Proteomics-based identification of cancer-associated proteins in chronic lymphocytic leukaemia

BACKGROUND: Chronic lymphocytic leukaemia (CLL) is a neoplasm of B-cells characterized by variable prognosis. Exploring the proteome of CLL cells may provide insights into the disease. Therefore, eleven proteomics experiments were conducted on eleven primary CLL samples. RESULTS: We reported a CLL proteome consisting of 919 proteins (false discovery rate (FDR) 1%) whose identification was based on the sequencing of two or more distinct peptides (FDR of peptide sequencing 1%). Mass spectrometry-based protein identification was validated for four different proteins using Western blotting and specific antibodies in different CLL samples. Small sizes of nucleolin (~57 kDa and ~68 kDa) showed a potential association with good prognosis CLL cells (n = 8, p < 0.01). Compared with normal B-cells, CLL cells over-expressed thyroid hormone receptor-associated protein 3 (THRAP3; n = 9; p = 0.00007), which is implicated in cell proliferation; and heterochromatin protein 1-binding protein 3 (HP1BP3; n = 10; p = 0.0002), which promotes cell survival and tumourogenesis. A smaller form of HP1BP3, which may correspond to HP1BP3 isoform-2, was specifically identified in normal B-cells (n = 10; p = 0.0001). HP1BP3 and THRAP3 predicted poor prognosis of CLL (p 0.05). Consistently, THRAP3 and HP1BP3 were found to be associated with cancer-related pathways (p 0.05). CONCLUSIONS: Our findings add to the known proteome of CLL and confirm the prognostic importance of two novel cancer-associated proteins in this disease.

Differential effect of the route of inoculation of rabies virus on NeuN immunoreactivity in the cerebral cortex of mice / Efecto diferencial de la vía de inoculación del virus de la rabia sobre la inmunorreactividad de NeuN en la corteza cerebral de ratones

Rabies is a lethal disease caused by a neurotropic virus that produces inconspicuous morphological changes hardly observable with conventional histopathology. The fatal outcome caused by rabies could be attributed to specific biochemical changes that severely impact neuronal function. The neuronal nuclear protein (NeuN) has become a widely used neuronal marker for the research and the histopathological diagnosis of nervous system diseases. To evaluate the distribution of the protein NeuN in the motor cortex of normal and rabies-infected mice adult ICR mice were inoculated with rabies virus either intramuscularly or intracerebrally. Rabies-infected mice were sacrificed at the terminal stage of the disease. Control mice were also euthanized at the same age. The brains were removed and cut into coronal sections on a vibratome. Immunohistochemistry was used to study the expression of NeuN in the motor area of the cerebral cortex. Neuronal counts, cellular optical densitometry and neuronal diameter measurements were performed to analyze the immunoreactivity of the protein. All parameters revealed decreased immunoreactivity for NeuN in cortical neurons of mice intracerebrally infected with rabies. In contrast, the changes were not statistically significant in mice inoculated intramuscularly. Either the immunoreactivity of NeuN or its expression is affected by the presence of rabies virus in the cerebral cortex depending on the inoculation route. These results contribute to the knowledge of the dynamics of cellular infection on rabies pathogenesis.

La rabia es una enfermedad mortal causada por un virus neurotrópico que produce discretos cambios morfológicos difícilmente observables con la histopatología convencional. El desenlace fatal causado por la rabia puede atribuirse a cambios bioquímicos específicos que afectan gravemente la función neuronal. La proteína nuclear neuronal (NeuN) es un marcador ampliamente utilizado para la investigación y el diagnóstico histopatológico de enfermedades del sistema nervioso. Este trabajo se realizó con el propósito de evaluar la distribución de la proteína NeuN en la corteza motora de ratones normales y ratones infectados con virus de la rabia. Ratones ICR adultos fueron inoculados con virus de la rabia por vía intramuscular o por vía intracerebral. Los animales infectados con rabia fueron sacrificados en la etapa terminal de la enfermedad. Ratones de la misma edad no inoculados con el virus (controles) fueron sacrificados simultáneamente. Se extrajeron los cerebros y se obtuvieron cortes coronales en un vibrátomo. Mediante inmunohistoquímica se estudió la expresión de NeuN en el área motora de la corteza cerebral. Se realizaron conteos neuronales, densitometría óptica celular y mediciones del diámetro de los perfiles neuronales para analizar la inmunorreactividad de la proteína. En los ratones inoculados por vía intracerebral hubo disminución significativa de la inmunorreactividad de NeuN manifestada en los diferentes parámetros evaluados. En contraste, estos cambios no fueron estadísticamente significativos en los cerebros de ratones inoculados por la ruta intramuscular. La inmunorreactividad de NeuN o su expresión es afectada por la presencia del virus de la rabia en la corteza cerebral pero dependiendo de la vía de inoculación. Estos resultados contribuyen al conocimiento de las dinámicas de infección celular en la patogénesis de la rabia.

