Analysis of Olig2 and YKL-40 expression: a clinicopathological/immunohistochemical study for the distinction between subventricular zone II and III glioblastomas.

Glioblastomas (GBs) are the most common and lethal primary brain tumors in the adults. Glioblastomas originates either from astrocytes that have accumulated mutations and de-differentiated or from neural stem cells within the subventricular zone (SVZ) in close contact with the vasculature. Recently, several studies have hypothesized that gliomagenesis occurs in perivascular niches with highly invasive peripheral proliferating zones. The purpose of our study was to investigate the pathological and clinical significance of Olig2 and YKL40 immunoexpression in 152 GBs in relationship to the SVZ II and III. Olig2 expressions were successfully detected in 12 (15.58%) of 77 SVZ type II GBs and 16 (21.3%) of 75 SVZ type III GBs, respectively. YKL-40 expression was observed in 45 (58.4%) of 77 SVZ type II GBs and in 17 (22.6%) of 75 SVZ type III GBs, respectively. Stepwise multivariate Cox proportional hazards models were used, and the prognostic factors to significantly impact OS were: PFS < 54 weeks (HR: 5.86; CI: 3.02-11.33; p = 0.00); radiotherapy (HR: 0.34; CI: 0.18-0.60; p = 0.00); radio- and chemotherapy (HR: 0.05; CI: 0.03-0.10; p = 0.0), and YKL-40+ GBs (HR: 1.61; CI: 1.28-2.31; p = 0.01).

STAT3 inhibition reduces toxicity of oncolytic VSV and provides a potentially synergistic combination therapy for hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is a refractory malignancy with a high mortality and increasing worldwide incidence rates, including the United States and central Europe. In this study, we demonstrate that a specific inhibitor of signal transducer and activator of transcription 3 (STAT3), NSC74859, efficiently reduces HCC cell proliferation and can be successfully combined with oncolytic virotherapy using vesicular stomatitis virus (VSV). The potential benefits of this combination treatment are strengthened by the ability of NSC74859 to protect primary hepatocytes and nervous system cells against virus-induced cytotoxicity, with an elevation of the VSV maximum tolerated dose in mice. Hereby we propose a strategy for improving the current regimen for HCC treatment and seek to further explore the molecular mechanisms underlying selective oncolytic specificity of VSV.

Biclonal gammopathies: Retrospective study of 47 patients. / Gammapatías biclonales: estudio retrospectivo de 47 pacientes.

OBJECTIVES: Biclonal gammopathies are characterized by the clonal proliferation of plasma cells or their B-lymphoid progenitors and are associated with the production of abnormal immunoglobulins (M proteins or paraproteins). There are no known studies that have analyzed this disease in Spain. We studied the underlying diseases, characteristics of paraproteins and the evolution of a series of patients with biclonal gammopathy. MATERIAL AND METHODS: We reviewed clonal gammopathies at the Department of Immunology of Hospital Puerta de Hierro in Madrid, between 1970 and 2011, selecting those patients with biclonal gammopathy in one reading. We collected data on the patient's epidemiology, underlying disease, associated diseases, therapies and paraprotein and immunoglobulin levels. RESULTS: Of the 1626 cases of clonal gammapathies, 47 were biclonal gammopathy (2.89%). The median follow-up was 2 years. The main associated condition was biclonal gammopathies of undetermined significance (BGUS). The most common paraprotein combination was IgG-IgG. Upon conducting a second paraprotein reading, 81% of the patients had lost at least 1 monoclonal component. A third of the patients had not undergone treatment. CONCLUSIONS: Biclonal gammopathy are fundamentally associated with biclonal gammopathies of undetermined significance. No biclonal gammopathies of undetermined significance evolved to a malignant disease. In a high percentage of patients, at least 1 of the 2 clonal components disappeared, sometimes spontaneously.

Serum IL-12 is increased in Mexican obese subjects and associated with low-grade inflammation and obesity-related parameters.

