Differential Expression of ADAM23, CDKN2A (P16), MMP14 and VIM Associated with Giant Cell Tumor of Bone.

Though benign, giant cell tumor of bone (GCTB) can become aggressive and can exhibit a high mitotic rate, necrosis and rarely vascular invasion and metastasis. GCTB has unique histologic characteristics, a high rate of multinucleated cells, a variable and unpredictable growth potential and uncertain biological behavior. In this study, we sought to identify genes differentially expressed in GCTB, thus building a molecular profile of this tumor. We performed quantitative real-time polymerase chain reaction (qPCR), immunohistochemistry and analyses of methylation to identify genes that are putatively associated with GCTB. The expression of the ADAM23 and CDKN2A genes was decreased in GCTB samples compared to normal bone tissue, measured by qPCR. Additionally, a high hypermethylation frequency of the promoter regions of ADAM23 and CDKN2A in GCTB was observed. The expression of the MAP2K3, MMP14, TIMP2 and VIM genes was significantly higher in GCTB than in normal bone tissue, a fact that was confirmed by qPCR and immunohistochemistry. The set of genes identified here furthers our understanding of the molecular basis of GCTB.

High-risk human papillomaviruses in two different primary tumors in the same patient.

Two cases of patients with high-risk human papillomavirus-related squamous cell carcinomas of the penis are reported. In both patients, a second high-risk human papillomavirus-related squamous cell carcinoma, of the same type (genotype 16), was detected: a carcinoma of the oropharynx 2 years after treatment of the squamous cell carcinomas of the penis in the first patient, and a carcinoma of the esophagus 1 year after the treatment of the squamous cell carcinomas of the penis in the second patient. To the best of our knowledge, this is the first time that multiple human papillomavirus-related tumors in the same patient are reported. It is suggested that a careful clinical investigation is necessary in patients with tumors attributable to high-risk human papillomavirus for the early detection of a possible second neoplasm related to this virus in a different organ.

Overexpression of ANXA1 in penile carcinomas positive for high-risk HPVs.

The incidence of penile cancer varies between populations but is rare in developed nations. Penile cancer is associated with a number of established risk factors and associated diseases including phimosis with chronic inflammation, human papillomavirus (HPV) infection, poor hygiene and smoking. The objective of this study was to identify genes related to this type of cancer. The detection of HPV was analyzed in 47 penile squamous cell carcinoma samples. HPV DNA was detected in 48.9% of penile squamous cell carcinoma cases. High-risk HPV were present in 42.5% of cases and low-risk HPV were detected in 10.6% of penile squamous cell carcinomas. The RaSH approach identified differential expression of Annexin A1 (ANXA1), p16, RPL6, PBEF1 and KIAA1033 in high-risk HPV positive penile carcinoma; ANXA1 and p16 were overexpressed in penile squamous cells positive for high-risk HPVs compared to normal penile samples by qPCR. ANXA1 and p16 proteins were significantly more expressed in the cells from high-risk HPV-positive penile carcinoma as compared to HPV-negative tumors (p<0.0001) independently of the subtype of the carcinoma. Overexpression of ANXA1 might be mediated by HPV E6 in penile squamous cell carcinoma of patients with high-risk HPVs, suggesting that this gene plays an important role in penile cancer.

Results of a control quality strategy in cervical cytology.

OBJECTIVE: To determine the efficacy of a quality control strategy in cervical cytology in the detection of high-grade squamous intraepithelial lesions. METHODS: Forty-two patients were selected who underwent a Pap smear and cervical uterine biopsy between April 2008 and December 2009, with evidence of a high-grade squamous intraepithelial lesion in one or both tests. The statistical parameters of the smear test were calculated before and after systematic meetings for review of the archived test results (6 years), in which the following was done: interobserver diagnostic consensus; cytohistological correlation, with the latter as gold standard; and evaluation of the therapeutic status of each patient. RESULTS: Once these controls were applied, it was noted that sensitivity and positive likelihood ratio of the test for high-grade squamous intraepithelial lesion increased 9.5% (34.5 to 44%) and 0.45% (1.64 to 2.09%), respectively, while specificity remained at 79%. Reduction in interference of false-negative results associated with errors in the analytical phase of the cytological productive process gave an estimate of failures in collection of the specimens (pre-analytical phase). CONCLUSION: In addition to improving the performance of the cytological diagnosis of the high-grade squamous intraepithelial lesion, the proposed quality control strategy allows a reflection on the causes of incorrect or conflicting scrutiny.