Clinicopathological values of NBS1 and DNA damage response genes in epithelial ovarian cancers

Epithelial ovarian cancers (EOCs) are highly lethal gynecological malignancies with a high recurrence rate. Therefore, developing prognostic markers for recurrence after chemotherapy is crucial for the treatment of ovarian cancers. As ovarian cancers frequently respond to DNA-damaging agents, we assessed the clinicopathological significance of key double-strand DNA break (DSB) repair genes, including BRCA1, BRCA2, BARD1, ATM, RAD51 and NBS1 in EOC cell lines and paraffin-embedded tissue sections from 140 EOC patients treated with cytoreductive surgery, followed by platinum-based chemotherapy. These samples were analyzed for the clinicopathological impact of DSB genes by western blot analysis, immunohistochemistry and quantitative real-time PCR. Of the DSB repair genes, BRCA1, ATM and NBS1, which are involved in the homologous recombination-mediated repair pathway, were related to aggressive parameters in EOC. When survival analysis was performed, NBS1 expression exhibited an association with EOC recurrence. Specifically, increased NBS1 expression was found in 107 out of 140 cases (76.0%) and correlated with advanced stage (P=0.001), high grade (P=0.001) and serous histology (P=0.008). The median recurrence-free survival in patients with positive and negative expression of NBS1 was 30 and 78 months, respectively (P=0.0068). In multivariate analysis, NBS1 was an independent prognostic factor for the recurrence of EOC. Together, these results suggest that NBS1 is a marker of poor prognosis for the recurrence of EOC and is associated with aggressive clinicopathological parameters.

Immunohistochemical detection of the latent nuclear antigen-1 of the human herpesvirus type 8 to differentiate cutaneous epidemic Kaposi sarcoma and its histological simulators / Detecção imuno-histoquímica do antígeno nuclear latente-1 do herpesvirus tipo 8 para diferenciar o sarcoma de Kaposi epidêmico cutâneo de seus simuladores histológicos

Kaposi's sarcoma is the most common neoplasia diagnosed in AIDS patients and the expression of the human herpesvirus-8 (HHV-8) latent nuclear antigen-1 has been useful for its histological diagnosis. The aim of this study is to confirm that immunohistochemistry is a valuable tool for differentiating KS from its simulators in skin biopsies of HIV patients. Immunohistochemical and histological analyses were performed in 49 Kaposi's sarcoma skin biopsies and 60 of its histological simulators. Positivity was present in the 49 Kaposi's sarcoma skin biopsies and no staining was observed in the 60 simulators analyzed, resulting in sensibility and specificity of 100%. HHV-8 immunohistochemical detection is an effective tool for diagnosing Kaposi's sarcoma, especially in early lesions in which neoplastic features are not evident. It also contributes to its histological differential diagnosis.

O sarcoma de Kaposi é a neoplasia mais diagnosticada em pacientes com SIDA e a expressão do antígeno nuclear latente-1 do herpesvírus humano tipo-8 (HHV-8) tem se mostrado útil no seu diagnóstico histológico. O objetivo deste estudo é confirmar que o método imuno-histoquímico é uma ferramenta útil para diferenciar o sarcoma de Kaposi cutâneo de seus simuladores histológicos em pacientes HIV positivos. Análise histológica e imuno-histoquímica foram realizadas em 49 casos de sarcoma de Kaposi cutâneo e 60 casos de seus simuladores histológicos. Positividade à imuno-histoquímica para o antígeno nuclear latente 1 do HHV-8 foi observada nos 49 casos de sarcoma de Kaposi e nenhuma reação foi detectada nos 60 simuladores analisados, resultando em 100% de sensibilidade e especificidade. A detecção do HHV-8 por imuno-histoquímica é uma ferramenta útil para o diagnóstico de sarcoma de Kaposi, especialmente na lesão inicial cujo caráter neoplásico não é evidente, e contribui para seu diagnóstico diferencial histológico.

Autoanticorpos em esclerodermia e sua associação ao perfil clínico da doença: estudo em 66 pacientes do sul do Brasil / Autoantibodies in scleroderma and their association with the clinical profile of the disease: a study of 66 patients from southern Brazil

