Análisis de microdensidad vascular y factores de crecimiento en carcinoma oral de células escamosas / Analysis of vascular microdensity and growth factors in oral squamous cell carcinoma

Introducción: El carcinoma oral de células escamosas (COCE) es una neoplasia epitelial maligna que se presenta frecuentemente entre la quinta y sexta década de la vida. Su compleja patogénesis incluye el proceso de angiogénesis y la regulación del microambiente tumoral como mecanismos de progresión tumoral. Objetivo: Determinar la relación entre las variables clínicas e histológicas del COCE con la inmunoexpresión de VEGF, FGF-1, FGFR-1, TGFB-1, TGFBR-II y CD105. Material y métodos: Nueve casos de COCE; tres bien (BD), tres moderado (MD) y tres pobremente diferenciados (PD) obtenidos del Departamento de Patología y Medicina Bucal, División de Estudios de Postgrado e Investigación. Se aplicó la técnica de inmunohistoquímica por peroxidasa para identificar la expresión de VEGF, FGF-1, FGFR- 1, TGFB-1, TGFBR-II y CD105. El análisis de inmunoexpresión se realizó con el programa ImageJ. Se aplicó la prueba de Kruskal-Wallis y correlación de Spearman (p < 0.05). Resultados: La inmunoexpresión de VEGF fue mayor en los COCE PD, FGFR-1 fue positivo en los BD, mientras que FGF, TGFB-1 y TGFBR-II fueron negativos. El análisis de microdensidad vascular (MVD) indicó mayor número de vasos CD105 positivos en los carcinomas BD, seguidos de los PD y MD. Conclusión: Considerando los resultados obtenidos podemos concluir que la angiogénesis es un fenómeno constante independiente del grado de diferenciación que durante el proceso de transformación de una neoplasia requerirá la formación de vasos sanguíneos y que este proceso puede ser modulado por factores de crecimiento tales como los analizados en este trabajo (AU)

Introduction: Oral squamous cell carcinoma (OSCC) is a malignant epithelial neoplasm that frequently occurs between the fifth and sixth decade of life. Its complex pathogenesis includes the angiogenesis process and the regulation of the tumor microenvironment as mechanisms of tumor progression. Objective: To determine the relationship between the clinical and histological variables of OSCC with the immunoexpression of VEGF, FGF-1, FGFR-1, TGFB- 1, TGFBR-II and CD105. Material and methods: Nine cases of OSCC; three well (WD), three moderate (MD) and three poorly differentiated (PD) obtained from the Oral Medicine and Pathology Department, Division of Graduate Studies and Research. The peroxidase immunohistochemistry technique was performed to identify the expression of VEGF, FGF-1, FGFR-1, TGFB-1, TGFBR-II and CD105. The immunoexpression analysis was performed with the ImageJ software. The Kruskal-Wallis and Spearman correlation test were performed (p < 0.05). Results: VEGF immunoexpression was higher in PD OSCC, while FGFR-1 was predominantly positive in WD; FGF, TGFB-1 and TGFBR-II were negative. Vascular microdensity analysis (MVD) indicated a greater number of CD105 positive vessels in WD carcinomas, followed by PD and MD. Conclusion: Considering the results obtained, we can conclude that angiogenesis is a constant phenomenon independent of the degree of differentiation; that during the transformation process of a neoplasm it will require the formation of blood vessels and that this process can be modulated by growth factors such as those analyzed in this work (AU)

High-risk mucosal HPV types associated with squamous cell carcinoma on the nose tip in an immunocompetent young man

Abstract: The oncogenic role of high-risk HPV in anogenital, head and neck, and cervical cancer is well recognized, but not in skin cancer in the general population. Some authors have demonstrated their appearance mainly on the hands and feet, particularly in the area of the nail bed, which could be due to contamination with HPV types from anogenital regions. Here, we describe a case of genital HPV associated with SCC on the nose tip in an immunocompetent young man, which was confirmed by histopathological findings and in situ hybridization. The importance of this report is to highlight the potential role of HPV in the etiology of skin cancer in an immunocompetent individual.

Inmunoexpresión de E-cadherina y vimentina en mucosa oral normal, displasia epitelial oral y carcinoma oral de células escamosas / Immunoexpression of E-cadherin and vimentin in normal oral mucosa, oral epithelial displasia and oral squamous cell carcinoma

El objetivo fue evaluar la inmunoexpresión de E-cadherina y Vimentina en mucosa oral normal (MON), displasia epitelial oral (DEO) y carcinoma oral de células escamosas (COCE). Se realizó un estudio descriptivo de una serie de casos analizandolos mediante técnica de inmunohistoquímica contra E-cadherina y Vimentina 16 muestras de MON, 16 de DEO y 19 de COCE. La inmunotinción fue evaluada cualitativamente considerando extensión e intensidad para E-cadherina e intensidad para Vimentina. El análisis de la extensión e intensidad de la inmunotinción de E-cadherina y Vimentina según diagnóstico reveló una asociación estadísticamente significativa (p<0,001). Siendo la expresión de E-cadherina más alta en MON, seguido por DEO y más baja en COCE, inversamente a lo que se observó con Vimentina. El presente estudio reveló la subregulación del marcador molecular E-cadherina junto con la expresión aberrante por parte de células epiteliales del marcador mesenquimal Vimentina en muestras de MON, DEO y COCE.

The aim was to evaluate the expression of E-cadherin and Vimentin in oral epithelial dysplasia (OED) and oral squamous cell carcinoma (OSCC), in comparison with normal oral mucosa (NOM) in a descriptive case study using immunohistochemistry. A total of fifty-one (N=51) histological samples were included; as follows: n = 16 (NOM), n = 16 (OED) and n = 19 (OSCC). All samples were analyzed using immunohistochemistry against the expression of E-cadherin and Vimentin. Immunostaining was qualitatively evaluated by extent and intensity of its expression for E-cadherin and intensity for Vimentin. Extension and intensity analysis of E-cadherin and Vimentin immunostaining according to group revealed a statistically significant association (r<0.001). E-cadherin expression was found to be highest in NOM followed by OED and lowest in OSCC, inverse to what was observed with Vimentin. The present study revealed the down regulation of the molecular marker E-cadherin, suggestive of reduction in dysplastic cells on comparison to NOM cells, and aberrant expression of the mesenchymal marker Vimentin by epithelial cells in samples of NOM, OED and OSCC; questioning their value as a prognostic marker.