Results of a control quality strategy in cervical cytology / Resultados de uma estratégia de controle de qualidade em colpocitologia

Objective: To determine the efficacy of a quality control strategy in cervical cytology in the detection of high-grade squamous intraepithelial lesions. Methods: Forty-two patients were selected who underwent a Pap smear and cervical uterine biopsy between April 2008 and December 2009, with evidence of a high-grade squamous intraepithelial lesion in one or both tests. The statistical parameters of the smear test were calculated before and after systematic meetings for review of the archived test results (6 years), in which the following was done: interobserver diagnostic consensus; cytohistological correlation, with the latter as gold standard; and evaluation of the therapeutic status of each patient. Results: Once these controls were applied, it was noted that sensitivity and positive likelihood ratio of the test for high-grade squamous intraepithelial lesion increased 9.5% (34.5 to 44%) and 0.45% (1.64 to 2.09%), respectively, while specificity remained at 79%. Reduction in interference of false-negative results associated with errors in the analytical phase of the cytological productive process gave an estimate of failures in collection of the specimens (pre-analytical phase). Conclusion: In addition to improving the performance of the cytological diagnosis of the high-grade squamous intraepithelial lesion, the proposed quality control strategy allows a reflection on the causes of incorrect or conflicting scrutiny.

Objetivo: Determinar a eficácia de uma estratégia de controle de qualidade em colpocitologia na detecção da lesão intraepitelial escamosa de alto grau. Métodos: Foram selecionadas 42 pacientes que realizaram Papanicolaou e biópsia cervicouterina entre abril de 2008 e dezembro de 2009, com evidência de lesão intraepitelial escamosa de alto grau em um ou em ambos os exames. Os parâmetros estatísticos do esfregaço foram calculados antes e após reuniões sistematizadas de revisão dos exames arquivados (6 anos), nas quais se procedeu a: consensualização diagnóstica interobservadores; correlação cito-histológica, sendo a última padrão-ouro; e avaliação do status terapêutico de cada paciente. Resultados: Aplicados tais controles, observou-se que a sensibilidade e a likelihood ratio positiva do teste para lesão intraepitelial escamosa de alto grau aumentaram 9,5% (34,5 para 44%) e 0,45% (1,64 para 2,09%), respectivamente, enquanto sua especificidade se manteve em 79%. A redução da interferência dos falso-negativos associados a erros na fase analítica do processo produtivo citológico traz estimativa das falhas de coleta do material (fase pré-analítica). Conclusão: Além de melhorar o desempenho do diagnóstico colpocitológico de lesão intraepitelial escamosa de alto grau, a estratégia de controle de qualidade proposta permite refletir sobre as causas de escrutínio incorreto ou discordante.

Differentially expressed genes in giant cell tumor of bone.

Giant cells tumors of bone (GCTB) are benign in nature but cause osteolytic destruction with a number of particular characteristics. These tumors can have uncertain biological behavior often contain a significant proportion of highly multinucleated cells, and may show aggressive behavior. We have studied differential gene expression in GCTB that may give a better understanding of their physiopathology, and might be helpful in prognosis and treatment. Rapid subtractive hybridization (RaSH) was used to identify and measure novel genes that appear to be differentially expressed, including KTN1, NEB, ROCK1, and ZAK using quantitative real-time polymerase chain reaction (qRT-PCR) and immunohistochemistry in the samples of GCTBs compared to normal bone tissue. Normal bone was used in the methodology RaSH for comparison with the GCTB in identification of differentially expressed genes. Functional annotation indicated that these genes are involved in cellular processes related to their tumor phenotype. The differential expression of KTN1, ROCK1, and ZAK was independently confirmed by qRT-PCR and immunohistochemistry. The expression of the KTN1 and ROCK1 genes were increased in samples by qRT-PCR and immunohistochemistry, and ZAK had reduced expression. Since ZAK have CpG islands in their promoter region and low expression in tumor tissue, their methylation pattern was analyzed by MSP-PCR. The genes identified KTN1, ROCK1, and ZAK may be responsible for loss of cellular homeostasis in GCTB since they are responsible for various functions related to tumorigenesis such as cell migration, cytoskeletal organization, apoptosis, and cell cycle control and thus may contribute at some stage in the process of formation and development of GCTB.

