Evaluation of Telomerase (hTert), Ki67 and p16ink4a expressions in low and high-grade cervical intraepithelial lesions / Avaliação da expressão de Telomerase (hTert), Ki67 e p16 ink4a em lesões intraepiteliais cervicais de baixo e alto graus

ABSTRACT Objective : to study the association between the histological grading of cervical intraepithelial neoplasia (CIN I, CIN II and CIN III) and the immunohistochemical expression for p16ink4a, hTert and Ki67, as well as to evaluate the relationship of these markers with the risk of recurrence after surgical treatment. Methods : we studied a historical cohort of 94 women with intraepithelial lesions CIN I (low grade), CIN II and CIN III (high grades) submitted to conization or electrosurgical excision of the transformation zone. We evaluated all surgical specimens for immunohistochemical expression of p16ink4a, hTert and Ki67. Results : the mean age was 38.2 years; p16ink4a was absent in most CIN I cases. In patients with CIN II or I/II (association of low and high-grade lesions), we observed p16ink4a ≤10%. In patients with CIN III, we found a higher expression frequency of p16ink4a >50%. In CIN I, the majority had Ki67≤10% and low frequency of Ki67>50%. In the CIN III category, there were fewer patients with Ki67≤10%, and Ki67 was absent in most patients of CIN II and III groups. There was no association between hTert expression and histologic grade. There were no statistically significant differences between the expression of the markers in patients with and without recurrence. Conclusion : there was a statistically significant association of p16ink4a and Ki67 with histological grade. The markers' expression, as for disease recurrence, was not statistically significant in the period evaluated.

RESUMO Objetivo: estudar a associação entre a graduação histológica das neoplasias intraepiteliais cervicais (NIC I, NIC II e NIC III) e a expressão imuno-histoquímica para p16ink4a, hTert e Ki67, assim como, avaliar a relação destes marcadores com o risco de recorrência após tratamento cirúrgico. Métodos: estudo de coorte histórica de 94 mulheres portadoras de lesões intraepiteliais NIC I (baixo grau), NIC II e NIC III (altos graus), submetidas à conização ou à excisão eletrocirúrgica da zona de transformação. Todas as peças cirúrgicas foram avaliadas quanto à expressão imuno-histoquímica para p16ink4a, hTert e Ki67. Resultados: a média de idade das pacientes foi 38,2 anos. Nas pacientes NIC I, a p16ink4a estava ausente na maioria dos casos; nas pacientes NIC II ou I/II (associação de lesões de baixo e alto graus), observou-se frequência de p16ink4a≤10%. Nas pacientes NIC III, observou-se maior frequência de expressão de p16ink4a>50%. Na categoria NIC I, a maioria apresentava Ki67≤10% e baixa frequência de Ki67>50%. Na categoria NIC III houve menor número de pacientes com Ki67≤10%, sendo que a maior parte das pacientes tinha Ki67 ausente nos grupos NIC II e III. Não houve associação entre a expressão do marcador imuno-histoquímico hTert e a graduação histológica. Não houve diferenças estatisticamente significativas entre as expressões dos marcadores em pacientes com e sem recorrência. Conclusão: houve associação estatisticamente significativa apenas de p16ink4a e Ki67 com a graduação histológica. A expressão dos marcadores em relação à recorrência da doença não foi estatisticamente significativa no período avaliado.

Overexpression of ezrin and galectin-3 as predictors of poor prognosis of cervical cancer

The aim of this study was to explore the correlation of ezrin and galectin-3 expressions with prognosis in cervical cancer. The immunohistochemical method was applied to detect ezrin and galectin-3 expressions in normal cervix tissues (n=30), cervicitis tissues (n=28), cervical intraepithelial neoplasia (CIN) tissues (classified as I-III, n=89), and cervical carcinoma tissues (n=84). Follow-up was conducted for 5 to 78 months to analyze the correlation of protein expressions with prognosis. Ezrin and galectin-3 expressions in cervical cancer were significantly higher than in normal cervix, cervicitis and CIN (all P<0.05), and expressions in CIN were significantly higher than in normal cervix and cervicitis (both P<0.05). The expressions of ezrin and galectin-3 were both related with histological grade, deep myometrial invasion and lymph node metastasis (all P<0.05). Spearman analysis showed that ezrin expression was positively correlated with galectin-3 expression in cervical cancer (r=0.355, P<0.05). The survival rate of patients with high expressions of ezrin and galectin-3 was significantly lower than those with low expressions of proteins (both P<0.05). The expressions of ezrin and galectin-3, histological grade, depth of stromal invasion, and lymph node metastasis are risk factors affecting the survival rate of patients with cervical cancer. The expressions of ezrin and galectin-3 were correlated with the development of cervical cancer, and overexpressions of those proteins were indicative of poor prognosis in patients with cervical cancer.