Immunoexpression of cell proliferation markers in oral squamous cell carcinoma / Inmunoexpresiones de marcadores de proliferación celular en carcinoma de células escamosas orales

This study aimed to assess the immunoexpression of cell proliferation markers (Ki-67 and Mcm-2) in oral tongue cancer, correlating it with patients' age and prognostic indicators. Sample was composed of 22 cases under 40 years and 22 over 50 years of age. Clinical staging and histological grade of malignancy were obtained. Cell proliferation was evaluated through labeling indices. Statistical analysis was performed (p<0.05 for statistical significance). Most young patients were stages III/IV (n=13/65 %) and most older patients were stages I/II (n=11/61.1 %) (p>0.05). Mean Ki-67-LI in young and older patients was 42.4 % and 44.15 %, respectively (p>0.05). Mean Mcm-2-LI was higher in older (63.6 %) than in young patients (55.75 %) (p<0.05). We found that young patients presented more aggressive lesions in comparison to older patients, however Mcm-2 expression was significantly higher in older than in young patients. SCC of tongue can be more aggressive in young patients, and this may not be related to cell proliferation. Our findings for Mcm-2 LI and Ki-67 LI suggests that Mcm-2 could be a more sensitive marker for cell proliferation.

Este estudio tuvo como objetivo evaluar la inmunoexpresión de marcadores de proliferación celular (Ki-67 y Mcm-2) en el cáncer de lengua oral, correlacionándolo con la edad de los pacientes y los indicadores pronósticos. La muestra estuvo compuesta por 22 personas menores de 40 años y 22 personas mayores de 50 años. Se identificaron los estadios clínicos y el grado histológico de malignidad. La proliferación celular se evaluó mediante índices de marcado. Se realizó análisis estadístico (p <0,05 para significación estadística). La mayoría de los pacientes jóvenes eran estadios III / IV (n = 13/65 %) y la mayoría de los pacientes mayores eran estadios I / II (n = 11 / 61,1 %) (p> 0,05). La media de Ki-67-LI en pacientes jóvenes y mayores fue 42,4% y 44,15%, respectivamente (p> 0,05). La media de Mcm-2-LI fue mayor en pacientes mayores (63,6 %) que en pacientes jóvenes (55,75 %) (p <0,05). Se encontró que los pacientes jóvenes presentaron lesiones más agresivas en comparación con los pacientes mayores, sin embargo la expresión de Mcm-2 fue significativamente mayor en pacientes mayores que en pacientes jóvenes. SCC de la lengua puede ser más agresivo en pacientes jóvenes, y esto no puede estar relacionado con la proliferación celular. Nuestros hallazgos para Mcm-2 LI y Ki-67 LI sugieren que Mcm-2 podría ser un marcador más sensible para la proliferación celular.

Identificação dos transcritos e proteínas glipicans 1, 3 e 4 em carcinoma escamocelular de boca: associação com moléculas Hedgehog e Vegfa / Identification of transcripts and proteins glypican 1, 3 and 4 in oral squamous cell carcinoma: association with Hedgehog and VEGFA molecules

INTRODUÇÃO: A Via Hedgehog (HH) está ativada em algumas neoplasias humanas, incluindo o Carcinoma Escamocelular de Boca (CEB), o qual corresponde a mais de 95% dos casos diagnosticados na cavidade bucal. Os glipicans (GPC) participam como reguladores desta cascata, atenuando (GPC1 e GPC3) ou regulando positivamente (GPC5) a via HH. OBJETIVO: O objetivo deste trabalho foi avaliar o perfil de expressão dos genes GPC1, 3 e 5, associando-os com genes da via HH (SHH, PTCH1 e SMO) e VEGFA, bem como caracterizar a imunoexpressão das proteínas GPC, em CEB. MATERIAL E MÉTODOS: Trinta e um casos de CEB foram submetidas a reações de qPCR para os genes SHH, PTCH1, SMO, VEGFA, GPC1, 3 e 5. O RNA total foi extraído utilizando uma coluna composta por membrana de silica (Rneasy Mini Kit). O DNA complementar foi obtido com auxílio da enzima Superscript Vilo™. As reações de qPCR foram conduzidas no aparelho ViiA™ 7 Real-Time PCR System utilizando o sistema Taqman, sendo a quantificação relativa avaliada pelo método comparativo de Cq (ΔΔCQ). Vinte e seis CEBs, 9 casos de margens tumorais (MAT) e 4 casos de mucosa bucal não neoplásica (MNN) foram submetidos à reação imuno-histoquímica para as proteínas GPC1, GPC3, GPC5, CD105 e MCM3...

INTRODUCTION: The Hedgehog pathway is activated in some human neoplasms, including Oral Squamous Cell Carcinoma (OSCC), which account for more than 95% of all oral cancers diagnosed. Glypicans are involved in the regulation of HH pathway through GPC3 e GPC1 downregulation or/and GPC5 upregulation. AIM: The aim of this study was to evaluate the expression profile of GPC1, 3 and 5 genes, correlating to HH and VEGFA gene, even as to characterize the immunoexpression of these proteins at OSCC. MATERIAL AND METHODS: A total of 31 cases of OSCC were assessed by qPCR for the SHH, PTCH1, SMO, VEGFA, GPC1, GPC3 and GPC5 genes. The total RNA were extracted using silica membrane column (Rneasy Mini Kit). Complementary DNA was obtained using of Superscript ™ Vilo enzyme. The qPCR reactions were performed in VIIA™ 7 Real-Time PCR System using the Taqman enzime, and relative quantification (RQ) was evaluated by the comparative method of Cq (ΔΔCQ). Immunohistochemical reactions for GPC1, GPC3, GPC5, MCM3 and CD105...