Interleukin-(IL-) 12 has been recently suggested to participate during development of insulin resistance in obese mice. Nevertheless, serum IL-12 levels have not been accurately determined in overweight and obese humans. We thus studied serum concentrations of IL-12 in Mexican adult individuals, examining their relationship with low-grade inflammation and obesity-related parameters. A total of 147 healthy individuals, 43 normal weight, 61 overweight, and 43 obese subjects participated in the study. Circulating levels of IL-12, tumor necrosis factor-alpha (TNF- α ), leptin, insulin, glucose, total cholesterol, and triglyceride were measured after overnight fasting in all of the study subjects. Waist circumference and body fat percentage were recorded for all the participants. Serum IL-12 was significantly higher in overweight and obese individuals than in normal weight controls. Besides being strongly related with body mass index (r = 0.5154), serum IL-12 exhibited a significant relationship with abdominal obesity (r = 0.4481), body fat percentage (r = 0.5625), serum glucose (r = 0.3158), triglyceride (r = 0.3714), and TNF- α (r = 0.4717). Thus, serum levels of IL-12 are increased in overweight and obese individuals and show a strong relationship with markers of low-grade inflammation and obesity in the Mexican adult population. Further research is needed to understand the role of IL-12 in developing obesity-associated alterations in humans.

Large deletions and splicing-site mutations in the STK11 gene in Peutz-Jeghers Chilean families.

Peutz-Jeghers syndrome (PJS) is an autosomal dominant disorder characterized by mucocutaneous melanocytic macules, gastrointestinal hamartomatous polyposis and an increased risk of various neoplasms. Germline mutations in the serine/threonine kinase 11 (STK11) gene have been identified as a cause for PJS. The aim of this study was to characterize the genotype of Chilean PJS patients. Mutation screening of 13 patients from eight PJS families was performed using a single strand conformation polymorphism analysis, DNA sequencing and multiplex ligation-dependent probe amplification assay. The breakpoints of the genomic rearrangements were assessed by a long-range polymerase chain reaction and sequencing. The results revealed the existence of seven different pathogenic mutations in STK11 gene in seven unrelated families, including three point mutations and four large genomic deletions. Three of these point mutations (43%, 3/7) may be considered as novel. Our results showed that a germline mutation is present in STK11 in 88% of probands fulfilling the diagnostic criteria of PJS. In this study, the combination of two different experimental approaches in the screening of the STK11 in PJS, led to a higher percentage of mutation detection.

High yield synthesis, purification and characterisation of the RNase L activators 5'-triphosphate 2'-5'-oligoadenylates.

Upon viral infection, double-stranded viral RNA is detected very early in the host cell by several cellular 2'-5' oligoadenylate synthetases, which synthesize 2'-5' adenylate oligonucleotides that activate the cellular RNase L, firing an early primary antiviral response through self and non-self RNA cleavage. Transfecting cells with synthetic 2'-5' adenylate oligonucleotides activate RNase L, and thus provide a useful shortcut to study the early steps of cellular and viral commitments into this pathway. Defined 2'-5' adenylate oligonucleotides can be produced in vitro, but their controlled synthesis, purification, and characterisation have not been reported in detail. Here, we report a method suitable to produce large amounts of 2-5As of defined lengths in vitro using porcine OAS1 (pOAS) and human OAS2 (hOAS). We have synthesized a broad spectrum of 2-5As at the milligram scale and report an HPLC-purification and characterisation protocol with quantified yield for 2-5A of various lengths.

Acyclic phosphonate nucleotides and human adenylate kinases: impact of a borano group on alpha-P position.

Adenylate kinases are involved in the activation of antiviral drugs such as the acyclic phosphonates analogs PMEA and (R)PMPA. We examine the in vitro phosphorylation of PMEA and PMPA bearing a borano- or a H- group on the phosphorus atom. The alpha-borano or alpha-H on PMEA and PMPA were detrimental to the activity of recombinant human AMP kinases 1 and 2. Docking PMEA to the active site of AMP kinase 1 indicated that the borano group may prevent two conserved critical Arg interactions with the alpha-phosphate, resulting in substrate bad positioning.