Greater expression of the human leukocyte antigen-G (HLA-G) and interleukin-17 (IL-17) in cervical intraepithelial neoplasia: analytical cross-sectional study / Aumento da expressão do antígeno leucocitário humano-G (HLA-G) e interleucina-17 (IL-17) em neoplasia intraepitelial cervical: estudo transversal analítico

CONTEXT AND OBJECTIVE:Impaired local cell immunity seems to contribute towards the pathogenesis and progression of cervical intraepithelial neoplasia (CIN), but the underlying molecular mechanisms promoting its progression remain unclear. Identification of new molecular markers for prognosis and diagnosis of early-stage CIN may aid in decreasing the numbers of CIN cases. Several novel immunoregulatory molecules have been discovered over the past few years, including the human leukocyte antigen G (HLA-G), which through interaction with its receptors exerts important tolerogenic functions. Several lines of evidence suggest that T-helper interleukin-17 (IL-17)-producing cells (Th17 cells) may play a role in antitumor immunity. However, recent reports have implicated Th17 cells and their cytokines in both pro and anti-tumorigenic processes. The aim of the study was to evaluate the roles of HLA-G and Th17 in the immunopathogenesis of CIN I.DESIGN AND SETTING:Analytical cross-sectional study with a control group using 58 cervical specimens from the files of a public university hospital providing tertiary-level care.METHODS:We examined HLA-G and IL-17 expression in the cervical microenvironment by means of immunohistochemistry, and correlated these findings with clinical and pathological features.RESULTS:There was a greater tendency towards HLA-G and IL-17 expression in specimens that showed CIN I, thus suggesting that these molecules have a contribution towards cervical progression.CONCLUSION:These findings suggest that HLA-G and IL-17 expression may be an early marker for assessing the progression of cervical lesions.

CONTEXTO E OBJETIVO:A deficiência na imunidade celular localizada parece contribuir para a patogênese e progressão das neoplasias intraepiteliais cervicais (NIC), no entanto, ainda não está totalmente esclarecido o mecanismo molecular fundamental nesse processo de progressão. A identificação de novos marcadores moleculares de prognóstico e diagnóstico das NIC em estágios precoces pode ajudar a diminuir a quantidade de casos de NIC. Várias novas moléculas com função imunorregulatória foram descobertas nos últimos anos, inclusive o antígeno leucocitário humano G (HLA-G), que, através de interação com os receptores, tem importantes funções tolerogênicas. Diversas linhas de evidência sugerem que as células T-ajudantes produtoras de interleucina-17 (IL-17, células Th17), podem desempenhar um papel na imunidade antitumoral. Porém, recentes relatos implicaram as células Th17 e suas citocinas tanto em processos pro- quanto anti-tumorigênicos. O objetivo do estudo foi avaliar o papel do HLA-G e Th17 na imunopatogênese das NIC I.TIPO DE ESTUDO E LOCAL:Estudo transversal analítico com grupo controle em 58 espécimes cervicais dos arquivos de um hospital universitário público com assistência prestada no nível terciário.MÉTODOS:Avaliamos a expressão de HLA-G e IL-17 por imunoistoquímica no microambiente cervical, associando esses achados com as características clínico-patológicas.RESULTADOS:Houve tendência aumentada da expressão de HLA-G e IL-17 em espécimes que apresentaram NIC I, sugerindo que essas moléculas têm contribuição na progressão cervical.CONCLUSÃO:Estes resultados sugerem que a expressão do HLA-G e da IL-17 pode ser um marcador precoce para avaliar a progressão das lesões cervicais.