FUNDAMENTOS: A esclerodermia é uma colagenose relativamente rara, cujo perfil de autoanticorpos está associado a diferentes manifestações clínicas. A prevalência de autoanticorpos na esclerodermia sofre influência racial e genética. OBJETIVO: Estudar a prevalência dos anticorpos anti-Scl-70, anticentrômero e anti-U1-RNP em pacientes com esclerodermia do sul do Brasil e verificar suas associações às manifestações clínicas. MÉTODOS: Estudo retrospectivo de análise de 66 pacientes com esclerodermia para presença de anti-Scl-70, anticentrômero (ACA) e anti-U1-RNP e de manifestações clínicas como: Raynaud, cicatrizes estelares, necrose digital, telangiectasias, calcinose, fibrose pulmonar, pleurites, pericardites, miocardiopatias, artralgias e artrites, grau de esclerose da pele, contraturas articulares e atritos de tendão, hipertensão pulmonar, manifestações esofágicas e crise renal. RESULTADOS: A prevalência do anti-Scl-70 foi de 17,8 por cento, a do ACA, de 33,3 por cento, e a do U1 RNP foi de 11,8 por cento. O anti-Scl-70 estava associado à forma difusa da doença (p=0,015), presença de miocardiopatias (p=0,016) e de cicatrizes estelares (p=0,05); o anticentrômero foi mais comum na forma limitada, embora sem significância estatística e mostrou-se protetor para as miocardiopatias (p=0,005). O anti-U1-RNP foi mais comum nas formas de superposição (p=0,0004). CONCLUSÃO: A prevalência e o perfil de associações clínicas dos autoanticorpos em esclerodermia de pacientes brasileiros assemelham-se aos da literatura mundial.

BACKGROUND: Scleroderma is a fairly rare connective tissue disease whose autoantibody profile is associated with different clinical manifestations. The prevalence of autoantibodies in scleroderma is influenced by race and genetics. OBJECTIVE: To study the prevalence of anti-Scl-70, anti-centromere (ACA) and anti-U1-RNP antibodies in patients with scleroderma in southern Brazil and verify their association with clinical manifestations of the disease. METHODS: A retrospective study involving 66 patients with scleroderma for the presence of anti-Scl-70, anti-centromere and anti-U1-RNP and of clinical manifestations such as Raynaud's phenomenon, digital micro scars, digital necrosis, telangiectasias, calcinosis, pulmonary fibrosis, pleuritis, pericarditis, cardiomyopathy, arthralgia and arthritis, skin sclerosis, joint contractures, tendon friction rubs, pulmonary hypertension, esophageal disorders and renal crisis. RESULTS: The prevalence of anti-Scl-70 was 17.8 percent , that of ACA was 33.3 percent and the prevalence of U1 RNP was 11.8 percent. Anti-Scl-70 was associated with the diffuse form of the disease (p = 0.015), presence of cardiomyopathies (p = 0.016) and digital micro scars (p = 0.05). Anti-centromere was more common in the limited form, although it was not statistically significant, and had a protective role associated with cardiomyopathies (p = 0.005). Anti-U1-RNP was more common in the overlap forms (p = 0.0004). CONCLUSION: The prevalence and profile of clinical associations of autoantibodies in Brazilian patients with scleroderma are similar to those found in the literature.

The enigmatic role of nucleophosmin in malignant melanoma: Does it have an effect.

Background: Melanoma is quite a heterogeneous group of diseases of the skin. Prognostic markers of tumor behavior are important to precisely assign individual patients for appropriate treatment protocols. Aim: The aim of our first study was to investigate nucleophosmin expression in melanoma patients and to determine its relationship with the tumor characterictics and patient prognosis. Materials and Methods: We analyzed the immunohistochemical expression of nucleophosmin in 55 melanoma patients. The immunostaining pattern was classified into two groups: Diffuse nuclear and nucleolar relocalization. We also investigated the relationship between the expression of nucleophosmin and the clinicopathological parameters sucssh as Clark level, tumor thickness, stage, histological type, location, and survey. Results: In all cases the neoplastic cells were strongly positive for nucleophosmin (14 cases diffuse nuclear, 41 cases nucleolar relocalization). No correlation was demonstrated between the expression pattern of nucleophosmin and the clinicopathological parameters and survey. Conclusions: The implications of our results, nevertheless, are that the immunohistochemical detection of nucleophosmin is not a valuable tool for predicting the outcome of patients with melanoma or identifying subgroups of patients who may be at a higher risk.

Minichromosome maintenance 2 and 5 expressions are increased in the epithelium of hereditary gingival fibromatosis associated with dental abnormalities

INTRODUCTION: Gingiva fibromatosis is a relatively rare condition characterized by diffuse enlargement of the gingiva, which is caused by expansion and accumulation of the connective tissue. OBJECTIVE: The aim of the present study was to investigate proliferative and apoptotic biomarker expression in normal gingiva and two forms of gingival fibromatosis. METHODS: Archived tissue specimens of hereditary gingival fibromatosis, gingival fibromatosis and dental abnormality syndrome and normal gingiva were subject to morphological analysis and immunohistochemical staining. The results were analyzed statistically. RESULTS: Proteins associated with proliferation were found in the nuclei of epithelial cells from the basal and suprabasal layers, whereas apoptotic proteins were detected in the cytoplasm of the upper layers of the epithelium. Increased expressions of minichromosome maintenance proteins 2 and 5 were observed in the gingival fibromatosis and dental abnormality syndrome samples. In contrast, geminin expression was higher in normal gingiva samples. No difference in the expression of apoptotic proteins was observed among the groups. CONCLUSION: Our findings support a role for augmented proliferation of epithelial cells within the overgrown tissues associated with gingival fibromatosis or dental abnormality syndrome. However, our data suggest that different biological mechanisms may account for the pathogenesis of different types of gingival fibromatosis.