Associação entre a expressão de p16 e neoplasia intra-epitelial cervical / Association between p16 expression and cervical intraepithelial neoplasia

Introdução: Com aproximadamente 500 mil casos novos por ano no mundo, o câncer do colo do útero é o segundo tipo de câncer mais comum entre as mulheres, sendo responsável pelo óbito de, aproximadamente,230 mil mulheres por ano. Por meio de métodos imunohistoquímicos é possível evidenciar genes como o p16,envolvido diretamente na carcinogênese. Objetivo: Verificar a associação da expressão de proteína p16 em neoplasia intra-epitelial cervical (CIN) e prognóstico, relação com a persistência e graduação da lesão. Materiais e métodos: Neste estudo experimental, foram selecionadas 13 biópsias com CIN, de mulheres que, uma vez tratadas, permanecem livres de manifestação da doença há, no mínimo, 36 meses após o diagnóstico inicial. O grupo de estudo conta com 21 biópsias com CIN ou carcinoma invasor e que, mesmo tratadas, mantêm manifestação da CIN há pelo menos 4 anos. Utilizamos um biomarcador para a proteína p16, através de técnica imunohistoquímica e contamos as células marcadas. A seguir, verificamos o índice entre o númerode células imunocoradas dentre as células contadas em cada caso. Para o estudo estatístico, foi utilizado o teste Análise de Variância para Medidas repetidas. Resultados: Não há diferença na expressão deste marcador quanto à persistência da lesão (p=0,5024); ele marca de maneira diferenciada os casos de CIN III e de carcinoma espinocelular invasivo do colo do útero e, com menor intensidade, as CIN I (p=0,00000007), sendo p significante quando menor que 0,005. Verificamos que, quanto ao diagnóstico, o marcador para proteína p16 mostra-se altamente eficaz, marcando de maneira diferenciada os núcleos e citoplasmas de células dos casos de CIN III e, com menor intensidade, os de CIN I.

Introduction: With approximately 500,000 new cases worldwide each year, uterine cervical neoplasm is the second most common type of cancer among women. It is responsible for about 230,000 deaths yearly. By immunohistochemical methods, it is possible to reveal genes such as p16, which is directly involved incarcinogenesis. Objective: To study the association of p16 protein expression in cervical intraepithelial neoplasia (CIN) and prognosis compared with persistence and degree of injury. Materials and methods: In this experimental study, 13 biopsies with CIN were selected. All the biopsies were from women who have been treated and are now free of signs and symptoms of the disease for at least 36 months after the initial diagnosis. The study group consists of 21 biopsies from women with CIN or invasive carcinoma and that,even treated, remain with signs and symptoms of CIN for at least 4 years. We used a biomarker for protein p16 and an immunohistochemistry technique, and we counted the labelled cells as well. Then, we have checked the ratio of the amount of immuno stained cells to the cells counted in each case. We used the repeated measures analysis of variance to analyze data. Results: There is no difference in the expression of this marker as for the persistence of the lesion (p = 0.5024); it makes a mark differently in cases of CIN III and invasive spindle cell carcinoma of the uterine cervix, and to a lesser extent, in cases of the CIN I (p = 0.00000007). We considered p < 0.005 as statistically significant. We note that regarding the diagnosis, the marker for protein p16 appears to be highly effective, marking differently nuclei and cytoplasm of cells of all cases of CIN III and, to a lesser extent, those of CIN I.

Incidence of HPV infection in the uterine cervix in HIV positive pregnant women seen at the Hospital de Base in São José do Rio Preto, Brazil / Incidência de HPV em colo do útero de gestantes HIV positivas atendidas no Hospital de Base de São José do Rio Preto, SP