Segmentation of immunohistochemical staining of beta-catenin expression of oral cancer using EM algorithm

Oral Cancer, also called Oral Squamous Cell Carcinoma [OSCC], has been one of the serious cancers that affect the South Asian countries. A range of diagnostic strategies are available including biopsy of the affected part. The Wnt/beta-catenin pathway plays important roles in morphogenesis, normal physiological functions, and tumor formation. This study examined the accumulation of beta-catenin in the nuclei and cytoplasm of oral cancer. The accuracy of histopathological results is hampered by considerable inter and intra-reader variability even by expert pathologists. In order to get both qualitative and quantitative results, we developed a system for diagnosis of oral cancer using Expectatione Maximization [EM algorithm]. The microscopic images of immunohistochemical staining of beta-catenin expression were segmented using Iterative Method of [EM] algorithm to extract the cellular and extracellular components of an image. The segmentation process of the system uses unitone conversion to obtain a single channel image using Principal Component Analysis [PCA] with the highest contrast. Finally, the unitone image is normalized to [0-1] range. Based on the segmentation process we conclude that beta-catenin expression using EM algorithm is an efficient technique to help the pathologist to evaluate the histological changes on microscopic images of oral cancer

Via Hedgehog em carcinoma escamocelular de boca: associação com macrófagos CD163+, angiogênese, ploriferação e diferenciação celular / Hedgehog pathway in mouth squamous cell carcinoma: association with CD163 + macrophages, angiogenesis, and cell differentiation ploriferação

INTRODUÇÃO/OBJETIVO: O Carcinoma Escamocelular de Boca (CEB) corresponde a mais de 95% dos casos de câncer diagnosticados na cavidade bucal e consiste numa neoplasia invasiva e agressiva. Sabendo-se que a via Hedgehog (HH) está envolvida na patogênese de diversos tumores, o presente trabalho propôs-se a avaliar a expressão de componentes desta via em CEB, associando a expressão destas moléculas com aspectos clínicos, angiogênese, graus de diferenciação tumoral, potencial proliferativo e macrófagos CD163+. MATERIAL E MÉTODOS: Vinte e oito casos de CEB, 9 casos de margens tumorais (MAT) e 4 casos de mucosa bucal não neoplásica (MNN) foram submetidos à reação imuno-histoquímica para as proteínas MCM3, SHH, IHH, GLI1, CD163 e CD105 utilizando o sistema polimérico AdvanceTM. A co-localização das proteínas IHH/CD163 e GLI1/CD105 foi avaliada através de dupla marcação imuno-histoquímica. As análises das proteínas MCM3, SHH, IHH e GLI1 foram realizadas em 5 áreas coincidentes de cada caso, de acordo com os parâmetros semi-quantitativos descritos por Gurgel et al. (2008). A densidade de macrófagos (DM) e microdensidade vascular (MDV) foram mensuradas considerando-se a população destas células e vasos neoformados em 5 áreas e os resultados expressos em cel/mm² e vasos/mm². A análise estatística foi realizada utilizando GraphPad Prism versão 6.03. RESULTADOS: Todos os casos de CEB foram positivos para a proteína MCM3, em citoplasma e núcleo de células do parênquima tumoral, sendo o escore 4+ predominante (n=19; 67,85%)...

INTRODUCTION/OBJETIVE: The Oral Squamous Cell Carcinoma (OSCC) accounts for over 95% of all cancers diagnosed in the oral cavity and it consists on an invasive and aggressive type of tumor. The Hedgehog pathway (HH) has been involved in the pathogenesis of different tumors. The aim of this study was to evaluate the components of the HH pathway in OSCC, correlating the results with clinical aspects, angiogenesis, tumor differentiation, proliferative potential and macrophages CD163+. MATERIAL AND METHODS: Twenty-eight cases of OSCC, 9 cases of tumor margins (TM) and 4 cases of non-neoplastic oral mucosa (NNM) were submitted to immunohistochemical reaction for MCM3, SHH, IHH, GLI1, CD163 and CD105 proteins using the AdvanceTM polymer system. Co-localization for IHH/GLI1 and CD163/CD105 proteins was evaluated using double staining method. The analysis of MCM3, SHH, IHH and GLI1 proteins were conducted in 5-matching areas and data described using the semi-quantitative parameters described by Gurgel et al. (2008). The density of macrophages (MD) and microvessel density (MVD) were measured considering the population of these cells and newly formed vessels in 5-matching areas and the results expressed in cells/mm² and vessels/mm², respectively. Statistical analysis were performed using GraphPad Prism v. 6.03. RESULTS: All cases of OSCC were positive for MCM3 protein on the cytoplasm and nucleus of tumor cells, and 4+ was the main score (n= 19; 67.85%)...

Expressão imuno-histoquímica das proteínas ki-67, MCM3 e p27 em leucoplasias e carcinoma epidermóide de boca / Imunohistochemical expression of ki-67 protein, MCM3 and p27 in leukoplakia and squamous cell carcinoma of the mouth

