Immunosurveillance profile of oral squamous cell carcinoma and oral epithelial dysplasia through dendritic and T-cell analysis.

Oral squamous cell carcinomas (OSCCs) can arise from potentially malignant disorders, such as leukoplakia. The immune system plays an important role recognizing tumour precursor cells. However, due to immuno-editing mechanisms cancer cells are able to escape immune system surveillance. OBJECTIVE: To evaluate the profile of dendritic (Langerhans and plasmacytoid) and T cells in OSCC and oral epithelial dysplasia (OED) and correlate these findings with clinical data. MATERIALS AND METHODS: Fifty cases of OSCC and 48 of OED were immunostained for CD1a and CD83 dendritic Langerhans cells (DLC), CD303 plasmacytoid dendritic cells (pDC) and CD8 followed by quantitative analysis. RESULTS: Analysis revealed a significant decrease in the number of mature CD83 DLC in OSCC compared with OED. CD303 positivity was significantly increased in the OSCC group when compared to OED. CD8-positive lymphocytes were significantly decreased in OSCC compared with OED lesions. No statistical correlation was found with clinical data. CONCLUSION: The number of mature dendritic cells (DC) was decreased in OSCC compared with OED lesions suggesting that either these cells might have migrated to lymph nodes to present the tumour antigens and activate the immune system or cytokines secreted by the tumour microenvironment are inhibiting the adequate maturation of DLC. The numbers of pDC were significantly increased in the OSCC group compared with the OED group. This suggests they may play an important role in the defence against tumours although it is not clear whether this is promoting or inhibiting malignant progression.

Respuesta inmune celular en pacientes con lesiones benignas y malignas del cuello uterino / Cellular immune response in patients presenting with benign and malignant lesions of cervix

Objetivo: estudiar parámetros inmunológicos en pacientes con lesiones intraepiteliales (NIC) y carcinoma in situ del cuello uterino en el Instituto Nacional de Oncología y Radiobiología durante el año 2009. Métodos: se realizó un estudio en 20 pacientes donde se determinaron las características inmunofenotípicas de los linfocitos de sangre periférica mediante citometría de flujo y la capacidad funcional frente a diversos mitógenos utilizando el método de síntesis de DNA. El análisis de correlación entre variables inmunológicas y epidemiológicas se realizó mediante el cálculo del coeficiente de correlación de Pearson. Para las pruebas estadísticas se utilizó el paquete estadístico SPSS (versión 11.5). Resultados: la subpoblación de los linfocitos Tc CD8+, mostró valores superiores estadísticamente significativos (p=0,004) solo para las pacientes con NIC I. En todas las pacientes, independientemente del estadio de la enfermedad y del mitógeno utilizado, los índices de estimulación (IE) resultaron inferiores a los valores del grupo control. Conclusión: las alteraciones en el sistema inmune en las pacientes con patología de cuello están asociadas al progreso de la enfermedad y las células T son fundamentales en el control de la progresión de las lesiones

Objective: To study the immunologic parameters in patients presenting with intraepithelial lesions (IEL) and carcinoma in situ of cervix in the National Institute of Oncology and Radiotherapy over 2009. Methods: A study was conducted in 20 patients to determine the immuno-phenotypical of lymphocytes in peripheral blood by flow-cytometry and the functional ability in face of diverse mitogen using the AND synthesis method. The correlation analysis among the immunologic and epidemiologic variables was carried out by an estimation of Pearson's correlation coefficient. For the statistic test the SPSS statistical package was used (version 11.5). Results: The subgroup of Tc + CD8 lymphocytes showed higher values statistically significant ( p= 0.004) only for patients presenting with IEL. In all patients, independently of disease stage and of the mitogen used, the stimulation rates (SR) were lower than the values of controls. Conclusions: The alterations in the immune system in patients with cervix pathology are associated with the progress of lesions

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.

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.

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.

Expression of the nonclassical HLA-G and HLA-E molecules in laryngeal lesions as biomarkers of tumor invasiveness.

INTRODUCTION: HLA-G and HLA-E are two nonclassical class I molecules, which have been well recognized as modulators of innate and adaptive immune responses, and the expression of these molecules in virus infected cells has been associated with subversion of the immune response. OBJECTIVE: In this study we performed a cross-sectional study, systematically comparing the expression of HLA-G and HLA-E in benign, pre-malignant and malignant laryngeal lesions, correlating with demographic and clinical variables and with the presence of high-risk and low-risk HPV types. MATERIALS AND METHODS: Laryngeal lesions were collected from 109 patients and stratified into 27 laryngeal papillomas, 17 dysplasias, 10 in situ laryngeal carcinomas, 27 laryngeal carcinomas without metastases, 28 laryngeal carcinomas with metastasis along with their respective draining cervical lymph nodes, and 10 normal larynx specimens. The expression of HLA-G and HLA-E molecules was determined by immunohistochemistry. HPV DNA detection and typing was performed using generic and specific primers. RESULTS: HLA nonclassical molecules showed a distinct distribution pattern, according to the larynx lesion grade. HLA-G expression increased in benign and premalignant lesions, and gradually decreased in invasive carcinomas and in respective draining cervical lymph nodes. Conversely, HLA-E expression increased as far as lesion grade increased, including increased molecule expression in the draining lymph nodes of malignant lesions. Only 17 (15.6%) patients were HPV DNA positive. CONCLUSIONS: Overexpression of HLA-E and underexpression of HLA-G appear to be good markers for malignant larynx lesion.

