Diagnostic value of α-enolase expression and serum α-enolase autoantibody levels in lung cancer.

OBJECTIVE: To investigate the diagnostic value of α-enolase (ENO1) and serum ENO1 autoantibody levels in lung cancer. METHODS: Immunohistochemistry staining and ELISA were performed to detect ENO1 expression in lung tissue and serum ENO1 autoantibody levels, respectively. RESULTS: The expression of ENO1 was higher in lung cancer tissues than in benign lung disease tissues (p < 0.001). The proportion of lung cancer samples expressing ENO1 was not significantly different among the various pathological classification groups. The proportion of samples expressing ENO1 was higher in lung cancer patients in stages I/II than in those in stages III/IV (χ2 = 5.445; p = 0.018). The expression of ENO1 in lung cancer tissues was not associated with age, gender, or smoking history. Serum ENO1 antibody levels were significantly higher in the lung cancer group than in the benign lung disease and control groups (p < 0.001). The differences among the pathological classification groups were not statistically significant. Serum ENO1 antibody levels were also in lung cancer patients in stages I/II than in those in stages III/IV (p < 0.01). Serum ENO1 antibody levels were not associated with age, gender, or smoking history in lung cancer patients. The ROC curve representing the diagnosis of lung cancer based on ENO1 antibody levels had an area under the curve of 0.806. CONCLUSIONS: Our results suggest that high levels of ENO1 are associated with the clinical stage of lung cancer and that ENO1 expression and its serum autoantibody levels show diagnostic value in lung cancer.

Identification of the alpha-enolase P46 in the extracellular membrane vesicles of Bacteroides fragilis.

BACKGROUND: Members of the Bacteroides fragilis group are the most important components of the normal human gut microbiome, but are also major opportunistic pathogens that are responsible for significant mortality, especially in the case of bacteraemia and other severe infections, such as intra-abdominal abscesses. Up to now, several virulence factors have been described that might explain the involvement of B. fragilis in these infections. The secretion of extracellular membrane vesicles (EMVs) has been proposed to play a role in pathogenesis and symbiosis in gram-negative bacteria, by releasing soluble proteins and other molecules. In B. fragilis, these vesicles are known to have haemagglutination and sialidosis activities, and also contain a capsular polysaccharide (PSA), although their involvement in virulence is still not clear. OBJECTIVE: The aim of this study was to identify proteins in the EMV of the 638R B. fragilis strain by mass spectrometry, and also to assess for the presence of Bfp60, a surface plasminogen (Plg) activator, previously shown in B. fragilis to be responsible for the conversion of inactive Plg to active plasmin, which can also bind to laminin-1. METHODS: B. fragilis was cultured in a minimum defined media and EMVs were obtained by differential centrifugation, ultracentrifugation, and filtration. The purified EMVs were observed by both transmission electron microscopy (TEM) and immunoelectron microscopy (IM). To identify EMV constituent proteins, EMVs were separated by 1D SDS-PAGE and proteomic analysis of proteins sized 35 kDa to approximately 65 kDa was performed using mass spectrometry (MALDI-TOF MS). FINDINGS: TEM micrographs proved the presence of spherical vesicles and IM confirmed the presence of Bfp60 protein on their surface. Mass spectrometry identified 23 proteins with high confidence. One of the proteins from the B. fragilis EMVs was identified as an enolase P46 with a possible lyase activity. MAIN CONCLUSIONS: Although the Bfp60 protein was not detected by proteomics, α-enolase P46 was found to be present in the EMVs of B. fragilis. The P46 protein has been previously described to be present in the outer membrane of B. fragilis as an iron-regulated protein.