O propósito deste estudo foi identificar o padrão de expressão de Ki-67, MCM3 e p27 em mucosa normal (MN), em leucoplasias de orais ( leves, moderadas e intensas ) e em carcinomas epidermoides de boca (CEB), com objetivo de sugerir a participação dessas proteínas na aquisição do fenótipo malígno. Os espécimes foram removidos de 37 pacientes com leucoplasias bucais (13 com displasias leves- DL, 12 com displasias moderadas-DM e 12 com displasias intensas-DI). Onze amostras de mucosa normal de soalho bucal (MN) e 50 amostras de CEB de soalho e língua de pacientes não tratados foram incluídas neste estudo. As amostras foram imunomarcadas com anticorpos contra, Ki-67, MCM3 e p27. O teste Kruskal Wallis e o teste de Dunns foram usados para determinar a diferença dos grupos entre si. Para avaliar a correlação das proteínas entre si foi utilizado o teste de correlação de Pearson. Ki-67 mostrou índices de marcação estatisticamente maior em casos de CEB quando comparados com o grupo que continha MN (p<0.001) e DL (p<0.01) e índices de marcação estatisticamente menor na MN quando comparada com os grupos que continham DM e DI (p<0.05). A proteína p27 mostrou índice de expressão estatisticamente menor quando as amostras com CEB foram comparadas com os grupos que continham a MN, DL; DM e DI (p<0.001). A proteína MCM3 mostrou índices de marcação estatisticamente menor no grupo que continha a MN quando se correlacionou com o que apresentava DI e CEB (p<0.001) e o grupo com DL mostrou índices estatisticamente menores quando foi correlacionado tanto com o grupo de DI (p0.01)Outros resultados dignos de nota foram a significante correlação entre as proteínas MCM3 e p27 ( r=0.31 e p< 0.03 ), Ki-67 e p27 ( r=0.31 e p< 0,03281 ) e MCM3 e ki- 67 ( r=0,69 p< 0,01 ) em CEB e MCM3 e p27 ( r=0.58 e p< 0.05 ) em displasia ...

The aim of this study was to identify the expression of MCM3, Ki-67 and p27 in normal mucosa (MN) in leukoplakia and Oral Squamous Cell Carcinoma Epidemiology (OSCCE), and to determine whether altered expression could serve as a prognostic marker of a malignant progression of dysplasia lesions. The samples were collected from 37 patients with oral leukoplakia (13 with mild dysplasia - MLD, 12 with moderate dysplasia - MD and 12 with severe dysplasia - SD). Eleven samples of mouth floor mucocele (M) and 50 floor and tongue samples OSCCE of untreated patients were included in this study. Immunohistochemical MCM3, Ki-67 and p27 expression of all the groups was analyzed. Kruskal Wallis and Dunns test were used to determine differences among groups, and a Pearson correlation test was used to evaluate the correlation between the proteins. Ki-67 expression was higher in OSCCE than M (p0.001) and MLD (p0.01) groups, and there was a lower expression in MN compared with MD and ID (p0.05). Ki-67 expression did not show significant difference between MD, SD and OSCCE. Regarding p27, its expression was lower in OSCCE compared with M, MD and SD (p<0,001). MCM3 expression was lower in M compared with SD and OSCCE (p0.001) and MLD showed a lower expression when compared SD (p0.01) and OSCCE (p0.001). The present study showed that MCM3 could be a better marker than Ki67 for evaluation dysplastic oral lesions.

Immunohistochemical evaluation of MMP-2, MMP-9 and CD31/microvascular density in squamous cell carcinomas of the floor of the mouth

The aim of this study was to evaluate the immunoexpression of MMP-2, MMP-9 and CD31/microvascular density in squamous cell carcinomas of the floor of the mouth and to correlate the results with demographic, survival, clinical (TNM staging) and histopathological variables (tumor grade, perineural invasion, embolization and bone invasion). Data from medical records and diagnoses of 41 patients were reviewed. Histological sections were subjected to immunostaining using primary antibodies for human MMP-2, MMP-9 and CD31 and streptavidin-biotin-immunoperoxidase system. Histomorphometric analyses quantified positivity for MMPs (20 fields per slide, 100 points grade, ×200) and for CD31 (microvessels <50 µm in the area of the highest vascularization, 5 fields per slide, 100 points grade, ×400). Statistical design was composed by non-parametric Mann-Whitney U test (investigating the association between numerical variables and immunostainings), chi-square frequency test (in contingency tables), Fisher's exact test (when at least one expected frequency was less than 5 in 2×2 tables), Kaplan-Meier method (estimated probabilities of overall survival) and Iogrank test (comparison of survival curves), all with a significance level of 5%. There was a statistically significant correlation between immunostaining for MMP-2 and lymph node metastasis. Factors associated negatively with survival were N stage, histopathological grade, perineural invasion and immunostaining for MMP-9. There was no significant association between immunoexpression of CD31 and the other variables. The intensity of immunostaining for MMP-2 can be indicative of metastasis in lymph nodes and for MMP-9 of a lower probability of survival.

O objetivo deste estudo foi avaliar a imunoexpressão de MMP-2, MMP-9 e CD31/densidade microvascular em carcinomas espinocelulares de soalho bucal e correlacionar os resultados com variáveis demográficas, de sobrevida, clínicas (estadiamento TNM) e histopatológicas (grau de diferenciação tumoral, invasão perineural, embolização e invasão óssea). Dados de prontuários e de diagnósticos de 41 pacientes foram revisados. Cortes histológicos foram submetidos à imunomarcação usando anticorpos primários para MMP-2, MMP-9 e CD31 humanos e sistema streptoavidina-biotina-imunoperoxidase. Análise histomorfométrica quantificou a positividade para MMPs (20 campos, grade de 100 pontos por lâmina, ×200) e para CD31 (microvasos <50 µm na área de maior vascularização, 5 campos, grade de 100 pontos por lâmina, ×400). O planejamento estatístico foi composto pelo teste não paramétrico U de Mann-Whitney (verificação da associação entre variáveis numéricas e imunomarcações), teste de frequências do qui-quadrado (em tabelas de contingência), teste exato de Fisher (quando pelo menos uma frequência esperada foi menor do que 5 em tabelas 2×2), método de Kaplan-Meier (estimativa de probabilidades de sobrevida global) e teste de Iogrank (comparação das curvas de sobrevida), todos com nível de significância de 5%. Houve correlação estatisticamente significante entre imunomarcação para MMP-2 e metástase em linfonodo. Os fatores relacionados negativamente com a sobrevida foram estadiamento N, gradação histopatológica, invasão perineural e imunomarcação de MMP-9. Não houve associação significativa entre imunoexpressão de CD31 e as demais variáveis. A intensidade de imunomarcação para MMP-2 pode ser indicativa de metástase em linfonodo e para MMP-9 de uma menor probabilidade de sobrevida.