Tinción dual inmunocitoquímica de p16INKa/Ki-67 para la detección de lesiones del cuello uterino asociadas a infección por el virus del papiloma humano / p16INK4a/Ki-67 immunocytochemical dual staining for detection of cervical lesions associated to papillomavirus infection

Este estudio se llevó a cabo para examinar el patrón de inmunoexpresión simultánea de p16INK4a/Ki-67y establecer su posible utilidad clínica para la detección precoz del cáncer de cuello uterino. Las muestras celulares de cuello uterino fueron seleccionadas de la pesquisa de rutina de cáncer cervical. La detección inmunocitoquímica de p16INK4a/Ki-67se realizó con el kit de trabajo CINtec® Plus. Todos los casos tenían una prueba de virus papiloma humano (VPH). Ciento quince muestras citológicas fueron incluidas: 11(9,6%) fueron negativas para lesión intraepitelial o malignidad (NILM), 32(27,8%) presentaron células escamosas con atipias de significado indeterminado (ASC-US), 62(53,9%) mostraron lesión intraepitelial escamosa de bajo grado (LSIL) y 10(8,7%) lesión intraepitelial escamosa de alto grado (HSIL). La prevalencia general de infección por VPH fue de 81,7% (94/115). En 42 casos (45,0%) se identificaron los siguientes genotipos específicos de VPH: VPH16 (26,2%), VPH51 (21,4%), VPH52 (14,3%) y el genotipo VPH66 (7,1%). De 115 muestras celulares, 42(36,5%) fueron positivas para la tinción dual de p16INK4a/Ki-67, siendo ésta muy frecuente en las muestras citológicas con HSIL (70,0%), disminuyendo en las LSIL (44,0%) y existiendo inmunopositividad en una minoría de ASC-US (25,0%). Ningún caso NILM mostró inmunopositividad para p1(6INK4a)/Ki-67 (p<0,001). En 40/115 casos (34,8%) hubo positividad tanto para infección por VPH oncogénico como para la tinción dual p16INK4a/Ki-67, incluyendo 6/32 (18,8%) con ASC-US, 26/62 (42,0%) con LSIL and 8/10 (80,0%) con HSIL. Esta metodología podría ser utilizada para detectar lesiones en cuello uterino que aún no han sido diagnosticadas o han pasado inadvertidas.

We aimed to explore the expression pattern of p16INK4a/Ki-67 immunocytochemical dual-staining and to establish the potential clinical utility for early detection of cervical lesions. Liquid-based cytologies of cervical specimens of cervical cancer screening were processed for p16INK4a/Ki-67 immunocytochemical dual-staining using the CINtec® Plus Kit. HPV testing was performed with the INNO-LiPA HPV genotyping Extra Reverse Hybridization Line Probe Assay kit. One hundred and fifteen cervical cytologies were analyzed with the following results: 11(9.6%) were negative for intraepithelial lesions or malignancy (NILM); 32(27.8%) presented atypical squamous cells of undetermined significance (ASC-US); 62(53.9%) exhibited low grade squamous intraepithelial lesions (LSIL) and 10(8.7%) showed high grade squamous intraepithelial lesions (HSIL). No cases of cervical cancer were detected. The overall prevalence of DNA HPV detection was 81.7% (94/115). The following specific HPV genotypes were identified in 42 (45.0%) cases: HPV16 (26.2%), HPV51 (21.4%), HPV52 (14.3%) and HPV66 (7.1%). Viral sequences of an unknown single HPV were detected in 23.8%of the cases. A total of 42/115 (36.5%) were p16INK4a/Ki-67 dual-staining-positive, being more frequent in HSIL (70.0%), decreasing in LSIL (44.0%), detected in a minority of ASC-US (25.0%) and negative in NILM cases (p<0.001). 40/115 cases (34.8%) were positive for both oncogenic HPV and p16INK4a/Ki-67 dual-staining, including 6/32 (18.8%) ASC-US, 26/62 (42.0%) LSIL and 8/10 (80.0%) HSIL, which represent a strong association between positivity for HPV, p16INK4a/Ki-67 staining and severe cytological abnormalities (p<0.001). This methodology could be used to detect unnoticed cervical lesions.

Immunohistochemical expression of MIB-1 and PCNA in precancerous and cancerous lesions of uterine cervix.