Diagnostic value of α-enolase expression and serum α-enolase autoantibody levels in lung cancer / Valor diagnóstico da expressão de α-enolase e dos níveis séricos de autoanticorpos contra α-enolase no câncer de pulmão

ABSTRACT Objective: To investigate the diagnostic value of α-enolase (ENO1) and serum ENO1 autoantibody levels in lung cancer. Methods: Immunohistochemistry staining and ELISA were performed to detect ENO1 expression in lung tissue and serum ENO1 autoantibody levels, respectively. Results: The expression of ENO1 was higher in lung cancer tissues than in benign lung disease tissues (p < 0.001). The proportion of lung cancer samples expressing ENO1 was not significantly different among the various pathological classification groups. The proportion of samples expressing ENO1 was higher in lung cancer patients in stages I/II than in those in stages III/IV (χ2 = 5.445; p = 0.018). The expression of ENO1 in lung cancer tissues was not associated with age, gender, or smoking history. Serum ENO1 antibody levels were significantly higher in the lung cancer group than in the benign lung disease and control groups (p < 0.001). The differences among the pathological classification groups were not statistically significant. Serum ENO1 antibody levels were also in lung cancer patients in stages I/II than in those in stages III/IV (p < 0.01). Serum ENO1 antibody levels were not associated with age, gender, or smoking history in lung cancer patients. The ROC curve representing the diagnosis of lung cancer based on ENO1 antibody levels had an area under the curve of 0.806. Conclusions: Our results suggest that high levels of ENO1 are associated with the clinical stage of lung cancer and that ENO1 expression and its serum autoantibody levels show diagnostic value in lung cancer.

RESUMO Objetivo: Investigar o valor diagnóstico da α-enolase (ENO1) e dos níveis séricos de autoanticorpos contra ENO1 no câncer de pulmão. Métodos: Marcação imuno-histoquímica e ELISA foram realizados para detectar a expressão de ENO1 no tecido pulmonar e os níveis séricos de autoanticorpos contra ENO1, respectivamente. Resultados: A expressão de ENO1 foi maior nos tecidos de câncer de pulmão que nos tecidos de doença pulmonar benigna (p < 0,001). Não houve diferença significativa entre os diversos grupos de classificação patológica quanto à proporção de amostras de câncer de pulmão que expressaram ENO1. A proporção de amostras que expressaram ENO1 foi maior nos pacientes com câncer de pulmão nos estágios I/II que naqueles com câncer de pulmão nos estágios III/IV (χ2 = 5,445; p = 0,018). Não houve relação entre a expressão de ENO1 em tecidos de câncer de pulmão e idade, sexo ou histórico de tabagismo. Os níveis séricos de anticorpos contra ENO1 foram significativamente maiores no grupo câncer de pulmão que nos grupos doença pulmonar benigna e controle (p < 0,001). As diferenças entre os grupos de classificação patológica não foram estatisticamente significativas. Os níveis séricos de anticorpos contra ENO1 foram também significativamente maiores nos pacientes com câncer de pulmão nos estágios I/II que naqueles com câncer de pulmão nos estágios III/IV (p < 0,01). Nos pacientes com câncer de pulmão, não houve relação entre os níveis séricos de anticorpos contra ENO1 e idade, sexo ou histórico de tabagismo. A curva ROC do diagnóstico de câncer de pulmão baseado nos níveis de anticorpos contra ENO1 apresentou área sob a curva = 0,806. Conclusões: Nossos resultados sugerem que há relação entre níveis elevados de ENO1 e o estágio clínico do câncer de pulmão e que a expressão de ENO1 e os níveis séricos de autoanticorpos contra ENO1 têm valor diagnóstico no câncer de pulmão.

Selective Reaction Monitoring Mass Spectrometry for Quantitation of Glycolytic Enzymes in Postmortem Brain Samples.

Patients with psychiatric disorders exhibit dysfunctions in peripheral and central metabolism. This may be a root cause of impaired neuronal function, manifested as changes in mood, behavior, and cognitive capabilities in patients suffering with these conditions. Here we describe a selective reaction monitoring mass spectrometry (SRM-MS)-based targeted proteomic protocol for precise simultaneous quantitation of three glycolytic enzymes in postmortem brain tissue extracts. The SRM-MS approach has several advantages in terms of sensitivity, reproducibility, and reduced sample consumption, compared to traditional MS methods.

Histogenesis of Abrikossoff tumour of the oral cavity.