Hernia hiatal gigante: reporte de un caso / Giant hiatal hernia: a case report

La hernia de hiato es una situación patológica que se da cuando parte del estómago se introduce hacia el tórax. El esófago entra en el abdomen desde el tórax a través de un agujero o hiato que se encuentra en el diafragma. El estómago protruye a través de este hiato debilitado y produce ardores (pirosis) y dolor torácico. La persistencia en el tiempo de esta hernia, puede provocar una metaplasia de la mucosa esofágica dando al lugar al esófago de Barret el cual puede malignizar produciéndose cáncer de esófago. (1) La hernia de hiato es relativamente frecuente, afectando hasta un 20 por ciento de la población. Del total de pacientes con el trastorno, cerca del 10 por ciento son asintomáticos, dependiendo del grado de protrución estomacal y de que tanto esté afectado el esfinter esofágico inferior (EEI). Un 40 por ciento de las hernias de hiato son hernias deslizantes, en las que el EEI protruye conjuntamente con una porción del estómago y solo un 5 por ciento son paraesofágicas, en la que solo una porción del estómago se hace intratorácica mientras tanto que el EEI permanece intraabdominal. Los síntomas más comunes de una hernia de hiato incluyen pirosis, dolor torácico, disfagia, palpitaciones y ocasionalmente regurgitación o reflujo gastroesofágico. El diagnóstico de una hernia hiatal comienza con el éxamen físico por razón de la sintomatología. Los estudios radiográficos y la endoscopia digestiva demuestran la hernia hiatal y ayudan a descartar otras causas de molestias digestivas altas. (2) Se recomienda un procedimiento quirúrgico conocida como fundoplicación de Nissen, (3) Cuando los síntomas causados por una hernia de hiato son tan severas que pueden resultar en lesiones al esófago o incluso cáncer del esófago. (6) Se trata de lactante mayor de seis meses de edad quien es natural de la localidad y procedente de Cordero, quien inicia enfermedad actual el día de hoy en la madrugada según refiere la madre, caracterizado por vómitos en número incontables.

Cierre diferido en onfalocele no roto: reporte de un caso / Close deferred not broken in omphalocele: a case report

El término onfalocele fue definido por Benson en 1949 y se atribuye a Paré su descripción inicial en la primera mitad del siglo XVI. Se trata de una herniación de las vísceras abdominales que están cubiertas por un saco formado por peritoneo, membrana amniótica y gelatina de Warthon y que se continúa con el cordón umbilical. (1) Esta herniación es consecuencia de una alteración en la formación de la pared abdominal anterior del embrión. El defecto se sitúa soempre en la región umbilical y puede ser de tamaño variable, considerándose un onfalocele gigante cuando el defecto supera los 10 cm. Generalmente contiene asas de intestino delgado, y en los de mayor tamaño, el hígado. (2) La presencia de otras malformaciones asociadas no es rara y puede condicionar el pronóstico del recién nacido. El 25 por ciento presentan una cardiopatía congénita, siendo más frecuente la tetralogía de Fallot. La frecuencia de esta entidad varía de unas regiones a otras y oscila entre 1/6.000 hasta 1/60.000 de los recién nacidos vivos, incluyendo los casos de gastrosquitis. Puede diagnosticarse mediante las ecografías prenatales a partir del cuarto mes, y en un futuro serán subsidiarias de corregirse mediante cirugía fetal. (3) Existe controversia en relación a la necesidad de una cesárea programada para el nacimiento. La cesárea previene la lesión de las vísceras herniadas, pero el parto vaginal logra una mayor madurez del feto. En general los niños afectados de onfaloceles grandes deben nacer mediante cesárea electiva, pero los que presentan onfaloceles pequeños pueden nacer mediante parto vaginal. Con independencia de la decisión adoptada es importante que los neonatólogos y cirujanos pediátricos estén alertados del nacimiento para realizar una adecuada valoración del recién nacido lo antes posible. El tratamiento tiene por misión reintroducir las vísceras en la cavidad abdominal restaurando la integridad de la misma antes de que se produzca la contaminación del contenido herniario.

High-yield production of short GpppA- and 7MeGpppA-capped RNAs and HPLC-monitoring of methyltransfer reactions at the guanine-N7 and adenosine-2'O positions.

Many eukaryotic and viral mRNAs, in which the first transcribed nucleotide is an adenosine, are decorated with a cap-1 structure, (7Me)G5'-ppp5'-A(2'OMe). The positive-sense RNA genomes of flaviviruses (Dengue, West Nile virus) for example show strict conservation of the adenosine. We set out to produce GpppA- and (7Me)GpppA-capped RNA oligonucleotides for non-radioactive mRNA cap methyltransferase assays and, in perspective, for studies of enzyme specificity in relation to substrate length as well as for co-crystallization studies. This study reports the use of a bacteriophage T7 DNA primase fragment to synthesize GpppAC(n) and (7Me)GpppAC(n) (1 < or = n < or = 9) in a one-step enzymatic reaction, followed by direct on-line cleaning HPLC purification. Optimization studies show that yields could be modulated by DNA template, enzyme and substrate concentration adjustments and longer reaction times. Large-scale synthesis rendered pure (in average 99%) products (1 < or = n < or = 7) in quantities of up to 100 nmol starting from 200 nmol cap analog. The capped RNA oligonucleotides were efficient substrates of Dengue virus (nucleoside-2'-O-)-methyltransferase, and human (guanine-N7)-methyltransferase. Methyltransfer reactions were monitored by a non-radioactive, quantitative HPLC assay. Additionally, the produced capped RNAs may serve in biochemical, inhibition and structural studies involving a variety of eukaryotic and viral methyltransferases and guanylyltransferases.