BACKGROUND AND OBJECTIVE: The present study was done to analyze the immunoexpression of diagnostic markers (MIB-1: molecular immunology borstel and PCNA: proliferating cell nuclear antigen) in grading cervical intraepithelial lesion (CIN) and squamous cell carcinoma (SCC) in cervix. SETTING AND DESIGN: Total 150 cervical biopsies were divided into four groups respectively; Group I-Normal (n = 32), Group II- CIN (n = 60), Group III- SCC (n = 44), Group IV- CA cervix (n = 14) respectively. MATERIALS AND METHODS: These biopsies were stained with monoclonal antibodies by streptavidin-- biotin method. Mean labeling index was calculated and grading was performed using the I--III scoring system. STATISTICAL ANALYSIS: Findings were correlated with age and menopausal status. Statistical analysis was done by using student sample‘t’ test and analysis of variance (ANOVA) by SPSS 10 package. RESULTS: MIB-1 immunostaining was positive in 112/150 (74.6%) cases and PCNA in 118 /150 (78.6%) cases. Labeling indices showed linear progression from normal to CIN to SCC to cancer lesion. Few cases of low-grade CIN lesion had high proliferative index. A significant positive correlation was found between age and PCNA and MIB-1 values (P < 0.05) when comparison was made for all the cases. CONCLUSION: These markers may be useful in identifying low-grade CIN lesion with high proliferative index. These cases should be kept for follow up studies so that proper intervention can be taken at an early stage. This method is simple and cost effective and can easily be done in formaline-fixed paraffin embedded tissues in a clinical laboratory for grading CIN and SCC lesions in cervix.

An evaluation of p16INK4a expression in cervical intraepithelial neoplasia specimens, including women with HIV-1

Although several studies have evaluated the role of p16INK4a as a diagnostic marker of cervical intraepithelial neoplasia (CIN) and its association with disease progression, studies regarding the role of p16INK4a in human immunodeficiency virus (HIV)-infected patients remain scarce. The present study was designed to determine the potential utility of p16INK4a as a diagnostic marker for CIN and invasive cervical cancer in HIV-positive and negative cervical specimens. An immunohistochemical analysis of p16INK4a was performed in 326 cervical tissue microarray specimens. Performance indicators were calculated and compared using receiving operating characteristics curve (ROC)/area under the curve. In HIV-1-negative women, the percentage of cells that was positive for p16INK4a expression was significantly correlated with the severity of CIN (p < 0.0001). A ROC curve with a cut-off value of 55.28% resulted in a sensitivity of 89%, a specificity of 81%, a positive predictive value of 91% and a negative predictive value of 78%. HIV-seropositive women exhibited decreased expression of p16INK4a in CIN2-3 specimens compared with HIV-negative specimens (p = 0.031). The ROC data underscore the potential utility of p16INK4a under defined conditions as a diagnostic marker for CIN 2-3 staging and invasive cervical cancer. HIV-1 infection, however, is associated with relatively reduced p16INK4a expression in CIN 2-3.

p16INK4a como predictor de evolución clínica en pacientes con neoplasia intraepitelial cervical de bajo grado (NIC-I): serie de casos / p16INK4a as a predictor of clinical outcome in patients with low grade-cervical intraepithelial neoplasia

El diagnóstico morfológico de neoplasia intraepitelial cervical de bajo grado (NIC-I) no permite determinar su real riesgo de progresión a cáncer de cérvix, llevando a seguimiento estricto a un alto número de pacientes cuyas lesiones, en su mayoría, regresarán espontáneamente. La detección de p16INK4A se ha propuesto como biomarcador que permite diferenciar entre infección productiva-autolimitada por virus del papiloma humano (VPH) y transformación celular inducida por cepas oncogénicas del VPH, sin embargo, es necesario aportar evidencias de su capacidad predictiva. Se presenta una serie de 14 casos de NIC-I con detección inmunohistoquímica de p16INK4a en biopsia de cérvix (8 positivos y 6 negativos), cuya revisión retrospectiva de historial clínico de más de 12 meses de seguimiento cito-histopatológico, permite describir regresión espontánea en todos los casos p16INK4a negativo y en 3 casos p16INK4a positivo (37,5 por ciento). Los hallazgos en esta serie coinciden con lo descrito en estudios previos en los que se ha demostrado el valor predictivo negativo del marcador para descartar riesgo de progresión-persistencia y evitar seguimientos y tratamientos innecesarios. Además señalan debilidades a evaluar respecto al valor predictivo positivo de la prueba, al no discriminar algunos factores independientes de la transformación celular, determinantes en la progresión-persistencia de la NIC-I, como lo son variables genéticas y/o inmunológicas del huésped.