BACKGROUND: Abrikossoff or granular cell tumour (GCT) is a relatively rare neoplasia, benign in most of the cases. It may occur in any part of the human body, but it has an oral location in 70% of the cases. Its origin has been discussed for decades, and it is not yet definitively determined. Immunohistochemical techniques suggest its origin in the Schwann cells, while more recent studies with new markers indicate an origin related to neuroendocrine cells. OBJECTIVE: Contribute to the clarification of histogenesis of oral Abrikossoff tumour studying immunohistochemical marking of 11 oral Brazilian cases. MATERIALS AND METHODS: Samples of tissues from the oral mucosa, tongue and lips placed in paraffin blocks, from eleven patients with a histopathological diagnosis of benign GCT were studied. Four different anti-serums (S-100, vimentin, PGP9.5 and ENE) were used for immunoperoxydase technique. RESULTS: A clear positivity for S-100 protein and vimentin was observed, with markers indicating origin from the Schwann cells. Less intense positivity was found in some cases, for ENE and PGP9.5, which suggests a neuroendocrine origin. CONCLUSIONS: The results obtained suggest an origin from Schwann cells, but also arise the possibility of neuroendocrine origin. New methods and more specific immunohistochemical markers are needed to elucidate the origin of the Abrikossoff tumour.

Enolasa específica de neurona en dos recién nacidos con depresión ligera al nacer / Neuron- specific enolase in two newborns presenting with moderate depression at birth

La enolasa específica de neurona es una isoenzima que se vierte al torrente sanguíneo después de un episodio de daño neuronal. En los procesos de hipoxia neonatal estos valores enzimáticos en suero suelen estar alterados. El objetivo del presente artículo fue estudiar la enolasa específica de neurona en suero de 2 recién nacidos con Apgar bajo y determinar si, en el seguimiento durante un año, dichos pacientes presentaban trastornos del desarrollo psicomotor. MÉTODOS. Se tomaron muestras de suero al momento del nacimiento y a las 72 h siguientes. Se determinaron los niveles de enolasa específica de neurona por un método inmunoenzimá tico de tipo ELISA. Cada muestra fue evaluada por el método de reacción en cadena de la polimerasa para citomegalovirus, en el Instituto de Inmunología de Wuersburg (Alemania). También se cuantificaron anticuerpos contra citomegalovirus de clase IgM e IgG, en el Laboratorio de Neuroquímica de la Universidad Georg August de Goettingen (Alemania). Se recogieron los datos clínicos de interés de cada recién nacido y al a±o se citaron a estos pacientes y se les realizó un examen físico para evaluar su neurodesarrollo. RESULTADOS. Las cifras de enolasa estuvieron incrementadas tanto al nacimiento como a las 72 h, con anticuerpos anticitomegalovirus de clase IgG que fueron transferidos de la madre a través de la placenta. No se encontró presencia de este virus en el momento del nacimiento. En el examen físico y neurológico realizado al año se constató que los niños evolucionaban satisfactoriamente hasta esa fecha. CONCLUSIONES. Se recomienda extender el estudio hasta los 3 años de vida y aumentar el número de pacientes estudiados, con énfasis en aquellos casos cuyo Apgar es menor de 5 a los 5 min del nacimiento.

Neuron-specific Enolase of is an isoenzyme present in blood stream after a neuronal damage episode. In processes of neonatal hypoxia, these enzymatic values in serum may be altered. The aim of present paper was to study Enolase specific of neuron in the serum of two newborns with low Apgar score and to determine if, during a one year follow-up, such patients presented disorders of psychomotor development. METHODS: We took serum samples at birth and at 72 hours. Levels of Enolase specific of neuron by immunoenzymatic method type ELISA were determined. Each sample was assessed by polymerase chain reaction (PCR) to cytomegalovirus in Wuersburg Institute of Immunology (Germany). Also, we quantified antibodies to IgM and IgG cytomegalovirus in Neurochemistry Institute of Georg August of Goettingen University (Germany). Interesting clinical data of each newborn were collected and after a year, all these patients were cited for a physical examination to evaluate tits neurodevelopment. RESULTS: Enolase figures were increased both at birth and at 72 hours, with IgG anticytomegalovirus antibodies from mother through placenta. At birth there was not presence of this virus. At physical and neurological examination performed at 1 year it was possible to confirm that children evolved adequately until now. CONCLUSIONS: Study must to be prolonged until 3 years of life, and to increase number of study patients, emphasizing on those with an Apgar score lower than 5 at 5 minutes post-birth.

Nerve fibers in uterosacral ligaments of women with deep infiltrating endometriosis.