Downregulation of C3 and C4A/B complement factor fragments in plasma from patients with squamous cell carcinoma of the penis

To investigate the use of ClinProt technique to identify cancer markers in plasma of patients suffering from squamous cell carcinoma of the penis (SCCP).

Plasma of 36 healthy subjects and 25 patients with penile carcinoma who underwent surgical treatment between June 2010 and June 2011 was collected and analyzed by the ClinProt/MALDI/ToF technique. Then the peptides were identified from the C8 MB eluted fraction of patients' and control subjects' plasma by LIFT MS/MS.

A cluster of 2 peptides (A=m/z 1897.22 ± 9 Da and B=m/z 2021.99 ± 9 Da) was able to discriminate patients from control subjects. Cross validation analysis using the whole casuistic showed 62.5% and 86.76% sensitivity and specificity, respectively. The cluster also showed very high sensitivity (100%) and specificity (97%) for SCCP patients that died due to the disease. Furthermore, patients with lymph node involvement presented sensitivity and specificity of 80% and 97%, respectively. These two peptides were identified by the proteomic approach based on a MALDI-TOF/TOF as fragments of C3 (m/z 1896.17) and C4a/b (m/z 2021.26) complement proteins.

The results showed that as the disease progresses, the fragments C3 and C4 A/B are less expressed in comparison with healthy subjects. These results may be useful as prognostic tools. .

Expressão de imunomarcadores em carcinomas epidermoides de boca / Expression of biological markers in oral squamous cell carcinomas

Squamous cell carcinomas are the most commonly diagnosed oral malignancy, accounting for about 90% of all malignant oral lesions. Detection of the condition at early stages is rare; as a result, the clinical and histological characteristics and prognosis of this tumor have not been extensively investigated. The objective of this study was to evaluate clinical and microscopic features of squamous cell carcinomas using immunohistochemical analysis and assessing biological markers of angiogenesis and tumor vascular activity (anti-CD31, anti-CD34, Factor VIII), cell proliferation (Ki-67), and loss of cell suppression (p53). Tolonium chloride 1% was used to determine the optimal biopsy site. Six patients seen at the Stomatology Service of a university hospital in Canoas, southern Brazil, with a suspected diagnosis of squamous cell carcinoma were analyzed. All patients were male, with a mean age of 56.6 years, and four had a white skin color. Lesions were detected in the tongue4 and tonsillar pillar2. All diagnoses were confirmed by microscopy (hematoxylin-eosin staining). Immunohistochemical analysis revealed p53 expression in 5 of the cases, Ki-67 in 6, and anti-CD34 in 1; anti-CD31 and Factor VIII were not detected in any patient. Our findings suggest an important contribution of tumor markers in the diagnosis and prognosis of these malignancies, as well as in treatment planning.

O carcinoma epidermoide é a neoplasia maligna bucal mais frequentemente diagnosticada, correspondendo a aproximadamente 90% de todas lesões malignas bucais. Sua detecção nos estágios iniciais da doença é rara; como resultado, tem sido difícil avaliar o comportamento clínico e histológico e o prognóstico deste tumor. O objetivo deste estudo foi avaliar clínica e microscopicamente o carcinoma epidermoide através de análise imuno-histoquímica, avaliando imunomarcadores tumorais angiogênicos e de atividade vascular tumoral (anticorpos anti-CD31, anti-CD34, Fator VIII), de proliferação celular (Ki-67) e de perda de supressão celular (p53). Azul de toluidina a 1% foi utilizado para determinar o local ideal da biópsia incisional. Foram analisados seis pacientes oriundos do Serviço de Estomatologia do hospital universitário localizado em Canoas, sul do Brasil, com hipótese diagnóstica de carcinoma epidermoide. Todos os pacientes eram do sexo masculino, com idade média de 56,6 anos, e quatro tinham pele branca. As lesões foram detectadas na língua4 e no pilar amigdaliano2. Todos os diagnósticos foram confirmados por microscopia (hematoxilina-eosina). A análise imuno-histoquímica revelou expressão de p53 em 5 dos casos, Ki-67 em 6 e anti-CD34 em 1; anti-CD31 e Fator VIII não apresentaram expressividade. Nossos resultados sugerem uma contribuição importante dos marcadores tumorais no diagnóstico e prognóstico dessas neoplasias, bem como no planejamento terapêutico.

IFNα2b augments immune responses of cisplatin+5-fluorouracil treated tongue squamous cell carcinoma patients - A preliminary study.

Background & objectives: Interferon alpha 2b (IFNα2b) has been reported to regulate several immune functions efficiently to enhance the cytotoxic activity of NK and T cells towards various forms of tumours. The objective of the present study was to evaluate the efficacy of IFNα2b in overcoming disease induced and/or treatment associated imunosuppression of tongue squamous cell carcinoma (TSCC) patients undergoing chemotherapy for better clinical outcome. Methods: Seven TSCC patients under cisplatin + 5-fluorouracil chemotherapy in combination with IFNα2b were assessed for various immunohaematological parameters before treatment, after chemotherapy and after IFNα2b therapy. Results: Deterioration of the haematological and immune responses was detected in immunosuppressed TSCC patients after chemotherapy. IFNα2b treatment led to a recovery in these parameters in most of the patients. Greater number of T/NK cells and enhanced secretion of type 1 cytokines were also noted. Haematological complications were reduced after completion of the therapy. Immune- and haematostimulation were also observed in patients with partial response. No positive clinical response was detected in one patient. Interpretation & conclusions: IFNα2b appears to be an effective immunostimulator having clinical impact to combat the immunosuppression in TSCC patients. Successful immunostimulation by IFNα2b may help TSCC patients in clinical improvement. The findings of this preliminary study need to be confirmed on a large number of patients with TSCC.