Rabdomiosarcoma embrionario hepatico: a propósito de un caso / Hepatic embrionary Rhabdomysarcoma: presentation of a case

El Rabdomiosarcoma es el sarcoma de tejidos blandos de origen musculoesqueléticos más frecuente en niños menores de 15 años y uno de los más comunes en adolescentes y adultos jóvenes. (1) Las áreas del cuerpo más comunes donde puede alojarse este tumor son la cabeza, el cuello, la vejiga, la vagina, los brazos, las piernas y el tronco. Aunque también puede encontrarse en la próstata, el oído medio y el sistema de conductos biliares. (2) En los niños con rabdomiosarcoma embrionario, esta anomalía se encuentra en el cromosoma 11, mientras el alveolar en los cromosomas 2 y 13. (1) Se reconocen tres subtipos: embrionario, alveolar y pleomórfico. El embrionario se observa en niños y adolescentes menores de 15 años y se localiza principalmente en la cabeza y en el cuello, tracto urogenital, retroperitoneo y extremidades. Los síntomas pueden incluir una masa visible o palpable que puede ser doloroso o no, parestesia, dolor. (2) El diagnóstico se establece a través del examen físico, historia médica completa, así como estudios imagenológicos, biopsia y punción de medula ósea. (3) El tratamiento específico será dado por la localización del tumor primario y el estadio del tumor, en algunos pacientes se administra quimioterapia preoperatorio en un intento de reducir la extensión de la intervención y de preservar órganos vitales. El pronóstico se relaciona con la edad del paciente, sitio de origen, resecabilidad. (4)

Caracterización del factor de transferencia a partir de leucocitos no inducidos con virus Sendai / Characterization of the transfer factor obtained from Sendai virus non-induced leucocytes

El uso extensivo en la práctica clínica del factor de transferencia (FT) se ha visto limitado por aspectos relacionados con su caracterización bioquímica y la garantía de seguridad, pues este producto debre cumplir con las regulaciones que aseguran la inocuidad de los productos hemoderivados. El hecho de que su proceso de obtención dependa del procesamiento de donaciones de sangre con menos de 24 horas de almacenamiento limita su disponibilidad. Con el objetivo de incrementar la cantidad de unidades de FT disponibles para el tratamiento clínico, se realizó la caracterización de varios lotes de FT obtenidos de donaciones de hasta 48 horas; el producto fue pasteurizado, como garantía de la inactivación de posibles contaminantes virales. La evaluación de sus propiedades bioquímicas y de sus efectos biológicos permitió demostrar la identidad y potencia de este producto. Se demostró además su inocuidad mediante ensayos preclínicos de tolerancia local y toxicidad.

Systemic administration of phenyl-N-tert-butylnitrone protects the retina from light damage.

PURPOSE: This study was conducted to test the hypothesis that phenyl-N-tert-butylnitrone (PBN), a spin-trapping agent known to cross the blood-brain barrier and protect the brain from ischemia-reperfusion injury, is incorporated into the retina after intraperitoneal injection and protects photoreceptor cells from the damaging effects of constant visible light. METHODS: Albino rats were injected intraperitoneally with PBN (aqueous solution) or water, or were not injected, and then were placed in constant light (2700 lux) for 24 hours. The incorporation of PBN into the retina was determined by high-performance liquid chromatography. Electroretinograms (ERGs) were recorded before light treatment and 1 and 15 days after the cessation of exposure to constant light. Eyes were taken for histology at each time point and outer nuclear layer (ONL) thickness determined. RESULTS: PBN was incorporated into the retina after intraperitoneal injection. Both control (water-injected and uninjected) groups exposed to constant light maintained only 28% of ONL thickness and 20% of retinal function, compared with the unexposed control group. In contrast, the PBN-treated animals maintained 80% of ONL thickness and exhibited 87% of retinal function. CONCLUSIONS: PBN protects the albino rat retina from the damaging effects of constant light stress. That light-induced and hereditary retinal degenerations share certain morphologic hallmarks and follow a similar apoptotic mechanism of degeneration raises the possibility of pharmacologic therapy for hereditary and environmentally induced neurodegenerative disorders.

Regulation of type II phosphatidylinositol phosphate kinase by tyrosine phosphorylation in bovine rod outer segments.