Compromiso pulmonar en esclerosis sistémica / Lung involvement in systemic sclerosis

El objetivo del estudio fue determinar las características clínicas de los pacientes con esclerodermia y compromiso pulmonar y evaluar si existen factores clínicos predictores de mayor riesgo de enfermedad intersticial. Se estudiaron en forma retrospectiva 40 pacientes con esclerodermia. Fueron divididos en 2 grupos: capacidad de difusión del monóxido de carbono (DLCO) normal (n = 22) y DLCO disminuida (n = 18, 45%). Los pacientes con DLCO disminuida no fueron diferentes en edad (51.1 más o menos 13.5 vs. 53.5 más o menos 9.3 años, p = 0.5182), sexo (varones 13.6%, p = 0.6088 ), presencia de Raynaud (86.6% vs. 85%, p = 0.6272), síndrome de ojo seco (6.2% vs. 10.5%, p = 1.0000) prevalencia de enfermedad difusa (94.1% vs. 83.3%, p = 0.6026) o de dilatación esofágica. El tiempo de evolución de la enfermedad no fue diferente. La sensibilidad de la disnea para detectar una DLCO alterada fue 46.6% con una especificidad del 90% y la de la caída de la saturación de O2 (SaO2) del 71.4% y 80% respectivamente. Los pacientes con DLCO baja tuvieron mayor prevalencia de anticuerpos anti-Scl 70 positivos (5/9 vs. 0/11, p = 0.0081) y de incapacidad ventilatoria restrictiva aunque en 56.7% de los pacientes con DLCO disminuida la capacidad pulmonar total (CPT) era normal. La presencia de hipertensión pulmonar medida por ecocardiograma Doppler fue idéntica (11/13 vs. 10/11, p = 1.0000). Los pacientes con DLCO disminuida tuvieron una prevalencia muy superior de tomografía computada de tórax con evidencias de compromiso intersticial (82.3% vs. 5.8%, p menor o igual a 0.0001). En conclusión, nuestros datos sugieren que la disminución de la DLCO es un hallazgo, muy frecuentemente asociado a TAC de tórax con compromiso intersticial y que no hay variables clínicas que permitan predecir su anormalidad.

The objective of this study was to determine clinical predictors of interstitial lung disease in patients with systemic sclerosis (SSc) and pulmonary involvement as defined by presence of a decreased diffusing capacity for carbon monoxide (DLCO). Forty subjects with SSc were retrospectively evaluated. Patients were categorized according to their level of DLCO (less than or more than or equal to 80% of predicted). Sensitivity of dyspnea to detect a decreased DLCO was 46.6% and specificity 90%, whereas oxygen desaturation showed a sensitivity of 71.4% and a specificity of 80%. Patients with decreased DLCO (n = 18) were not different in age (51.1 more or less than 13.5 vs. 53.5 more or less than 9.3 y, p = 0.5182), sex (male 13.6%, p = 0.6088), prevalence of Raynaud (86.6% vs. 85%, p = 0.6272), sicca syndrome (6.2% vs. 10.5% p = 1.0000) diffuse cutaneous involvement (94.1% vs. 83.3%, p = 0.6026) or esophageal dilatation. The duration of symptoms since diagnosis was no different. Prevalence of pulmonary hypertension assessed by Doppler echocardiography or abnormal nailfold capillaroscopic findings were identical in both populations. Patients with low DLCO had a significatly higher prevalence of anti topoisomerase antibodies. (5/9 vs. 0/11, p = 0.0081) and restrictive lung disease. Patients with low DLCO showed a significantly higher prevalence of abnormal HRCT findings suggestive of ILD (82.3% vs. 5.8%, p less than or equal to 0.0001). We conclude that a low DLCO is a frequent finding in SSc patients, strongly associated with HRCT signs of ILD. We have not found clinical factors predictive for a low DLCO.