The morphological diagnosis of low grade-cervical intraepithelial neoplasia (CIN-1) can not determine its true risk of progression to cervical cancer, leading to strict adherence to a high number of patients whose lesions, most will return spontaneously. The detection of p16INK4a has been proposed as biomarker to differentiate between productive infection self-limiting by human papillomavirus (HPV) and cell transformation induced by oncogenic strains of HPV, however, it is necessary provide evidence of its predictive capacity. We present a series of 14 cases of CIN-1 with immunohistochemical detection of p16INK4a in cervical biopsy (8 positive and 6 negative), whose retrospective review of medical records of more than 12 months of cyto-histopathological follow, can describe spontaneous regression in all cases p16INK4a negative and in 3 cases p16INK4a positive (37.5 percent). The findings in this series of cases coincide with that described in previous studies that has been demonstrated the negative predictive value of the marker to exclude risk of progression-persistence and avoid follow-ups and unnecessary treatments. Additionally, these indicates weaknesses to evaluate about the positive predictive value of the test, by not discriminating factors independent of cellular transformation, determining the progression-persistent CIN-1, such as genetic and / or immunologic variables of the host.

Detección de virus papiloma humano (HPV) a partir de muestras celulares de cuello uterino en base líquida: Correlación con la inmunorreactividad de la proteína p16INK4a / Detection of human papilloma virus (HPV) in liquid-based cervical samples: Correlation with protein p16INK4a expression

La citología del cuello uterino en base líquida mejora la calidad de la muestra y el material residual podría ser utilizado para realizar pruebas complementarias, como la detección del virus papiloma humano (HPV) y estudio inmunocitoquímico de biomarcadores. El propósito de este estudio fue correlacionar la presencia de HPV y la inmunoexpresión de p16INK4a en las muestras citológicas en base líquida para examinar la utilidad de estas nuevas herramientas en la detección de cáncer de cuello uterino. Las pacientes incluidas (n=67) presentaban una citología anormal o patología cervical previa. La detección y genotipificación de HPV se realizó con PCR-SPF10/LiPA (INNOLiPA Extra Amp) y para la inmunodetección de p16INK4a se utilizó el anticuerpo clon E6H4. La citología convencional proporcionó los mismos hallazgos citológicos que la citología en base líquida. La prevalencia general del HPV fue de 43,3% (29/67). El HPV16 fue el tipo viral mas frecuente (31,03%) y el 48,3% de los casos presentó infección múltiple. En el 35,8% de las muestras cervicales se detectó inmunoexpresión de la p16INK4a y ésta fue significativamente (p<0,020) asociada a la presencia de HPV. Estos resultados apoyan la evidencia que la implementación de nuevas tecnologías en la rutina diaria del laboratorio contribuye significativamente en la detección precoz del cáncer de cuello uterino y en el aporte de datos importantes para facilitar en el manejo clínico adecuado de la paciente. La detección de HPV combinada con la p16INK4a podría ser utilizado en la evaluación de pacientes con mayor riesgo a desarrollar lesiones cervicales significativas.

The liquid-based cervical cytology improves the quality of the sample and the residual sample could be used efficiently to carry out complementary tests, such as the detection of HPV DNA and the immunocytochemical biomarkers study. The purpose of this study was to correlate the presence of HPV and immunoexpression of p16INK4a in liquid-based cervical samples to examine the utility of these new tools in the detection of cervical cancer. The included patients (n=67) presented an abnormal cytology or previous cervical pathology. The HPV detection and genotyping were carried out with PCR-SPF10/LiPA (INNOLiPA Extra Amp) and for p16INK4a immunodetection was used antibody clone E6H4. The conventional cytology provided the same cytologic interpretations that those of liquid-based cytology. The overall HPV prevalence was 43.3% (29/67). HPV16 was the most frequent viral type (31.03%) and 48.3% of the cases were infected with multiple HPV types. p16INK4a immunoexpression was observed in 35.8% of liquid-based cytological samples and this was significantly (p < 0.020) associated to the HPV presence. These results support the evidence that the implementation of new technologies in the daily routine of the laboratory, contribute significantly in the early detection of cervical cancer and provide important data to help in the patient’s efficient management. The combined use of HPV detection and p16INK4a expression could be used for evaluation of patients with more risk to develop significant cervical lesions.

