Sphingosine Kinase 1 urothelial expression is increased in patients with neurogenic detrusor overactivity

Objectives: To evaluate the expression of sphingosine kinase 1 (SPK1) in the bladder wall in patients with neurogenic lower urinary tract dysfunction and its association with clinical, urodynamic and pathological features. Materials and Methods: The expression of SPK1 was studied in bladder wall specimens obtained from cystectomy using immunohistochemistry in ten patients with spinal cord injury (n=8) or multiple sclerosis (n=2) with urodynamically proven neuropathic bladder dysfunction, and in controls (n=5). Inflammation and fibrosis were analysed with histological criteria and SPK1 expression was determined by individual immunohistochemical staining. Results: Significant increased SPK1 urothelial immunoreactivity was shown in patients compared to control group (p=0.03). By contrast, SPK1 immunoreactivity in patients was significantly decreased in the sub-urothelium, muscles and nerves, p=0.02; 0.01 and 0.003, respectively. Patients with neurogenic detrusor overactivity (NDO) had higher SPK1 urothelium expression than those without any DO (p=0.04). Conclusions: SPK1 is expressed in the human bladder wall, specifically the urothelium, in bladder specimens from patients with NDO. The role of SPK1 in the pathophysiology of NDO needs further elucidation.

Assessment of primary health care: health professionals’ perspective / Evaluación de la atención primária em la salud: vision de los profissionales de la salud / Avaliação da Atenção Primária à Saúde: visão dos profissionais de saúde

Objective To assess primary health care attributes of access to a first contact, comprehensiveness, coordination, continuity, family guidance and community orientation. Method An evaluative, quantitative and cross-sectional study with 35 professional teams in the Family Health Program of the Alfenas region, Minas Gerais, Brazil. Data collection was done with the Primary Care Assessment Tool - Brazil, professional version. Results Results revealed a low percentage of medical experts among the participants who evaluated the attributes with high scores, with the exception of access to a first contact. Data analysis revealed needs for improvement: hours of service; forms of communication between clients and healthcare services and between clients and professionals; the mechanism of counter-referral. Conclusion It was concluded that there is a mismatch between the provision of services and the needs of the population, which compromises the quality of primary health care.


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Objetivo Evaluar la atención primaria de salud a través de las cualidades: Acesso de Primero Contacto, Intregidad, Coordinación, Longitudinalidad, Orientación Familiar, Orientación Comunitaria. Método Se trata de una evaluación cuantitativa y estudio transverso con 35 equipos de profesionales de la Estrategia de Salud de la Familia, de región de Alfenas, Minas Gerais, Brasil. Para recopilar los datos, se utilizó el Instrumento de Evaluación de la Atención Primaria - Brasil , la versión Professional. Resultados Los datos revelaron un bajo porcentaje de especialistas médicos en Atencion Primaria de Salud. Los participantes evaluó las calidades con puntajes altos, con la excepción de Acceso Primero Contacto. El análisis de datos reveló una mejora necesidades: horarios de apertura de los servicios; las formas de comunicación entre el usuario y el servicio y entre el usuario y el profesional, la remissión y consulta. Conclusión Existe un desajuste entre la oferta de servicios y las necesidades de la población, lo que compromete la calidad de la Atención Primaria de Salud.
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Objetivo Avaliar a Atenção Primária à Saúde por meio dos atributos: Acesso de Primeiro Contato, Integralidade, Coordenação, Longitudinalidade, Orientação Familiar, Orientação Comunitária. Método Estudo avaliativo, quantitativo e transversal, realizado com 34 profissionais de equipes da Estratégia de Saúde da Família da microrregião de Alfenas, Minas Gerais, Brasil. Para a coleta de dados, foi utilizado o Primary Care Assessment Tool – Brasil, versão profissionais. Resultados Os dados revelaram baixo percentual de profissionais médicos especialistas em Atenção Primária à Saúde. Os participantes avaliaram os atributos com altos escores, com exceção do Acesso de Primeiro Contato. A análise dos dados revelou necessidades de aperfeiçoamento: o horário de funcionamento dos serviços; as formas de comunicação entre usuário e serviço, e entre usuário e profissionais; o mecanismo de contrarreferência. Conclusão Existe um descompasso entre a oferta de serviços e as necessidades da população que compromete a qualidade da Atenção Primária a Saúde.
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Síndrome de Sengers: comunicación de dos casos en Chile / Sengers syndrome: report of two cases in chile