HLA and skin cancer / HLA e câncer de pele

Skin cancer - melanoma and non melanoma - are common neoplasm with rising incidence over the last decades. It is an important public health problem. Its pathogenesis is not completely understood and the same happens with the genetic factors involved. The genes that encode the HLA are associated with some tumors and they may be responsible for one of the mechanisms that take part in the development of the before mentioned cancers. We have reviewed the literature on the subject of HLA antigens, melanoma and non melanoma skin cancer.

Os cânceres da pele - melanoma e não-melanoma - são neoplasias comuns e com incidência crescente ao longo de décadas. Representam um importante problema de saúde pública. A patogênese destas neoplasias não é completamente compreendida, assim como não o são os fatores genéticos envolvidos. Os genes HLA estão associados a alguns tumores e podem representar um dos mecanismos implicados no desenvolvimento do câncer de pele. Apresenta-se uma revisão atualizada sobre a relação entre antígenos HLA, câncer da pele não-melanoma e melanoma.

Immunomorphological assessment of regional lymph nodes for predicting metastases in oral squamous cell carcinoma.

Background: Oral squamous cell carcinoma is the most common neoplasm and comprises of approximately 80% of the cancers occurring in the oral cavity. The role of the host response to this neoplasm has been recognized, and for many years the regional lymph node in tumor-bearing hosts has been considered as an anatomic barrier to the systematic dissemination of tumor cells. Morphological evaluation of the regional nodes has aided in understanding the immune response. Aim: The current study was carried out to observe the morphological changes occurring in the regional lymph nodes and to evaluate whether these features could be helpful in assessing the immunological status of the patient, and thereby, the prognosis of the patient. Materials and Methods: The study was based on lymph nodes from 63 patients with oral squamous cell carcinoma, who underwent radical neck dissection or modified neck dissection. In the lymph node, four morphological patterns were observed that included lymphocyte predominance, germinal center predominance, mixed pattern (sinus Histiocytosis), and an unstimulated pattern. The cases were then divided into four groups according to the predominant immunoreactivity pattern based on the World Health Organization (WHO) standardized system for reporting human lymph node morphology. Results: Revealed that risk of metastases to cervical lymph nodes in patients with lymphocyte predominance was less (28.6%) when compared to the high risk of metastases with germinal center predominance (68%), and these results were statistically significant (P < 0.05). Patients with a mixed pattern showed less risk of metastases (45.4%), while those with an unstimulated pattern had increased risk of metastases (66.6%), but the results were not statistically significant. It was also found that in the positive nodes, germinal center hyperplasia (50.2%) was the predominant pattern. Conclusion: The present study revealed that patients with lymphocyte predominance had less risk of metastases and patients with germinal center predominance had a high risk of metastases to the lymph node.

Morphometric analysis of dendritic cells from anal mucosa of HIV-positive patients and the relation to intraepithelial lesions and cancer seen at a tertiary health institution in Brazil / Análise morfométrica das células dendríticas da mucosa anal de pacientes HIV-positivos e relação com as lesões intraepiteliais e o câncer numa instituição de saúde terciária no Brasil

PURPOSE: To morphometrically quantify CD1a+ dentritic cells and DC-SIGN+ dendritic cells in HIV-positive patients with anal squamous intraepithelial neoplasia and to evaluate the effects of HIV infection, antiretroviral therapy and HPV infection on epithelial and subepithelial dendritic cells. METHODS: A prospective study was performed to morphometrically analyze the relative volume of the dendritic cells and the relationship between anal intraepithelial neoplasia and cancer in HIV-positive patients from the Tropical Medicine Foundation of Amazonas, Brazil. All patients were submitted to biopsies of anorectal mucosa to perform a classic histopathological and immunohistochemical analysis, employing antibodies against CD1a and DC-SIGN for the morphometric quantification of dendritic cells. RESULTS: HIV-negative patients displayed a CD1a DC density significantly higher than that of HIV-positives patients (3.75 versus 2.54) (p=0.018), and in patients with severe anal intraepithelial neoplasia had correlated between DC CD1a density with levels of CD4 + cells (p: 0.04) as well as the viral load of HIV-1 (p: 0.035). A not significant rise in the median density of CD1a+ DC was observed in the HIV positive/ HAART positive subgroup compared to the HIV positive/ HAART negative subgroup. The CD1a+ DC were also significantly increased in HIV-negative patients with anorectal condyloma (2.33 to 3.53; p=0.05), with an opposite effect in HIV-positive patients. CONCLUSIONS: Our data support an enhancement of the synergistic action caused by HIV-HPV co-infection on the anal epithelium, weakening the DC for its major role in immune surveillance. Notoriously in patients with severe anal intraepithelial neoplasia, the density of CD1a+ epithelial dendritic cells was influenced by the viral load of HIV-1. Our study describes for the first time the density of subepithelial DC-SIGN+ dendritic cells in patients with anal severe anal intraepithelial neoplasia and points to the possibility that a specific therapy for HIV induces the recovery of the density of epithelial DC.