The role of macrophages and mast cells in lymphangiogenesis and angiogenesis in cervical carcinogenesis.

During carcinogenesis it is known that growth factors and cytokines from stromal and inflammatory cells from the microenvironment promote angiogenesis and lymphangiogenesis. However, the participation of macrophages and mast cells in these processes is not well understood. The aim of this study was to evaluate the relationship between mast cell and macrophage density with blood and lymphatic vessels in various stages of carcinoma of the uterine cervix. Tissue sections from archival paraffin-embedded samples from cases with cervical intraepithelial neoplasias (CIN) 1, 2, 3, carcinoma in situ, and invasive carcinoma were used. Immunohistochemical staining was done using the following antibodies: anti-LYVE-1; anti-CD31; anti-CD68, and anti-tryptase. Our results showed a significant increase in the number of macrophages in carcinoma in situ, a correlation between lymphatic vessels and macrophages in premalignant lesions CIN 2, and a correlation between mast cells and blood vessels in both CIN 2 and carcinoma in situ. In conclusion, our data underscore the importance of the recruitment of macrophages and mast cells in the development of tumor-associated blood and lymphatic capillaries.

Immunohistochemical detection of Ki-67 is not associated with tumor-infiltrating macrophages and cyclooxygenase-2 in oral squamous cell carcinoma.

BACKGROUND: An inflammatory component consisting of cells and chemical mediators may influence the proliferation and dissemination of the oral squamous cell carcinoma (OSCC). In the present study, we evaluated the possible relationship between Ki-67, tumor-associated macrophages (TAMs), and COX-2 in OSCCs. In addition, the immunodetection of these proteins was associated with different histological grades of malignancy, including invasive and in situ tumors. METHODS: Twenty-seven OSCC cases were examined by light microscopy using criteria adopted WHO, and immunohistochemistry for Ki-67, CD68, and COX-2 using EnVision System in invasive and in situ lesions. Immunohistochemical detection of these proteins was assessed and scored for COX-2, and results were compared with their histological grades of malignancy. RESULTS: A correlation between Ki-67, COX-2, and CD68 was not found. Histological grade of malignancy (HDM) was associated with the Ki-67 immunostaining (P = 0.00), but this was not observed regarding both CD68 (P = 0.51) and COX-2 (P = 0.89). Furthermore, there was a COX-2 overexpression in 62.96% of the sample, and a high density of TAMs in both OSCCs and in situ carcinomas. CONCLUSIONS: Imunolabeling for Ki-67 was directly correlated with less-differentiated tumors, suggesting that this marker may contribute to understand the biological behavior of OSCC, and help to distinguish risk groups of OSCC. Furthermore, the lack of correlation between Ki-67, COX-2, and CD68 indicates that the latter two markers may play a pivotal role in oral carcinogenesis. However, further studies are needed to clarify their contribution for cell proliferation and tumor differentiation.

Ki-67 expression in anal intraepithelial neoplasia in AIDS.

CONTEXT: AIDS is one of the most important risk factors for progression and recurrence of anogenital condyloma. In a previous work, we observed that patients with warts and high-grade AIN (HAIN) had recurrences more frequently than did patients with warts without AIN. The mechanisms of this increased incidence of high-grade lesions in AIDS are not known. OBJECTIVE: We studied the expression of the proliferative marker Ki-67 by immunohistochemical methods, in specimens of anal condyloma from HIV+ patients to clarify whether its expression can be associated to the grade of AIN. DESIGN: A retrospective study of histological specimens. SETTING: University referral unit. SAMPLE: 34 patients were divided into two groups: (1) condylomas with low grade AIN (LAIN), with 25 patients; and (2) condylomas with HAIN, with 9 patients. In this latter group we examined two areas: 2A (HAIN area) and 2B (LAIN area). MAIN MEASUREMENTS: The immunohistochemical reaction for Ki-67 was done on histological sections. Slices were lightly stained with hematoxylin, to help us in Ki-67 positive cell counting. The percentage of Ki-67 marked nuclei was calculated. We applied one-way variance analysis for statistics. RESULTS: The mean number of Ki-67 positive cells in group 1 was 19.68 +/- 10.99; in group 2 (area A) it was 46.73 +/- 10.409; and in area B it was 36.43 +/- 14.731. There were statistical differences between groups 1 and 2A and between groups 1 and 2B. Ki-67 positive cells predominated in the lower layer in LAIN. Positive Ki-67 cells were found in all layers in group 2A, and in group 2B they predominated in the two lower or in all layers of the epithelium. CONCLUSIONS: Our results suggest that LAIN areas (using routine staining techniques) in HAIN can have a biological behavior more similar to HAIN.