Objetivo: Avaliar a incidência de coinfecção HPV-HIV em gestantes e relacioná-la à quantidade de células CD4+ no sangue periférico; à expressão da proteína p16 no epitélio do colo uterino; e, finalmente, à presença de DNA de HPV nas biópsias cervicais. Métodos:Foram selecionadas gestantes da mesma idade em dois grupos com 70 pacientes em cada um: um HIV soropositivo (grupo de estudo) e outro soronegativo (controle I). Outro grupo (controle II) foi formado por 36 pacientes HIV negativas de idades variadas e com diagnóstico de neoplasia intraepitelial cervical, no período de 2000 a 2007. Foram analisadas colpocitologias e/ou biópsias dessas pacientes quanto à ocorrência de características morfológicas sugestivas de infecção pelo HPV; contagem de células CD4+ no sangue periférico; a expressão da proteína p16 no epitélio do colo uterino, elas foram comparadas com a mesma expressão em um grupo de 36 mulheres HIV soronegativas com neoplasia intraepitelial cervical; e PCR das amostras do grupo de estudo para confirmar a presença de DNA de HPV. Resultados: As 70 pacientes gestantes HIV positivas, tinham idade entre 15 e 45 anos, com média etária de 28 anos (mediana = 28,2 anos). Delas, 22 (31,4%) apresentaram alterações morfológicas compatíveis com infecção cervical pelo HPV; dessas pacientes, 16 apresentaram menos de 500 células CD4 positivas (p=0,03). No grupo HIV soronegativo, uma paciente (1,4%) apresentou lesão intraepitelial cervical. Dez biópsias cervicais do grupo de estudo apresentaram DNA de HPV pela PCR. O Grupo Controle II mostrou resultados semelhantes ao de estudo. Conclusões: A incidência de coinfecção HPV-HIVfoi relevante em relação ao grupo de gestantes HIV negativas. Nas gestantes HIV positivas, a incidência de HPV mostrou-se diretamente relacionada com baixa imunidade. A expressão da proteína p16 mostrou índices de marcação maiores nos casos com diagnóstico de lesões do colo do útero mais graves (NIC 2 e 3). A presença de DNA de HPV foi confirmada em lesões cervicais de gestantes HIV positivaspor meiode PCR. O presente estudo pôde constatar que existe necessidade de avaliações ginecológicas e colpocitológicas periódicas no pré-natal de pacientes HIV positivas, visando o diagnóstico precoce da infecção pelo HPV.

Controle da qualidade em colpocitologia: visão rápida com campo marcado / Cervical cytology quality control: rapid pre-screening with marked field

INTRODUÇÃO: No Departamento de Patologia e Medicina Legal da Faculdade de Medicina de São José do Rio Preto (FAMERP), adotou-se um sistema de gestão da qualidade (SGQ) segundo as normas brasileiras preconizadas pela International Organization for Standardization (NBR ISO) 9001: 2000. Esse procedimento foi adotado para assegurar a confiabilidade técnica dos exames realizados, em que a qualidade é considerada como julgamento do cliente, ou seja, solução àss suas expectativas. Neste estudo, focamos a qualidade dos exames colpocitológicos realizados pelo Departamento. Embora a evolução de lesões do colo do útero para carcinoma invasor seja lenta, o êxito do tratamento depende do seu diagnóstico precoce. Assim, entre as possíveis falhas do exame preventivo, diagnóstico falso-negativo é a que mais agravo causa, pois posterga o início do tratamento. OBJETIVO: Testar tr�s métodos diferentes, procurando aquele com menor número de diagnóticos falso-negativos. MATERIAL E MÉTODO: Com a mesma equipe examinadora e com os mesmos critérios de diagnóstico, testamos o método 1 (M1), preconizado pelo Ministério da Saúde (MS), que faz revisão aleatória de 10 por cento dos casos negativos, em 3.500 colpocitologias; o método 2 (M2): revisão rápida (RR), proposto na literatura, que consiste de revisão rápida de 100 por cento dos casos, após o escrutínio, em 7.373 casos; e o m�todo 3 (M3): visão rápida (VR) com campo marcado, delineado pela pesquisadora, propõe o exame rápido pré-escrutínio de todos os casos, marcando-se um campo em que existam células alteradas, quando visualizadas, em 8.096 citologias ginecológicas. RESULTADOS: Foram encontradas as seguintes porcentagens de diagnósticos falso-negativos: M1: 2,4 por cento; M2: 1,7 por cento e M3: 1,2 por cento. CONCLUSÃO: O méodo M3 foi o melhor, apresentando menor quantidade de diagnósticos falso-negativos.