OBJETIVO: Quantificar morfometricamente as células dendríticas DC CD1a+ e DC DC-SIGN+ em pacientes HIV positivos portadores de neoplasia escamosa intraepitelial anal e avaliar os efeitos da infecção pelo HIV, da terapia antirretroviral e da infecção pelo HPV sobre as células dendríticas epiteliais e subepiteliais. MÉTODOS: Um estudo prospectivo foi realizado para analisar morfometricamente o volume relativo das células dendríticas e as relações entre neoplasia intraepitelial anal e o câncer em pacientes HIV positivos da Fundação de Medicina Tropical do Amazonas, Brasil.Todos os pacientes foram submetidos a biópsia da mucosa retal para realizar uma análise clássica histopatológica e imunohistoquímica utilizando anticorpos contra anti-CD1a e anti-DC-SIGN, para a quantificação morfométrica das células dendríticas. RESULTADOS: Os pacientes HIV negativos apresentaram densidade das DC CD1a+ significativamente maior do que a dos pacientes HIV positivos (3,75 versus 2,54) (p:0,018), e os pacientes com severa apresentaram correlação das DC CD1a com os níveis de células TCD4(p:0,04) assim como a carga viral do HIV-1 (p:0,035). Observamos no subgrupo HIV-positivo/HAART positivo elevação não significativa na mediana da densidade das DC CD1a+ em relação ao grupo HIV-positivo/HAART negativo. As DC CD1a+ também se elevaram nos pacientes HIV negativo portadores de condiloma anorretal(2,33 para 3,53; p:0,05), com efeito inverso nos pacientes HIV positivos. CONCLUSÕES: Nossos dados confirmam a potencialização da ação sinérgica representada pela coinfecção HIV-HPV sobre o epitélio anal, fragilizando as DC em sua função primordial de vigilância imune. Notoriamente nos pacientes com neoplasia intraepithelial anal grave, a densidade das DC CD1a+ epiteliais sofreu influência da carga viral do HIV-1. Nosso estudo descreveu pela primeira vez a densidade das DC subepiteliais DC-SIGN+ em pacientes com neoplasia intraepithelial anal severa e apontamos para a possibilidade de que a terapia específica para o HIV induza a recuperação da densidade das DC epiteliais.

Response of peripheral blood mononuclear cells to conditioned medium from cultured oral squamous cell carcinomas

The current study investigated the capacity for tumor factors secreted by head and neck squamous cell carcinoma (HNSCC) cell lines, KB, KB16, and HEP, to induce the secretion of various cytokines from peripheral blood mononuclear cells (PBMCs). PBMCs were isolated from blood samples collected from six healthy volunteers and these cells were incubated for 6, 24, 48, or 72 hours in the presence of 50 percent conditioned medium collected from cultured cell lines pretreated with, or without, stimulants such as phytohemagglutinin (PHA) or lipopolysaccharides (LPS). Aliquots of each supernatant were then assayed for levels of IFN-Γ, vascular endothelial growth factor (VEGF), TNF-α, and IL-4 using enzyme linked immunosorbent assays (ELISAs). Data collected were analyzed using Student's t-test, an ANOVA test followed by Tukey's test, and tests of Pearson's Correlation. PBMCs cultured with KB16-conditioned medium produced the highest levels of IFN-Γ. VEGF was also detected in conditioned media collected from all of the squamous cell carcinoma (SCC) cell lines used, and a significant difference in VEGF levels between control and KB- or KB16-conditioned media was observed. TNF-α was secreted by all PBMC groups within 6 hours of receiving conditioned media, and these levels increased up to the 24 hour timepoint, after which levels of TNF-α stabilized. In contrast, none of the supernatant samples contained detectable levels of IL-4. In combination, these data suggest that direct contact between fresh human PBMCs and conditioned media from tumor cells induces the secretion of TNF-α and VEGF by PBMCs, and this represents an initial angiogenic response.

Immunohistochemical staining of Langerhans cells in HPV-positive and HPV-negative cases of oral squamous cells carcinoma

The Human Papillomavirus (HPV) has been strongly implicated in development of some cases of oral squamous cell carcinoma (OSCC). However, the immunological system somehow reacts against the presence of this virus. Among the cells involved in such mechanism of defense Langerhans cells (LC) stand out, which are responsible for processing and presenting antigens. OBJECTIVES: The purposes of this study were to investigate the presence of HPV DNA and to evaluate the immunohistochemical reactivity for Langerhans cells between HPV-positive and HPV-negative OSCC. Twenty-seven cases of OSSC were evaluated. MATERIAL AND METHODS: DNA was extracted from paraffin-embedded tissue samples and amplified by Polymerase Chain Reaction (PCR) for the detection of HPV DNA. Viral typing was performed by dot blot hybridization. Immunohistochemistry was performed by the Streptavidin-biotin technique. RESULTS: From the 27 cases, 9 (33.3 percent) were HPV-positive and 18 (66.0 percent) HPV-negative. HPV 18 was the most prevalent viral type (100 percent cases) and infection with HPV-16 (co-infection) was detected in only 1 case. In the OSCC specimens examined, immunoreactivity to S-100 antibody was detected in all cases, with a mean number of 49.48±30.89 Langerhans cells positive for immunostaining. The mean number of immunostained Langerhans cells was smaller in the HPV-positive cases (38 cells/case) than in the HPV-negative cases (42.5 cells/case), but this difference was not significant (p=0.38). CONCLUSIONS: The low frequency of detection of HPV DNA in OSCC indicates a possible participation of the virus in the development and progression of only a subgroup of these tumors. There was no association between the immunohistochemical labeling for Langerhans cells (S-100+) and HPV infection of in OSSC. These findings suggest that the presence of HPV in such OSCC cases could not alter the immunological system, particularly the Langerhans cells.

Effect of suicide gene therapy in combination with immunotherapy on antitumour immune response & tumour regression in a xenograft mouse model for head & neck squamous cell carcinoma.

Background & objectives: Prodrug activation strategy as well as immunotherapy have been widely used for cancer gene therapy. In the present study, using a head and neck squamous cell carcinoma (HNSCC) xenograft nude mouse model, we have investigated whether the two therapies in combination could improve tumour cell kill. We also investigated induction of immune effector cells viz., NK (DX5+) and DC (CD11c+) in vivo, post-combination gene therapy. Methods: A retroviral vector producing cell line (PLTK47.1 VPC) carrying Herpes simplex virus thymidine kinase gene (HSVtk) was used for intratumoural injection into NT8e xenograft tumours followed by the prodrug ganciclovir (GCV). IL-2 plasmid DNA was injected intramuscularly. Immune cells were analyzed by flow-cytometry. Non parametric ANOVA was performed with Kruskal Wallis test. Results: IL-2 could induce proliferation of both NK cells (DX5+) and dendritic cells (CD11c+) in vivo. Apoptosis was higher in combination therapy group as compared to HSVtk/GCV alone or IL-2 alone and was mediated through caspase-3 dependent pathway. Significant reduction in tumour volume was seen in all 3 treatment arms as compared to controls. Interpretation & conclusions: Combination of suicide gene therapy and immunotherapy leads to successful tumour regression in a HNSCC xenograft mouse model. Immunotherapy could help in a systemic long lived anti-tumour immune response which would prove powerful for the treatment of metastatic cancers, and also for minimal residual disease. The results of this study may form the basis for Phase 1 clinical trials.