Ki-67 cell proliferation in familial and in esporadic unilateral retinoblastoma: case report

PURPOSE: Ki-67 is a nuclear protein that is expressed at all phases of the cell cycle except the resting phase. This study is a clinicopathologic observational case report that aims to report on the cell proliferation rates, as measured by the Ki-67 antigen, in two enucleated retinoblastoma eyes. METHODS: One unilateral familial (mother with unilateral disease - patient 1) and one unilateral sporadic retinoblastoma (patient 2) patients were submitted to enucleation without previous treatment. The tumor cell proliferation rate was assessed by the Ki-67 antigen labeling index (stained cells / 100 cells) in five different fields of the tumor. RESULTS: Patient 1 was 23 months old and the tumor was exophytic with associated neovascularization of the iris; patient 2 was 6 years old and the tumor was endophytic with coarse vitreous seeds. Both enucleated eyes presented optic nerve with free surgical margins. Positive Ki-67 cell index in patient 1 varied from 75 to 90 (MD ± SD: 79.5 ± 6.61) and in patient 2 from 38 to 60 (MD ± SD: 46.6 ± 8.2). CONCLUSIONS: The familial retinoblastoma, besides the earlier age presentation, showed 45.8 percent more Ki-67 positive cells than the same stage sporadic one. This proliferation rate may explain the earlier presentation age of the tumor in the inherited disease.

OBJETIVO: O Ki-67 é antígeno nuclear que se expressa em todas as fases do ciclo celular, exceto no período de repouso. Este é um estudo de casos com correlação clínico-patológica que visa avaliar a taxa de proliferação celular, medida pelo antígeno Ki-67, em 2 olhos enucleados com retinoblastoma. MÉTODOS: Um paciente com retinoblastoma unilateral familiar (mãe com doença unilateral - paciente 1) e outro com retinoblastoma unilateral esporádico (paciente 2) foram submetidos à enucleação ocular sem outro tratamento prévio. A taxa de proliferação celular foi avaliada segundo índice obtido pela contagem de células marcadas com Ki-67, em 5 campos sob microscópia óptica (células marcadas/100 células). RESULTADOS: O paciente 1, com 23 meses de idade, apresentou tumor exofítico com neovascularização de íris associada; o paciente 2, de 6 anos, apresentou tumor de crescimento endofítico, com sementes vítreas importantes. Ambos os olhos enucleados apresentaram margens cirúrgicas do nervo óptico livres de neoplasia. O índice de células positivas no paciente 1 variou de 75 a 90 (Média ± DP: 79,5 ± 6,61), e no paciente 2, de 38 a 60 (Média ± DP: 46,6 ± 8,2). O retinoblastoma familiar, além de sua manifestação em idade mais precoce, apresentou 45,8 por cento mais células positivas que o retinoblastoma esporádico com o mesmo estadiamento. CONCLUSÃO: O retinoblastoma familiar, além de surgimento mais precoce, apresentou 45,8 por cento mais células em proliferação que o retinoblastoma esporádico em mesmo estádio. Essa taxa de proliferação pode explicar a menor idade de aparecimento do tumor nos casos de doença herdada.

Evaluation of nucleolar organizer regions in maxillary osteosarcoma

Maxillary osteosarcomas are a relatively frequent malignant tumor of the oral cavity. Similarly to other skeletal osteosarcomas, they exhibit different cellular differentiation patterns, i.e. chondroblastic, osteoblastic, or fibroblastic. Although their histological features resemble those of osteosarcomas of the long bones, their pattern of evolution usually differs. Morphometric variations in silver stained Nucleolar Organizer Regions (AgNOR) have proved of value to study the biology of several tumors. However, information on the analysis of AgNOR in maxillary tumors is scarce. The aim of the present study was to analyze the variations of different morphological parameters related to AgNOR in a series of 32 cases of maxillary osteosarcoma. In each case we analyzed 100 nuclei corresponding to the prevalent cellular differentiation type, selecting the most aggressive area. We employed software previously developed at our laboratory that yields information on different AgNOR-related parameters. The results were compared with those previously reported in a study on 12 cases of osteosarcoma of long bones. Six cases of oral mucosa squamous cell carcinoma were also included for comparative purposes. Single AgNOR volume proved to be the most discriminatory and informative parameter. The value of single AgNOR volume was considerably lower in mandible osteosarcomas than in osteosarcomas of the upper maxilla (p=0.02). The values were significantly lower in maxillary osteosarcomas than in long bone osteosarcomas and in oral carcinomas. This finding would suggest a slower rate of cell activity in maxillary osteosarcomas, associated in turn to its known lower degree of aggressiveness. The present results suggest that the analysis of AgNOR is a valuable and easily applicable marker to determine the degree of malignancy and biology of maxillary osteosarcomas.