p16INK4 expression in precursor lesions of squamous cell cervical cancer related to the presence of HPV-DNA

The purpose of the present study was to identify the expression of p16INK4 in cervical cancer precursor lesions by immunohistochemistry and to correlate it with lesion grade and presence of human papillomavirus (HPV) infection. Cervical specimens from 144 women seen consecutively at the gynecology outpatient clinic of our institution from December 2003 to May 2005 were analyzed by cytopathology, histopathology, polymerase chain reaction for HPV-DNA, and p16INK4 immunostaining. Histologically normal biopsies, HPV-DNA negative by polymerase chain reaction, were used as control. HPV-DNA prevalence, including the control group, was 68.1 percent and the prevalence of p16INK4 expression was 55.0 percent. The percentage of cells stained by p16INK4 ranged from 10 to 100 percent, both in the group consisting of cervical intraepithelial neoplasia (CIN)1/HPV specimens and in the group of CIN2/CIN3 specimens with P value of 0.0001. p16INK4 expression was 48.3 percent in the CIN1/HPV group, as opposed to 94.3 percent in the CIN2/CIN3 group (P = 0.001), showing a statistically significant difference between the two groups. The quantitative method used here is simple and less subjective than the different semiquantitative methods described in the literature. In view of the different definitions of a p16INK4-positive case, it is almost impossible to compare the findings reported by different investigators. This study confirms the association between p16INK4 and CIN2 and CIN3 lesions. Moreover, it shows that some low grade lesions expressed high levels of this protein. This may indicate that such low grade lesions may be predisposed to progress to high grade lesions. This means that p16INK4 may be a strong marker for "neoplastic lesions" induced by HPV and not just an infection marker.

Associação entre a carga viral de HPV de alto risco, expressão de p16INK4a e lesões intra-epiteliais escamosas do colo uterino / Association between high risk HPV viral load, p16ink4a expression and intra-epithelial cervical lesions

OBJETIVO: Determinar a carga viral do HPV de alto risco em lesões intra-epiteliais escamosas do colo uterino e sua associação com a expressão da proteína p16INK4a. MÉTODOS: Foram analisadas 109 amostras de biópsias de colo uterino (57 de tecido normal, 26 de lesões intra-epiteliais escamosas de baixo grau LSIL, e 26 de lesões intra-epiteliais escamosas de alto grau - HSIL). A determinação da presença do HPV de alto risco e sua carga viral foi feita por biologia molecular, com captura de híbridos, por meio de coleta de células de colo uterino previa à biópsia. O material histológico foi preparado e testado por imunohistoquímica, utilizando p16INK4a kit (clone E6H4). RESULTADOS: Encontrou-se 57,8 por cento de positividade para HPV de alto risco nos 109 casos estudados, sendo 29,8 por cento nos casos normais, 80,8 por cento nos casos com LSIL e 96,1 por cento nos casos de HSIL. A expressão da proteína p16INK4a ocorreu em 23,8 por cento do total casos com SIL (15,4 por cento dos casos com LSIL e 84,6 por cento dos casos com HSIL), porém não houve qualquer caso positivo nos tecidos sem lesão intra-epitelial. A carga viral foi significativamente superior para os casos positivos para p16INK4a, sendo a média de 669,9 RLU/PCB (9,47 a 2814,9), em relação aos casos negativos em que a média foi de 253,94 RLU/PCB (1.07 a 1882,21) (p<0.05). Mas, analisando apenas as HSIL, a diferença não foi significativa. CONCLUSÃO: No presente estudo, embora a carga viral de HPV de alto risco tenha demonstrado uma diferença significativa entre casos positivos e negativos para p16, nos casos de HSIL isto não foi confirmado. Novos estudos, com maior número de casos, são necessários para conclusões mais consistentes.