El Síndrome de Sengers es una enfermedad mitocondrial autosómica recesiva, producida por mutación del gen de la Acil-Glicerol Kinasa (AGK), recientemente descubierto. Se caracteriza por cataratas congénitas bilaterales, miocardiopatía hipertrófica y acidosis láctica. Puede tener miopatía esquelética leve, intolerancia al ejercicio y desarrollo mental normal. Los pacientes fallecen tempranamente debido a falla cardíaca. Dada la alta letalidad, lo infrecuente de este síndrome y la presencia de un diagnóstico confirmado, se presenta el caso clínico de 2 hermanos chilenos, fallecidos por la enfermedad, que se presentaron con el cuadro característico de cataratas congénitas bilaterales, miocardiopa-tía hipertrófica y acidosis láctica. El mayor, se operó las cataratas a los 4 meses de edad y falleció a la edad de 13 meses debido a falla cardíaca severa refractaria y falla orgánica múltiple, descompensado por una infección respiratoria. El menor se diagnosticó a los 3 meses de edad y se le confirmó la mutación del gen de AGK en Alemania. Se decidió no operarlo de las cataratas dado el mal pronóstico vital. Presentó progresión de la miocardiopatía hipertrófica y falleció súbitamente a los 8 meses de edad.

Senger's Syndrome is a recessive autosomal mitochondrial disease due to a recently discovered mutation of the Acyl-Glycerol Kinase (AGK) gen,. It is characterized by congenital bilateral cataracts, progressive hypertrophic cardiomyopathy and lactic acidosis. It may present skeletal myopathy, exercise intolerance and usually normal mental development. Patients die early in life due to heart failure. The clinical cases of two brothers with a confirmed diagnosis of Senger's syndrome are reported. The older brother was operated on for cataracts at the age of 4 months and he died when he was 13 months old due to severe refractory heart failure and multi-organ failure, decompensated by a respiratory infection. The younger brother was diagnosed at 3 month of age and the AGK gene mutation was confirmed in Germany. Cataracts were not operated on due to the the patient's extremely poor prognosis. He had progressive hypertrophic cardiomyopathy and died suddenly at 8 month of age.

The subsidiary GntII system for gluconate metabolism in Escherichia coli: Alternative induction of the gntV gene

Two systems are involved in the transport and phosphorylation of gluconate in Escherichia coli. GntI, the main system, consists of high and low-affinity gluconate transporters and a thermoresistant gluconokinase for its phosphorylation. The corresponding genes, gntT, gntU and gntK at 76.5 min, are induced by gluconate. GntII, the subsidiary system, includes IdnT and GntV, which duplicate activities of transport and phosphorylation of gluconate, respectively. Gene gntV at 96.8 min is divergently transcribed from the idnDOTR operon involved in L-idonate metabolism. These genetic elements are induced by the substrate or 5-keto-D-gluconate. Because gntV is also induced in cells grown in gluconate, it was of interest to investigate its expression in this condition. E. coli gntK, idnOokan mutants were constructed to study this question. These idnO kan-cassete inserted mutants, unable to convert gluconate to 5-keto-D-gluconate, permitted examining gntV expression in the absence of this inducer and demonstrating that it is not required when the cells grow in gluconate. The results suggest that E. coli gntV gene is alternatively induced by 5-keto-D-gluconate or gluconate in cells cultivated either in idonate or gluconate. In this way, the control of gntV expression would seem to be involved in the efficient utilization of these substrates.