INTRODUCTION: The Pathology and Forensic Medicine Department of the Faculdade de Medicina de S�o Jos� do Rio Preto, SP had decided to develop a Quality Control System following the NBR ISO 9001: 2000 guidelines. Some procedures were adopted to verify technical reliability for the exams performed, where quality is defined as the clients' judgement, i.e., the solution for the clients' requests and expectations. In this study we have focused on the quality of the cervical cytology performed in the Department. Although the development of cervical lesions through cervical carcinoma takes place slowly, the treatment will have a better chance of success in case of an early diagnosis. Considering all the possible diagnostic mistakes, the false-negative exam is the most dangerous, as it delays the beginning of the treatment. OBJECTIVE: To test three different methods looking for the one with the smaller number of false-negative results. MATERIAL AND METHODS: For the same team of technicians, and with the same diagnostic criteria the three different techniques were tested. Methodology 1 (M1): in accordance to the Brazilian Ministry of National Health, with a random review of 10 percent of the false-negative results, in the 3,500 cytologies; Methodology 2 (M2): rapid review (RR), as suggested in literature, consists of a rapid review of 100 percent of the smears, after their routine evaluation, and was done in 7,373 cytologies; Methodology 3 (M3): rapid prescreening (RP) with marked field, was proposed by the researcher and consists of a rapid screening of all cases, pointing out a field were there are abnormal cells, and after that the normal screening, in 8,096 cytologies. RESULTS: The three techniques tested showed the following false-negative rates: M1: 2.4 percent; M2: 1.7 percent and M3: 1.2 percent. CONCLUSION: The M3 was the best methodology, presenting less false-negative diagnosis.

The prevalence of human papillomavirus in the oropharynx in healthy individuals in a Brazilian population.

Oncogenic human papillomavirus (HPV), a causative agent of uterine cervical cancer, has also been detected in head and neck squamous cell cancers, especially in squamous cell carcinomas of the tonsils. However, the true HPV prevalence in normal and neoplasic oropharyngeal mucosa remains uncertain. To determine the prevalence of HPV DNA in normal oropharyngeal mucosa of cancer-free individuals, a study was carried out on 50 Brazilian subjects. PCR was performed to identify HPV DNA in samples from four sites in the oropharynx (tonsils, soft palate, base of the tongue, and back wall of the pharynx). For amplification of the HPV DNA, MY09/11 consensus primers were used, and specific genotypes were identified by dot-blot hybridization or cloning and sequencing. HPV DNA was present in 14.0% of the individuals, and the identified genotypes were 16, 18, 52, and 61. All these types are considered high-risk (HR) HPV. The tonsils and the soft palate were the sites with the highest HPV prevalence. This study shows the prevalence of HR HPV in the oropharynx of normal individuals. However, the prevalence of HPV is still unclear, and if HPV infection in a healthy it is not known individual predisposes to HPV-associated disease such as oropharyngeal cancer. Thus, it is important to assess the prevalence of HPV in cancer-free individuals, in order to compare it with the HPV prevalence in oropharyngeal carcinomas and to attempt to determine the true role of HPV in the development of head and neck squamous cell cancers.

Matriz extra-celular: fibronectina / Extracellular matrix: fibronectine

A fibronectina é uma glicoproteína presente no plasma sangüíneo e nos tecidos. Tem propriedade de ligar-se a si mesma e a várias outras substâncias diferentes, tais como a fibrina, a heparina, bactérias, colágeno, fibroblastos, e outras células. Por esta propriedade ligante, desempenha funções fisiológicas múltiplas e tem participação importante em diversos processos patológicos, tanto em sua forma intacta como pela presença de seus isômeros e fragmentos. Fisiologicamente, a fibronectina é responsável pela orientação da migração celular na embriogênese e participa na homeostase e a coagulação sangüínea e na condrogênese. A fibronectina está envolvida em processos osteoartríticos, na produção de anomalias nos membros e em condições patológicas que afetam a mucosa oral humana

Idade do paciente, hiperplasia nodular e adenocarcinoma de próstata: estudo estatístico / Patient age, nodular hyperplasia and prostate adenocarcinoma: a statistical study

A próstata humama é constituída por estroma e ácinos, distribuídos em dois lobos laterais e um central e pode apresentar três importantes lesoes: inflamatória, hiperplasia nodular e câncer. No presente trabalho investigamos a faixa etária em que ocorrem a hiperplasia nodular e o câncer. Verificamos, estatisticamente, que ambas as patologias ocorrem na mesma faixa etária, com maior incidência de ambas entre 65 e 69 anos.