Serological response to HPV16 in CIN-III and cervical cancer patients. Case-control studies in Spain and Colombia.

This study evaluates the association of antibodies against HPV-16-derived peptides with cervical cancer and estimates the sensitivity and specificity of the serological assays in relation to HPV DNA detection in cervical cells by PCR. Study subjects were derived from 4 case-control studies carried out in Spain and Colombia. Sera from 544 cases of CIN III and invasive cancer and of 543 age-matched controls were tested for antibodies to 5 peptides derived from E2, E7 (3 partially overlapping frames of HPV 16 denoted E7/ 1, E7/2, E7/3) and L2 open reading frames of HPV 16. HPV DNA was detected using a L1-PCR based method. Among cancer controls, antibody response to E2 and E7/1, E7/2, E7/3 was higher in Colombia (22.5%,7.2%,11.7%,12.6% respectively) than in Spain (17.1 %, 4.7%, 5.9%, 5.9%). E7 antibodies were related to stage, particularly in CIN III vs. invasive stages and less markedly within invasive stages. Detection of antibodies to the E7/1 was associated to CIN III (OR = 1.8). The risk of invasive cervical cancer was increased among those with antibodies to E2 (OR = 2.2), to E7/1 (OR = 4.2), to E7/2 (OR = 4.3), and to E7/3 (OR = 2.5). Presence of antibodies to all the 3 E7 peptides increased the risk of CIN III (OR = 5.6) and that of invasive cancer (OR = 17.5). High levels of antibodies to E7/1 or E7/2 or E7/3 increased the risk of invasive cervical cancer (OR for high levels of antibodies vs. negatives to E7/1 OR = 22.6; E7/2 OR = 7.5, E7/3 OR = 3.4). In the present analysis, antibodies to L2 were not associated with either CIN III or cervical cancer. Serological markers of HPV 16 detected less than half of the HPV-16-DNA-positive cases. It is concluded that antibodies to E2 and particularly E7 antigens are strongly associated with cervical cancer. Antibodies to E7 seem to be a moderate marker of tumor burden.

Cervical intraepithelial neoplasia is not associated with elevated serum neopterin levels.

Neopterin, a marker of cellular immune activation, was elevated in patients who had cervical cancer in previous studies. To examine neopterin in the presence of precursors to cervical cancer (i.e., cervical intraepithelial neoplasia) we measured serum levels in 185 colposcopy patients in Jamaica, a country with high cervical cancer incidence, and in 72 age-matched Jamaican women selected from a large population-based sample. We also measured serum levels of beta-2-microglobulin, another commonly used marker of immune activation. Neopterin and beta-2-microglobulin levels were not elevated in colposcopy patients; neither were they related to severity of cervical neoplasia. In multivariable analysis, neither adjustments for detection of cervical human papillomavirus DNA by PCR nor detection of antibodies to human T-cell lymphotropic virus type I (a retrovirus endemic to Jamaica) altered our findings. The absence of a serologically detectable increase in cellular immune activation linked to cervical intraepithelial neoplasia suggests that the immunological response to cervical intraepithelial neoplasia does not involve substantial systemic cellular immune activation.

Simple mucin-type carbohydrate antigens (T, sialosyl-T, Tn and sialosyl-Tn) in breast carcinogenesis.

Immunohistochemical analysis of the expression of simple mucin-type carbohydrate antigens (Tn, sialyl-Tn and T) was performed in a series of 43 cases of intraductal hyperplasia without atypia, 9 cases of intraductal hyperplasia with atypia, 54 cases of ductal carcinoma in situ (DCIS) and 26 cases of invasive breast carcinoma. We also studied 36 cases of isolated breast normal epithelium, 20 cases of "normal" breast epithelium adjacent to neoplasms and 14 cases of apocrine metaplasia. All antigens were detected in different frequencies in normal, hyperplastic, metaplastic and neoplastic breast epithelium. Tn and sialyl-Tn are expressed more frequently in malignant than in benign breast epithelium; while Tn expression increases from normal to invasive carcinomas, sialyl-Tn increases until DCIS and drops in invasive carcinomas, suggesting that either there is a failure of a proportion of DCIS to progress to invasive carcinoma or loss of expression of sialyl-Tn when some carcinomas become invasive. The high frequency of Tn and sialyl-Tn expression in breast intraductal proliferations probably reflects incomplete glycosylation in these lesions, which is a well-known tumour-associated phenomenon and supports the assumption that such lesions are putative precursors of breast cancer. T antigen was expressed in all groups studied, but its prevalence differed significantly between normal and neoplastic epithelium. The expression of these antigens in epithelium adjacent to carcinomas is similar to that found in isolated normal breast epithelium, whereas apocrine metaplasia has a pattern of simple mucin-type glycosylation that is specific and distinct from that of the normal breast epithelium, with a high frequency of marked expression of Tn and sialyl-Tn. The similarity of the pattern of expression of simple mucin-type antigens in metaplasia and malignant neoplasia reduces the usefulness of these markers from a diagnostic standpoint.