UNLABELLED: Endometriosis is a disease of high prevalence and enigmatic origin. One aspect not yet clarified is the relationship between endometriosis and nerve tissue. OBJECTIVE: To evaluate, by immunohistochemistry, the presence of sympathetic and parasympathetic nerve fibers in the uterosacral ligament and adjacent connective tissue in women with deep pelvic endometriosis and women without endometriosis. DESIGN: Cross-sectional study (Canadian Task Force II). SETTING: University Hospital. Obstetrics and Gynecology Department, Santa Casa Medical School, São Paulo, Brazil. METHODOLOGY: We selected 49 patients, 20 of them with deep endometriosis in the uterosacral ligament and 29 patients without endometriosis. Secondary antibodies to NSE (pan-neuronal marker), NPY (that identifies sympathetic nerve fibers), and VIP (that identifies parasympathetic nerve fibers) were used for the immunohistochemistry analyses. RESULTS: The immunohistochemical staining by the NSE antibody was positive in 40% of cases of women with endometriosis and in 20.7% of patients without endometriosis (non-significant). The immunohistochemical staining by the NPY antibody was positive in 60% of patients with endometriosis and in 20.7% of the control group (p=0.005), while staining by the VIP antibody was 60% in patients with endometriosis and 13.8% in patients without endometriosis (p=0.001). CONCLUSION: Immunoexpression of NPY (sympathetic fibers) and VIP (parasympathetic fibers) is higher in women with deep pelvic endometriosis than in women without endometriosis.

Oral alveolar soft-part sarcoma: review of literature and case report with immunohistochemistry study for prognostic markers.

OBJECTIVE: To describe a case of alveolar soft-part sarcoma (ASPS) affecting the tongue of a child and to study prognostic imunohistochemical markers for the disease. STUDY DESIGN: Tissue sections were incubated with primary antibodies reactive to neuron-specific enolase (NSE), vimentin, desmin, S-100 protein, cytokeratins AE1-AE3, EMA, neurofilament, synaptophysin, and muscle-specific actin (MSA), and for prognostic markers, including Ki-67, p53, bcl-2, bax, and nm23. RESULTS: Histologically, the tumor showed a proliferation of large polygonal cells with PAS-positive diastase-resistant intracytoplasmatic material, arranged in an alveolar growth pattern. Diffuse positive reaction for neuron specific enolase (NSE), focal reactivity for desmin and S-100 protein, strong positivity for nm23 and bax, but weak reaction for p53 and Ki-67 were found. No bcl-2-positive cells were noted. CONCLUSION: These immunohistochemical findings may reflect the less aggressive behavior of ASPS in oral tissues.

Constant light exposure causes dissociation in gonadotrophin secretion and inhibits partially neuroendocrine differentiation of Leydig cells in adult rats.

The objective of this work was to study the changes that occur in the Leydig cells of rats exposed to continuous light. The laboratory rat is considered a non-photoperiodic species because exposure to short photoperiod has little or no effect on the reproductive status. However, exposure of adult female rats to constant light induces polycystic ovaries, indicating that extreme changes in the photoperiod affect the reproductive function seriously. Adult male rats were placed under continuous light conditions for a duration of 15 weeks. After this period, the animals were killed and testicles were dissected and processed by routine histologic protocols. Follicle-stimulating hormone (FSH) and luteinizing hormone (LH) serum levels were determined by radioimmunoassay (RIA). The visualization of antigens was achieved by the streptavidin-peroxidase immunohistochemical method. Antibodies against chromogranin A, S-100 protein, P substance, synaptofisin, neurofilament protein-200, gliofibrillary acidic protein and neurone-specific enolase were used. The mean LH serum concentration was significantly lower, while the mean FSH level was significantly higher in treated animals. The expression of S-100, NSE, CrA, SP and SYN was significantly lower in treated animals. In conclusion, the constant light exposure acting directly at the pituitary level decreases LH secretion. The increased FSH secretion may be due to a partial reduction of the negative androgen feedback in the pituitary gland. Moreover, the constant light exposure affects the expression of some immunomarkers in Leydig cells, possibly because of the changes found in the gonadotrophin level and feedback mechanism.

