal oral epithelium and studied the relation of HPV prevalence with other factors obtained from the patient's records. Our material consisted of 291 tissue specimens from 258 individuals. From every individual formalin fixed and paraffin embedded tissues were examined by nested Polymerase Chain Reaction (NPCR) for the detection of HPV DNA and by immunohistochemistry (IHC) for the in situ detection of HPV L1 protein. Positive PCR products were sequenced in order to type HPVs. Also 33 fresh tissues were obtained, fixed and used to detect HPV particles by transitional electron microscopy (TEM). HPV was detected in 32.9% of the tissue specimens by NPCR, in 4.7% by immunohistochemistry and in 28.1% by TEM. In detail, by nested PCR HPV L1 DNA was detected in 40% of normal tissues, 40% of fibromas, 35.8% of non-dysplastic leukoplakias, 31.6% of dysplastic leukoplakias and 22.2% of oral squamous cell carcinomas. The HPV viral load of 96.5% of the samples was very low (1 viral copy per 10(2)-10(4) cells). HPV16 prevails in all histological groups in 89-100%. We conclude that HPV does not seem, from the specific sample examined, to play a substantial role in oral carcinogenesis. However, it cannot be excluded that HPV could be involved in oral carcinogenesis only in cases with high viral load or at early stages of carcinogenesis possibly through the hit-and-run mechanism.

Detection of Candida albicans ADH1 and ADH2 mRNAs in human archival oral biopsy samples.

The aim of this study was to investigate the relationship between expression of Candida albicans alcohol dehydrogenases (ADH) genes in archival formalin-fixed paraffin-embedded (FFPE) samples from biopsies of leukoplakia. Archival FFPE samples were obtained from four sample groups: normal oral mucosa, non-dysplastic leukoplakia, chronic hyperplastic candidosis (CHC), and non-CHC dysplastic leukoplakia. The presence of C. albicans was determined by periodic acid Schiff staining and by immunocytochemistry. C. albicans ADH1 and ADH2 mRNAs were detected using reverse transcription PCR. Candida albicans was detected in FFPE samples diagnosed as CHC (the histological diagnoses had been made by specialist oral pathologists, using uniform criteria), but not in any other sample group, including the non-dysplastic leukoplakias. RT-PCR confirmed a significant correlation between the expression of CaADH1 mRNA (P = 0.000), but not for CaADH2 mRNA (P = 0.056) in archival FFPE samples (n = 31) from biopsies of leukoplakia. Candida albicans was the predominant species in the lesions diagnosed as CHC, and the presence of C. albicans in CHC lesions was associated with a high expression of C. albicans ADH1 mRNA. There was no association between the presence of Candida and malignant transformation in the cases examined; however, the number of cases was limited and further studies are needed to further elucidate the role of C. albicans ADH1 in the pathogenesis of oral squamous cell carcinoma.

Evaluation of the efficacy of AgNOR as a proliferative marker in oral leukoplakia: A morphometric analysis

Background:Silver stainable nucleolar organizer regions (AgNORs) are replicatory markers which may have a place in objectively characterizing dysplasia.Materials and Methods:A study of various morphometric parameters related to AgNORs was performed in basal and parabasal layers of normal human oral epithelium, nondysplastic leukoplakia, and dysplastic leukoplakia employing photomicrographs of silver stained paraffin embedded sections using image analysis, to assess the usefulness of these parameters in distinguishing dysplastic leukoplakia from nondysplastic oral leukoplakia.Results:Out of various mean AgNOR related parameters, AgNOR count, area, perimeter, and proportion were found to be higher in dysplastic leukoplakia as compared to nondysplastic leukoplakia. On statistical analysis, AgNOR count showed statistically significant differentiation between dysplastic and nondysplastic leukoplakia. While other parameters can distinguish normal oral epithelium from dysplastic and nondysplastic leukoplakia.Conclusion:To conclude, the AgNOR count is the most appropriate marker to differentiate between dysplastic and nondysplastic leukoplakia.