Búsqueda de mutaciones en el gen UL 97 asociadas a resistencia a ganciclovir en citomegalovirus obtenidos desde muestras biológicas de pacientes chilenos / Presence of mutations associated with ganciclovir resistance in cytomegalovirus UL 97 gene

Background: Long term use of ganciclovir (GCV) is associated with acquired resistance to it. Ninety percent of the responsible mutations occur in cytomegalovirus (CMV) UL 97 gene. Aim: To search for these mutations, comparing nucleotide sequences of CMV-positive samples from post transplant and immunocompromised patients receiving GCV, with sequences of CMV isolates obtained from subjects not exposed to the drug. Patients and Methods: Codons 440 to 465 of gene UL 97, in-cluding the most common mutations causing resistance to GCV, were amplifed in 33 plasma samples from patients exposed to GCV and in 15 urine samples of newborns. Both populations and their nucleotide sequences were compared with the prototype strain CMV AD169. Results: Samples of exposed patients had multiple mutations but only one had a mutation associated with clinical resistance (M460I). Eight subjects had the D605E mutation, whose role in resistance is controversial. The remaining 150 mutations were silent mutations. Conclusions: A low frequency of mutations associated with CMV resistance to GCV was found in these exposed and unexposed samples. These mutations may refect coexistence of multiple genetic variants of CMV. The absence of clinical expression of resistance, even with these mutations, can be explained by the use of GCV for a shorter lapse than that associated with the appearance of resistance.

Gluconate as suitable potential reduction supplier in Corynebacterium glutamicum: Cloning and expression of gntP and gntK in Escherichia coli

Corynebacterium glutamicum is widely used in the industrial production of amino acids. We have found that this bacterium grows exponentially on a mineral médium supplemented with gluconate. Gluconate permease and Gluconokinase are expressed in an inducible form and, 6-phosphogluconate dehydrogenase, although constituvely expressed, shows a 3-fold higher specific level in gluconate grown cells than those grown in fructose under similar conditions. Interestingly, these activities are lower than those detected in the strain Escherichia coli Ml-8, cultivated under similar conditions. Additionally, here we also confirmed that this bacterium lacks 6-phosphogluconate dehydratase activity. Thus, gluconate must be metabolized through the pentose phosphate pathway. Genes encoding gluconate transport and its phosphorylation were cloned from C. glutamicum, and expressed in suitable E. coli mutants. Sequence analysis revealed that the amino acid sequences obtained from these genes, denoted as gntP and gntK, were similar to those found in other bacteria. Analysis of both genes by RT-PCR suggested constitutive expression, in disagreement with the inducible character of their corresponding activities. The results suggest that gluconate might be a suitable source of reduction potential for improving the efficiency in cultures engaged in amino acids production. This is the first time that gluconate specific enzymatic activities are reported in C. glutamicum.

Identificación molecular de enzimas modificantes de aminoglucósidos en cepas de Enterococcus spp. aisladas en hospitales de la Octava Región de Chile / Molecular identification of aminoglycoside-modifying enzymes among strains of Enterococcus spp. isolated in hospitals of the VIII Region of Chile

Background: Infectious diseases produced by Enterococcus spp, must be treated with a synergistic combination between a penicillin and an aminoglycoside. High level resistance to aminoglycosides is a serious therapeutic problem, since it predicts the loss of synergistic activity of this antimicrobial combination. Aim: To investigate the presence of genes encoding aminoglycoside-modifying enzymes (AMEs) among strains of Enterococcus spp with high level of resistance to aminoglycosides. Material and methods: The genes encoding some of the AMEs were investigated among 305 aminoglycoside-resistant strains of Enterococcus spp isolated in hospitals of the VIII region of Chile, by dot blot hybridization and Polymerase Chain Reaction (PCS). Results: High level resistance to some aminoglycosides was observed in 104 strains (34.1 percent) and 93 of these harbored at íeast one of the genes encoding the investigated AMEs. Three genes were detected: aac(6)Ie-aph(2")Ia (14.8 percent) encoding for the enzyme AAC(6)Ie-APH(2")Ia (resistance to all aminoglycosides, except streptomycin); aph(3)IIIa (26 percent), and ant(6)la (28.5 percent) encoding for the phosphorylating enzymes APH(3)Ilia (resistance to kanamycin, amikacin and neomycin), and ANT(6)-la (resistance only to streptomycin), respectively. None of the strains harbored the gene ant (4) which encode for the enzyme ANT (4). Conclusion: The low frequency of strains harbouring the bifunctional enzyme (<15 percent), conferring an extended resistance profile to aminoglycosides, allows us to propose the empirical use of aminoglycoside-aminocyclitols, associated to a penicillin, in the treatment of serious infections produced by species of enterococci.