Marcadores de diferenciação endócrina para diagnóstico do carcinoma de pequenas células extra-pulmonares / Markers of endocrine differentiation in extra-pulmonary small cell carcinoma

Introducao: estudos recentes confirmam a natureza endocrina do carcinoma indiferenciado de pequenas celulas do pulmao bem como das suas apresentacoes extra-pulmonares. O objetivo do estudo foi comparar a utilidade da Protein Gene Peptide com a enzima enolase neuronio especifica (NSE) o hormonio adrenocorticotropico e a calcitonina na diferenciacao endocrina do carcinoma indiferenciado de celulas extra-pulmonares. Materiais e metodos: analise de dados primarios nao publicados. A tecnica imunohistoquimica utilizada foi atraves do metodo peroxidase-antiperoxidase utilizando anticorpos PGP, NSE, ACTH e calcitonina...

Desmoplastic astrocytoma of infancy: a case report with histopathologic and immunohistochemistry profile.

The clinicopathological features and immunohistochemistry profile of desmoplastic cerebral astrocytoma of infancy are discussed in a 4 month old male infant who presented with an increasing head circumference more pronounced in the last two weeks prior to admission to the University Pediatric Hospital. This is a rare tumor that occurs in infants within the first two years of life and it is characterized by a massive, often cystic, supratentorial lesion usually in the frontoparietal region. It has a biphasic histologic pattern with an astrocytic and desmoplastic component and a good prognosis after total or near total surgical resection. This patient represents the first case of desmoplastic cerebral astrocytoma of infancy diagnosed in the Puerto Rico Medical Center.

Non increased neuron-specific enolase concentration in cerebrospinal fluid during first febrile seizures and a year follow-up in pediatric patients.

Febrile seizures are the commonest acute neurological disorder of early childhood. Studies suggested that febrile seizures are previous acute events from a more serious neurological problem. Due to neuron-specific enolase is generally accepted as a marker for neuropathological processes in the brain, 16 pediatric patients were studied during their first seizures and a year after it. Neuron-specific enolase in cerebrospinal fluid and blood were analysed by an immune enzyme assay. Non pathological neuron-specific enolase values were obtained in both periods in the group of patients. There were no significative differences when paired series statistics test was performed with 95% of confidence. Neuron-specific enolase appears not to be a marker for febrile seizures because its concentration not be increased in cerebrospinal fluid in this group of patients.

Non increased neuron-specific enolase concentration in cerebrospinal fluid during first febrile seizures and year follow-up in pediatric patients

Febrile seizures are the commonest acute neurological disorder of early childhood. Studies suggested that febrile seizures are previous acute events from a more serious neurological problem. Due to neuron-specific enolase is generally accepted as a marker for neuropathological processes in the brain, 16 pediatric patients were studied during their first seizures and a year after it. Neuron-specific enolase in cerebrospinal fluid and blood were analysed by an immune enzyme assay. Non pathological neuron-specific enolase values were obtained in both periods in the group of patients. There no significative differences when paired series statistics test was performed with 95 per cent of confidence. Neuron-specific enolase appears not to be a marker for febrile seizures because its concentration not be increased in cerebrospinal fluid in this group of patients.

Leishmania major-like parasite, a pathogenic agent of cutaneous leishmaniasis in Paraguay.

Leishmania parasites isolated from two patients with cutaneous leishmaniasis from geographically different localities in Paraguay have been characterized by enzyme electrophoresis (zymodeme) and digestion profiles of kinetoplast DNA with restriction enzymes (schizodeme). Both Paraguayan isolates showed identical zymodeme profiles to each other using 14 enzymes (glutamic pyruvate transaminase, glutamic oxaloacetic transaminase, enolase, fumarate hydratase, glucose phosphate isomerase, glucose-6-phosphate dehydrogenase, malate dehydrogenase, malic enzyme, mannose phosphate isomerase, nucleoside phosphorylase, peptidase-D, 6-phosphogluconate dehydrogenase, phosphoglucomutase, and pyruvate kinase). Although two Paraguayan isolates showed different zymodeme profiles from those of six Leishmania reference strains of Old and New World Leishmania species, they showed identical zymodeme profiles to those of an L. major-like parasite from Ecuador. These observations were confirmed by schizodeme analysis using three restriction endonucleases (Msp I, Hae III, and Taq I). These results indicate that Leishmania parasites isolated in Paraguay are identified as an L. major-like parasite, and it is necessary to consider the existence of L. major-like parasites when classifying Leishmania isolates from the New World.