Los osteoscaromas de maxilares son entidades relativamente frecuentes entre los tumores malignos de la cavidad bucal. Al igual que los osteosarcomas de otras localizaciones del esqueleto, pueden presentar diferentes patrones de diferenciación celular (condroblástico, osteoblástico o fibroblástico). Si bien sus características histológicas son similares, tienen generalmente un comportamiento evolutivo diferente al de los huesos largos. Las variaciones morfométricas de las regiones organizadoras del nucleoloidentificadas por impregnación argéntica (AgNOR) han demostrado ser marcadores útiles para el estudio de la biología de diversas entidades tumorales, pero hay muy escasa información de su análisis en tumores de los huesos maxilares. El objetivo de este trabajo fue analizar las variaciones de diferentes parámetros morfológicos de las AgNOR en una serie de 32 casos de osteosarcomas de maxilar. En cada caso se analizaron 100 núcleos en el patrón de diferenciación celular predominante, seleccionando la zona de mayor agresividad. Se utilizó un programa que aporta información sobre diferentes parámetros de AgNOR, desarrollado previamente en nuestro laboratorio. El parámetro más indicativo resultó ser el volumen individual de las AgNOR. Este parámetro en los osteosarcomas con localización mandibular fue considerablemente menor que aquellos localizados en maxilar superior (p=0.02). En los osteosarcomas de maxilar los valores fueron significativamente menores que en los de huesos largos y en los carcinomas bucales. Ellos podría ser indicativo de una menor actividad celular, a su vez asociada a su reconocida menor agresividad. Estos resultados sugieren que el análisis de AgNOR podría ser considerado como un marcador de utilidad y de fácil aplicación para determinar el grado de malignidad en osteosarcomas de maxilar y estimar su comportamiento biológico.

Distrofia muscular de Emery-Dreifuss: relato de caso / Emery-Dreifuss muscular dystrophy: case report

A distrofia muscular de Emery-Dreifuss é uma forma de distrofia muscular freqüentemente associada a contraturas articulares e defeitos de condução cardíaca, que pode ser causada pela deficiência da proteína emerina na membrana nuclear interna das fibras musculares. Descrevemos o caso de um homem de 19 anos com diminuição de força muscular, hipotrofia nas cinturas escapular e pélvica, disfagia, contraturas articulares em cotovelos e tornozelos, apresentando história familiar compatível com herança ligada ao cromossomo X. A investigação mostrou creatinaquinase sérica elevada, eletrocardiograma com bloqueio atrioventricular de primeiro grau e bloqueio de ramo direito, eletroneuromiografia normal, biópsia muscular com alterações miopáticas e a análise por imuno-histoquímica mostrou deficiência de emerina. São discutidas as manifestações clínicas e genéticas, alterações laboratoriais e eletroneuromiográficas, bem como, a importância do estudo do padrão de herança no aconselhamento genético destas famílias.

Significance of AgNOR score in benign and malignant soft tissue tumours.

AgNOR staining was employed on FNAC and histopathological sections obtained from patients with soft tissue tumours. The study comprised of 20 normal appearing soft tissues, 74 benign and 36 malignant soft tissue tumours. The slides were stained with AgNOR in order to differentiate between benign and malignant soft tissue tumours. The mean AgNOR count in normal appearing soft tissues, benign lesions and malignant lesions was 1.04+/-0.10 (0.94-1.14), 1.51+/-0.21 (1.1-2.1) and 4.96+/-1.33 (2.57-7.21) respectively. The mean AgNOR count was found to be higher in benign soft tissue tumours as compared to normal appearing soft tissues and the difference was found to be statistically significant. The mean AgNOR count in soft tissue sarcomas was found to be higher as compared to both normal appearing soft tissues and benign soft tissue tumours and the results were found to be statistically significant. The increased AgNOR score in both benign and malignant soft tissue tumours as compared to normal appearing soft tissues indicates high proliferative activity. Thus AgNOR staining is a simple and useful method for estimating tumour cell proliferation thereby differentiating normal appearing soft tissues from benign and malignant soft tissue tumours.

Role of proliferative markers in prostatic lesions.

Prostatic lesions on routine H&E stain sometimes cause diagnostic dilemma specially in premalignant lesions like A.A.H. and P.I.N. Proliferative markers (AgNOR, P.C.N.A) are of great help in this grey zone. Total 50 cases studied and provisional diagnosis after HIE stain revealed that 37 cases were B.H.P., 5 cases were A.A.H., 1 case was P.I.N and 7 cases were adenocarcinoma. Proliferative marker study revealed AgNOR count of B.H.P as (0.4 -2.5)/cell, of A.A.H as (1.5-3.2)/cell, of P.I.N as 4.8/cell and adenocarcinoma as (4.3-5.4)/cell. P.C.N.A index of B.H.P was (2-8)%, of A.A.H (17-35)%, of P.I.N 40% and of carcinoma (54-82)%. Proliferative marker study was of great help in distinguishing between benign and malignant and specially premalignant lesions like A.A.H and P.I.N, where routine histopathology diagnosis was confusing. In the study, P.C.N.A was found to be superior to AgNOR since the values for interpretation was wider.