OBJECTIVE: To determine the (HR-HPV) high risk HPV viral load in squamous intra-epithelial lesions and association with p16INK4a expression. METHODS: A series of 109 cervical biopsies were studied (57 normal tissue, 26 low grade squamous intra-epithelial lesions [LSIL] and 26 high grade squamous intra-epithelial lesions [HSIL]). Detection of high risk HPV and viral load in cervical cells was made by molecular biology using hybrid capture 2nd generation collected before the biopsy. The p16INK4a was identified by immunohistochemistry using the p16INK4a kit (clone E6H4). RESULTS: High risk HPV was positive in 57.8 percent of all cases (29.8 percent in normal tissue, 80.8 percent in LSIL and 96.1 percent in HSIL). Protein p16INK4a was expressed in 23.8 percent of squamous intra-epithelial lesions (15.4 percent in LSIL and 84.6 percent in HSIL). In normal tissue all cases were negative to p16INK4a. The viral load was higher in p16 positive cases than in negative cases (positive p16INK4a mean of 669.9 RLU/PCB [9.47 - 2814.9] and negative p16INK4a mean of 253.94 RLU/PCB [1.07 1882.21] (p<0.05). However when studying just the HSIL cases differences were not significant. CONCLUSION: In this study although the HR-HPV viral load had shown a significant difference between p16 positive and negative cases, in HSIL cases this finding was not confirmed. New studies with a larger number of cases are necessary for consistent conclusions.

Expression of Pin1 and Ki67 in cervical cancer and their significance / 华中科技大学学报（医学）（英德文版）

In order to investigate the expression levels of Pin1 mRNA and protein in cervical cancer and its association with Ki67 and their clinical significance, amplification of Pin1 gene was examined by RT-PCR, and the expression of both Pin1 and Ki67 protein was detected by immunohistochemistry in cervical cancer tissues. It was shown that the expression levels of Pin1 were higher in cervical cancer than in normal cervical tissues (P 0.05). The expression of Pin1 was significantly higher in adenocarcinoma than in squamous carcinoma of the uterine cervix (P < 0.05). In cervical cancer, the overexpression of Pin1 was positively correlated with that of Ki67 (P < 0.05). These results suggested that the overexpression of Pin1 was closely related with cancer cell proliferation or progression of cervical cancer and contributed to oncogenesis. Pin1 may serve as a potential marker for cervical cancer diagnosis.

Antioxidant Vitamins and Lipid Peroxidation in Patients with Cervical Intraepithelial Neoplasia

The purpose of this study was to investigate the implications of dietary intake and the level of plasma antioxidant, lipid peroxidation, and antioxidant capacity in Korean women with cervical intraepithelial neoplasia (CIN). From October 2002 to March 2003, 58 patients diagnosed with CIN (confirmed with colposcopy directed biopsy) and 86 patients without any cervical disease as control group were enrolled in the study at the Department of Gynecology cancer center at Samsung Cheil Hospital. The intake of antioxidant vitamins in both groups exceeded the amount recommended by the Korea RDA, 7th edition. The plasma concentration of Vitamin C was significantly lower in the CIN group (0.36 mg/dL) than in the control group (0.48 mg/dL) (p<0.05). The two groups showed similar plasma concentrations of beta-carotene, alpha-tocopherol, and retinol. The average concentration of malondialdehydes in the CIN group, 7.23 mmol/mL, was significantly higher than in the control group, 5.18 mmol/mL (p<0.01). The total radical trapping antioxidant potential concentration of plasma was significantly higher in the CIN group (1.15 mM) than in the control group (1.25 mM) (p<0.05). These results suggest that there is a possible correlation between cervical intraepithelial neoplastic processes and changes in the plasma antioxidative system.

Detection of mutagenicity in cervico-vaginal secretions--a plausible risk factor for cervical cancer.

The cervico-vaginal secretions from 51 women with various grades of dysplastic lesions of uterine cervix were assessed for mutagenic potential by Ames test using histidine deficient mutant strain of Salmonella typhimurium TA-98: with S-9 mix. Twenty three per cent of samples from women with cervical dysplasia were found significantly positive (P less than 0.001) for mutagenic activity compared to 3% positive from control. The frequency of mutagenic secretions detected were almost uniform, irrespective of the severity of cervical lesions. None of cervico-vaginal secretions, positive for mutagenicity could revert the tester strain when tested in absence of S-9 mix (liver microsomal enzymes). This indicates that mutagens in cervico-vaginal secretions are effective only when activated enzymatically.