Malignant neurocytic tumor.

Two patients, 14 and 46 years of age, presented with diffuse, rapidly growing intracerebral tumors leading to death 6 1/2 and 9 1/2 months, respectively, after diagnosis. Histological examination showed sheets of moderate-sized tumor cells with clear cytoplasm and central nuclei interrupted by delicate arciform vasculature, an appearance distinctly different from that of neuroblastoma. Malignant features were present in the form of significant nuclear pleomorphism, numerous mitotic figures, and small foci of necrosis with some suggestion of adjacent pseudo-palisading in one case. Ultrastructural examination showed neuronal differentiation, including prominent neuritic processes, microtubules, dense-core neurosecretory-type granules, and synaptic bouton-like structures containing small, empty-appearing synaptic-type vesicles and synapse-like membrane "thickenings." Immunohistochemistry showed focal immunopositivity for synaptophysin, neurofilaments, neuron-specific enolase, and S100 protein. Immunoreactivity for glial fibrillary acidic protein (GFAP) was found at the margins of the tumors adjacent to some intratumoral blood vessels and in some tumor cells. These tumors seem to occupy a nosological "middle ground" between neuroblastoma and central neurocytoma.

Neuron-specific enolase is a useful marker of neuroendocrine origin in pheochromocytoma cell culture

Neuron-specific enolase (NSE) has been used as a marker for neuroendocrine tumors either in immunocytochemical studies or in serum measurements. ln this paper NSE levels were determined in cultured pheochromocytoma cells to test whether it is also a useful marker ín cell culture of tumors derived from neuroendocrine system. Cultured pheochromocytoma cells came from a primary explant and were grown in RPMI supplemented with 20 per cent fetal calf serum, 100 mug/mL ampicillin and 100 mug/mL streptomycin. NSE was measured in culture medium and cell homogenates. Samples from different pheochromocytoma cultures were analyzed and compared to normal cultured fibroblast cells derived from human skin. NSE was measured by a commercially available radioimmunoassay kit. NSE levels were higher in cell homogenates as compared to those in culture medium, reaching levels as high as 6-fold in the former in TE cell line (26.46 ng/mL and 4.39 ng/mL respectively. Serial NSE measurements in culture medium from TE cell line evidenced decreasing values in subsequential subcultures (from 9.24 ng/mL during primary explant to 1.7 ng/ml. in the 10th subculture). In cultured normal fibroblasts, NSE levels in cultured media were definitely lower than those obtained from pheochromocytoma cultures. These preliminary data suggest that NSE may be a useful marker of neuroendocrine derived tumors, such as pheochromocytoma, in culture. Thus, the simplicity and availability of NSE radioimmunoassay provides an alternative to catecholamine measurement to better characterize pheochromocytoma cell lines in culture, with the advantage of faster results at lower costs.

Pulmonary endocrine cells of Aymara Indians from the Bolivian Andes.

INTRODUCTION: There is evidence to suggest that life at high altitude causes changes in the population of pulmonary endocrine cells, possibly because of exposure to chronic hypoxia. A study was made of the populations of pulmonary endocrine cells in three Aymara Indians and three Mestizos of La Paz (3600 m), Bolivia, which were compared with those in four white lowlanders. METHODS: Pulmonary endocrine cells were immunolabelled for neurone specific enolase and their two major secretory products, gastrin releasing peptide and calcitonin, and their numbers expressed per cm2 of tissue section. RESULTS: No differences in morphology, number, content, or distribution of immunoreactive cells were found when the native highlanders were compared with the lowlanders. CONCLUSIONS: If chronic hypoxia as such exerts an influence on human pulmonary endocrine cells it was not apparent in this morphological study. There was no increase in gastrin releasing peptide containing pulmonary endocrine cells, such as have previously been seen in patients with pulmonary hypertension characterised by plexogenic pulmonary arteriopathy. This may be due to the fact that in plexogenic pulmonary arteriopathy there is free migration of smooth muscle cells. Although three of the highlanders in this present study showed pulmonary vascular remodelling, this was in contrast only modest.