Utility of Thyroid Transcription Factor-1 and Cytokeratin 20 in Identifying the Origin of Metastatic Carcinomas of Cervical Lymph Nodes

The identification of primary location of a metastatic tumor is a difficult diagnostic problem and sometimes can be facilitated by the use of immunohistochemical markers. Thyroid transcription factor-1 (TTF-1) is a 38-kDa nuclear homeodomain transcription factor that is expressed specifically in lung or thyroid neoplasms. Cytokeratin 20 (CK20) is a 46-kDa low-molecular-weight cytokeratin that shows restricted expression in adenocarcinomas of the gastrointestinal tract (GIT) and transitional cell carcinomas of the urinary tract. We studied the immunohistochemical expression of TTF-1 and CK20 in 68 metastatic carcinomas in cervical lymph nodes. The primary sites were the lung in 29 cases, stomach in 13, colorectum in 3, and other sites in 23. TTF-1 expression was detected in 69.0% of metastatic lung carcinomas and none in metastatic GIT carcinomas, whereas CK20 expression was detected in 68.8% of metastatic GIT carcinomas and none of metastatic lung carcinomas. TTF-1 had a specificity of 0.95 and a sensitivity of 0.69 for metastatic lung carcinoma, whereas CK20 had a specificity of 1.00 and a sensitivity of 0.69 for metastatic GIT carcinoma. These results indicate that TTF-1 and CK20 should be the first choice as a component of antibody panel to prove or to exclude the lung and GIT origin, respectively, especially in patients presenting with metastatic carcinomas of unknown primary site.

Human Papillomavirus 16/18 Expression of Endocervical Glandular Lesions: Relationship with p53 and MIB-1 Expressions

The pathogenesis of endocervical glandular lesions are not clearly understood. The aims of this study are to evaluate the etiologic role of human papillomavirus (HPV) 16/18 and the relationship of HPV 16/18, p53 and MIB-1 expressions in endocervical glandular dysplasia (EGD), adenocarcinoma in situ (AIS) and adenocarcinoma. The materials included 14 endocervical adenocarcinoma and 5 AIS and 18 high grade EGD and 39 low grade EGD. Immunohistochemistry for p53 and MIB-1, and in situ PCR for HPV 16/18 were done. HPV 16/18 positivity was 84.2%, 16.7% and 17.9% in malignant glandular lesion (adenocarcinoma and AIS), high grade EGD and low grade EGD, respectively. P53 protein expression rates of malignant glandular lesions, high grade EGD and low grade EGD were 31.6%, 11.1%, and 0%, respectively. High MIB-1 labelling index was found in 73.7% of malignant glandular lesions, but in only 5.7% and 3.6% of high and low grade EGD, respectively. There were statistically significant differences in HPV 16/18, p53 and MIB-1 expressions between malignant endocervical glandular lesions and EGD, but no significant difference in p53 and MIB-1 expressions in relation to HPV 16/18 expression. In malignant endocervical glandular lesions, HPV 16/18 infection may be a major causative factor, but not be related to p53 and MIB-1 expressions.

Avaliaçäo da atividade secretora, proliferaçäo celular e expressäo da proteína p27 em tumores hipofisários / Evaluation of secretory activity, cell proliferation and expression of p27 protein in hypophyseal tumors

A finalidade deste estudo foi avaliar a atividade secretora, proliferaçäo celular e inibiçäo do ciclo celular das neoplasias hipofisárias. Os tecidos foram obtidos de 13 pacientes incluindo acromegalia (n=5), adenomas näo-secretores (n=4), hiperprolactinemia (n=3) e doença de Cushing (n=1). Os espécimes foram examinados por técnica imuno-histoquímicacom anticorpos anti-ACTH, anti-TSH, anti-LH, anti-FSH, anti-PRL, anti-hGH, anti MIB-1 (análise da proliferaçäo celular)e anti-p27(análise da inibiçäo do ciclo celular). A expressäo imunohistoquímica quanto à atividade secretora nos tumores hipofisários demonstrou que todos produziam mais que um hormôni. As células positivas para o marcador de proliferaçäo celular MIB-1 mostraram-se presentes em aproximadamente 46 por cento dos casos. A maior taxa proliferativa foi encontrada nos tumores com com quadro clínico de acromegalia (80 por cento dos casos). A taxa d positividade para a proteína p27 foi de aproximadamente 38 por cento, sendo, em média, maior no tumor responsável pela doença de Cushing. Esses resultados demonstram que os adenomas multisecretores säo relativamente comuns. Os índices de proliferaçäo celular baixos e os nveis de p27 próximos àqueles observados em tecidos normais expressam o baixo nível de proliferaçäo celulas destes tumores.