Rol de las características clínicas e histológicas como factores pronósticos para la sobrevida en pacientes con carcinoma de células escamosas de la cavidad oral / Oral squamous cell carcinoma: restrospective analysis of 36 cases

Background: Survival of patients with oral squamous cell carcinoma is low and depends mostly on TNM staging of the tumor. Aim: To perform a retrospective analysis of a series of patients with oral squamous cell carcinoma. Material and Methods: Retrospective review of patients with the diagnosis, seen between 1995 and 2006 in a regional hospital. Host inflammatory response and pattern of tumor invasion were assessed using the staging proposed by Bryne et al. Results: The medical records of 36 patients aged 39 to 89 years, were reviewed. During the study period, 15 patients died. Better survival was associated to a low pattern of tumor invasion and a high inflammatory response and the topographic location of the tumor. Conclusions: Inflammatory response, tumor invasion and location are associated with survival in oral squamous cell carcinoma.

Introducción: El carcinoma de células escamosas de la cavidad oral (CCECO) es una patología cuyo comportamiento es producto de interrelaciones con el huésped, esto es, por el patrón de invasión (PI) histopatológica y la respuesta inflamatoria (RI). El objetivo de este estudio es analizar las características clínicas e histopatológicas como factores pronóstico, en términos de supervivencia (SV) en pacientes con CCECO. Material y Método: Serie de casos. Se incluyeron pacientes diagnosticados en el Hospital Regional de Talca y Hospital Base de Curicó entre los años 1995 y 2006. Se revisaron las fichas clínicas y biopsias de 36 pacientes con CCECO. Se determinó el Frente Invasivo Tumoral (FIT), evaluándose los parámetros propuestos por el sistema de graduación de Bryne (PI y RI) y factores de importancia clínica como localización topográfica de la lesión, edad y género, relacionándolos con SV mediante curvas de Kaplan-Meier y Log Rank test. Posteriormente, se aplicó una regresión de Cox para obtener un análisis multivariado y cálculo de RR. Del total de casos del estudio, 15 pacientes fallecieron por CCECO. Resultados: La mayor SV se asoció a un bajo escore de PI y una alta RI respectivamente (RR 1,5). La localización topográfica de la lesión se relacionó significativamente con la SV, no así el grupo de edad. Conclusiones: Nuestros resultados sugieren que la ubicación de la lesión es un factor importante en el pronóstico de la enfermedad y que una respuesta inmune/inflamatoria adecuada, expresada en un bajo escore de RI, mejora el pronóstico de SV en pacientes con CCECO.

Field cancerization in head and neck squamous cell carcinoma: immunohistochemical expression of P53 and Ki67 proteins: clinicopathological study / Cancerização por campo em carcinoma de células escamosas: expressão imuno-histoquímica de proteínas p53 e Ki67: estudo clínico-patológico

OBJECTIVE: This study is aimed to evaluate the immunostaining influence of p53 and Ki67 proteins inareas of field cancerization of head and neck squamous cell carcinoma (HNSCC). We analyzed associationsof these proteins with clinicopathological parameters and the relation between their immunoexpression inHNSCC. MATERIAL AND METHOD: In a retrospecive analysis, 40 patients with HNSCC were selectedaccording to the recurrence of the disease, forming two groups: recurrent and non-recurrent HNSCC.Morphological gradations and imunnohistochemical analysis of p53 and Ki67 were performed in invasivefront and tumor adjacent epithelium. RESULTS: It was found significant associations between tumor recurrenceand p53 positivity in mucosa and invasive front. However, no association was found between p53immunostaining and the clinicopathological parameters. Ki67 was not related to any clinicopathologicalparameter either. The association between Ki67 and p53 expression was not significant. There was no significant inluence of recurrence in the clinicopathological parameters. Individuals with T1/T2 tumor size, non-recurrentand p53-negative in tumor adjacent epithelium presented better overall survival of HNSCC. CONCLUSION:p53 positivity in adjacent epithelium and invasive front of recurrent HNSCC is suggested in this study.

OBJETIVO: Avaliar a influência da imunoexpressão das proteínas p53 e Ki67 em áreas de campos de cancerização do carcinoma de células escamosas da cabeça e pescoço. Analisamos a associação dessas proteínas com parâmetros clínico-patológicos e a relação entre sua imunoexpressão.MATERIAL E MÉTODO: Em análise retrospectiva, 40 pacientes foram selecionados, de acordo com a recorrência da doença, formando dois grupos: com recorrência e sem recorrência da neoplasia. Gradações morfológicas e análises histoquímicas foram efetuadas na área de invasão e no epitélio adjacente ao tumor. RESULTADOS: Encontrou-se associações significativas entre a recorrência do tumor e positividade para p53 na mucosa e na área da lesão. Entretanto, não encontrou-se associação entre p53 e parâmetros clínico-patológicos. Ki67 não foi relacionada com qualquer parâmetro, igualmente. A associação entre expressão de Ki67 e p53 não foi significante e não houve influência significante de recorrência nos parâmetros clínico-patológicos. Indivíduos com tumores T1/T2, não recorrentes, e p53 negativos no epitélio adjacente ao tumor apresentaram melhor sobrevida à neoplasia. CONCLUSÃO: Positividade para p53 no epitélio adjacente e na área da neoplasia de carcinoma de células escamosas é sugerida por este estudo.