Type II phosphatidylinositol phosphate kinase (PIPKII) is an enzyme responsible for the synthesis of phosphatidylinositol-4,5-bisphosphate (PI-4,5-P(2)) from phosphatidylinositol-5-phosphate (PI-5-P). In this study, we demonstrate the presence of PIPKII alpha in bovine photoreceptor rod outer segments (ROS) and the involvement of tyrosine phosphorylation in the regulation of its activity. PIPKII activity in bovine ROS was verified by the preferential conversion of synthetic dipalmitoyl PI-5-P to PI-4,5-P(2), lack of effect of phosphatidic acid, inhibition by heparin, immunoreaction with an anti-PIPKII alpha antibody on Western blots, and immunocytochemical localization in bovine and rat ROS by anti-PIPKII alpha. Immunoprecipitates of bovine ROS with the anti-PIPKII alpha antibody possessed PIPK enzymatic activity and preferentially used PI-5-P as substrate for PI-4,5-P(2) biosynthesis. The activity of PIPKII was greatly increased under conditions favoring tyrosine phosphorylation in ROS, and PIPKII activity was immunoprecipitated with anti-phosphotyrosine (anti-PY) antibodies from tyrosine phosphorylated ROS. Preincubation of ROS with tyrosine kinase inhibitors almost abolished the kinase activity in the anti-PY immunoprecipitates. Immunoblot analysis showed that PIPKII alpha was present in anti-PY immunoprecipitates from phosphorylated ROS but not from nonphosphorylated controls. We conclude that PIPKII alpha is present in ROS and that its activity is regulated by tyrosine phosphorylation.

Association of the tyrosine phosphatase SHP-2 with transducin-alpha and a 97-kDa tyrosine-phosphorylated protein in photoreceptor rod outer segments.

Increasing evidence indicates that tyrosine phosphorylation, controlled by the concerted action of tyrosine kinases and protein tyrosine phosphatases (PTPs), plays important roles in retinal photoreceptor rod outer segments (ROS). We characterized PTP activity in isolated bovine ROS that is significantly inhibited by orthovanadate. Incubating ROS in the presence of exogenous Mg2+, ATP, and orthovanadate dramatically enhanced the tyrosine phosphorylation of several endogenous proteins. SHP-2, a PTP with two SH2 domains, was identified in ROS by immunoblot analysis and was found to associate with ROS membranes. Immunocytochemistry showed localization of SHP-2 in photoreceptor outer segments and possibly in the outer plexiform, inner nuclear, and inner plexiform cell layers of the retina as well. SHP-2 associated with transducin-alpha and a 97-kDa tyrosine-phosphorylated protein in ROS, suggesting the formation of a multimeric signaling complex. Based on its association with transducin-alpha and a 97-kDa protein, SHP-2 may regulate the tyrosine phosphorylation of endogenous proteins, including transducin-alpha, and may play a significant role in a novel signaling pathway in photoreceptors.

Detección por técnicas de biología molecular de Mycoplasma hominis y Ureaplasma urealyticum en muestras urogenitales / Detection by molecular biology techniques of Mycoplasma hominis and Ureaplasma urealyticum in urogenital samples

Mycoplasma homonis y Ureaplasma urealyticum están estrechamente relacionadas con enfermedades del tracto urogenital como pielonefritis, uretritis no-gonocóccica, cálculos urinarios, epididimitis, inflamaciones pélvicas, infertilidad, abortos y fiebre post-parto; en recién nacidos también pueden causar neumonías y meningitis. Estas bacterias pueden ser diagnosticadas por diferentes métodos. En este trabajo utilizamos la hibridación de ácidos nucléicos y la reacción en cadena de la polimerasa para analizar 22 muestras de pacientes con diferentes síntomas urogenitales, en busca de micoplasmas y ureaplasmas. Como resultado obtuvimos 10 muestras positivas y 12 negativas. Entre las muestras positivas se identificaron 2 como Mycoplasma hominis, 2 como Ureaplasma urealyticum y 6 con ambas especies. los resultados obtenidos por las técnicas moleculares fueron comparados con los métodos de referencia, encontrándose en 18 muestras resultados coincidentes, mientras que en 4 los resultados fueron discordantes, siendo esta diferencia estadísticamente no significativa

Vigilancia de la sensibilidad a antimicrobianos de streptococcus beta-hemolíticos, aislados entre 1982 y 1987 / Antimicrobial sensibility surveillance of betahemolytic streptococcus isolated between 1982 and 1987

El objetivo principal de este trabajo, fue determinar la resistencia o sensibilidad a antimicrobianos de uso habitual, en cepas de Streptococcus beta-hemolíticos grupos A, C y G aisladas de diversas localizaciones, en Santiago, entre los años 1982 y 1987