Morphological and biochemical analysis of anti-nuclear matrix protein antibodies in human sera

Autoimmune sera have been used in the diagnosis of autoimmune diseases as well as the analysis of nuclear substructures. In an attempt to study the biological characteristics of the nuclear matrix, we screened human sera using immunofluorescent staining and immunoblot. We detected antibodies against nuclear matrix (NM), a remnant nonchromatin protein compartment after the treatment of detergent, salt and nuclease, in 212 out of 284 tested sera (74.6%) by immunoblot. Peptides with molecular weights of 70 kDa, 50 kDa and 25 kDa were detected in the order of frequency. Clinical informations of 198 out of 212 cases were available and went as follows: 38 cases were autoimmune diseases, such as systemic lupus erythematosus and rheumatoid arthritis; 132 non-autoimmune and non-neoplastic diseases; 16 neoplastic diseases and 12 cases unclassified. The immunofluorescent staining intensity by anti-nuclear matrix protein (NMP) antibodies decreased variably, but fibrillogranular, speckled and nucleolar immunolocalization patterns were retained after in situ fractionation. Ku70 and La protein were detected by anti-NMP antibodies. Immunolocalization by anti-NMP antibodies indicates that the NMPs constitute a variety of characteristic nuclear substructures and may serve as autoantigens in diverse human diseases. In addition, the presence of Ku70 and La protein as NMPs suggests that the NM can be functionally active in association with DNA or RNA.

Expresion de MMP-2 (colagenasa tipo IV) en el carcinoma gastrico / MMP-2 expression (type IV collagenase) in gastric cancer

La MMP-2 (colagenosa tipo IV) es una proteína que pertence a la familia de las metaloproteinas, cuya función está relacionada con la degradación de la matriz extracelular. Su capacidad para degradar el colágeno IV de las membranas basales podría transformala en un agente facilitador de la diseminación neoplásica. Para ver su relación con algunas características clínico-patológicas e inmunohistoquímicas del cáncer gástrico se estudiaron 98 casos de adenocarcinoma de estómago determinado por inmunohistoquímica la presencia de MMP-2 en las células neoplásicas. Los resultados mostraron que había correlación entre la presencia de MMP-2 con el nivel de invasión parietal del tumor (p=0.03) y con la presencia de metástasis en ganglios regionales (p=0.05). En cambio no hubo asociación entre la expresión de MMP-2 con la frequencia por sexo, la localización dentro del estómago, el tamaño tumoral, el tipo histológico, el grado histológico, ni la expresión de las proteínas MIB-1, bcl-2, c-erbB-2 y p53. Tampoco se relacionó con la presencia de recidiva de la enfermedad ni con la sobrevida a los 5 años.

MIB-1 proliferative index in parathyroid adenoma & hyperplasia.

A retrospective study on 22 cases of parathyroid adenoma, 9 cases of primary parathyroid hyperplasia and 14 specimens of normal suppressed glandular tissue was undertaken to determine the usefulness of proliferative index (PI) for discriminating adenoma from hyperplasia, as an adjunct to the existing histological criteria. PI was determined by avidin-biotin-complex immunostaining after high temperature microwave antigen retrieval in paraffin sections, using monoclonal MIB-1 antibody which detects paraffin resistant analogue of cell cycle-associated Ki-67 antigen. PI expressed as percentage positive cell nuclei, was 1.36 +/- 0.62 (range 0.04-2.72) in adenoma, 1.17 +/- 0.83 (0.02-1.98) in hyperplasia and 0.03 +/- 0.02 (0.00-0.06) in normal suppressed glandular tissue. While the difference between normal suppressed glandular tissue and adenoma and hyperplasia was significant (P < 0.001), that between adenoma and hyperplasia was not. We conclude that although PI could distinguish between normal suppressed glandular tissue versus glands with primary hyperparathyroidism, it failed as an additional useful parameter for discriminating between adenoma and hyperplasia, both of which have low but similar proliferative activity.

Correlation of the growth fraction, nucleolar organizer region counts and epidermal growth factor receptor with histomorphological prognostic criteria in breast cancer.

A total of 60 cases of breast cancer were studied to find the correlation between newer parameters of prognosis viz growth fraction (GF), nucleolar organizer region (AgNOR) counts and epidermal growth factor receptor (EGFR) positivity with the various histomorphological factors such as tumour size, nuclear grade, histological grade and mitoses. Growth fraction measured by Ki67 monoclonal antibody varied from 3.6 to 73.7 percent. AgNOR counts ranged from 1.2 to 16.9 dots per tumour cell nucleus with a mean of 6.26 dots. EGFR positivity was seen in 66.7% of cases. Ki67 score correlated with all prognostic variables studied except the presence of axillary metastases. AgNOR counts correlated with size of tumour, axillary lymph-node metastases and Ki67 score but no correlation was seen with histologic or nuclear grade. EGFR positivity correlated strongly with size of the tumour and weakly with Ki67 score, AgNOR counts and mitoses.