Tipificación molecular de Brucella abortus cepa 19 y su aplicación al control de biológicos / Molecular characterization of Brucella abortus strain 19 and its application for controlling biologics

Brucella abortus es el agente etiológico de la brucelosis bovina. La cepa 19, utilizada en la elaboración de vacunas, puede ser identificada a través de una deleción en la región eri asociada con la sensibilidad al eritritol. Se optimizó un ensayo de PCR para caracterizar específicamente esta cepa. El método que describimos es un procedimiento rápido para identificar B. abortus y simultáneamente diferenciar la cepa 19 de otras cepas de B. abortus biovar 1. Hemos aplicado este ensayo para la detección de la cepa 19 en vacunas contra la brucelosis bovina elaboradas en Argentina. Los resultados indican que este método podría ser útil para el seguimiento de las cepas madres y semillas utilizadas en la producción industrial de esta vacuna. Esta metodología también contribuiría a la reducción del riesgo de la infección adquirida en el laboratorio y podría aplicarse como prueba de rutina para confirmar la presencia de B. abortus en vacunas no relacionadas.

Brucella abortus is the etiological agent of bovine brucellosis. The strain 19 used in vaccine elaboration can be identified through a deletion in the eri region associated with its susceptibility to erythritol. We optimized a PCR assay for specific characterization of this strain. The method described here is a rapid procedure that enables identification of B. abortus, and simultaneous differentiation of the strain 19 from other B. abortus biovar 1 strains. We applied the assay to detect the strain 19 in vaccines against B. abortus produced in Argentina. The results show this method could be used to follow vaccine seed cultures of this strain. The methodology could also contribute to reduce the risk of a laboratory-acquired infection and could be of great help as a routine test for confirmation of B. abortus in non related vaccines.

Cyclic AMP increases the survival of ganglion cells in mixed retinal cell cultures in the absence of exogenous neurotrophic molecules, an effect that involves cholinergic activity

Natural cell death is a well-known degenerative phenomenon occurring during development of the nervous system. The role of trophic molecules produced by target and afferent cells as well as by glial cells has been extensively demonstrated. Literature data demonstrate that cAMP can modulate the survival of neuronal cells. Cultures of mixed retinal cells were treated with forskolin (an activator of the enzyme adenylyl cyclase) for 48 h. The results show that 50 æM forskolin induced a two-fold increase in the survival of retinal ganglion cells (RGCs) in the absence of exogenous trophic factors. This effect was dose dependent and abolished by 1 æM H89 (an inhibitor of protein kinase A), 1.25 æM chelerythrine chloride (an inhibitor of protein kinase C), 50 æM PD 98059 (an inhibitor of MEK), 25 æM Ly 294002 (an inhibitor of phosphatidylinositol-3 kinase), 30 nM brefeldin A (an inhibitor of polypeptide release), and 10 æM genistein or 1 ng/ml herbimycin (inhibitors of tyrosine kinase enzymes). The inhibition of muscarinic receptors by 10 æM atropine or 1 æM telenzepine also blocked the effect of forskolin. When we used 25 æM BAPTA, an intracellular calcium chelator, as well as 20 æM 5-fluoro-2'-deoxyuridine, an inhibitor of cell proliferation, we also abolished the effect. Our results indicate that cAMP plays an important role controlling the survival of RGCs. This effect is directly dependent on M1 receptor activation indicating that cholinergic activity mediates the increase in RGC survival. We propose a model which involves cholinergic amacrine cells and glial cells in the increase of RGC survival elicited by forskolin treatment

Is aging related to cell signalling alterations in amphibian oocytes?

O armazenamento de oócitos de sapo Bufo arenarum diminuiu sua habilidade para serem fertilizados "in vitro". A estimulaçäo com carbachol em "oócitos jovens" mostrou uma hidrólisis persistente de fosfatidilinositol 4,5 bifosfato (PIP2) näo foram hidrolizados até um nível significativo. Estes resultados e os menores níveis de 32P-fosfoinositideos achados nos "oócitos envelhecidos" num tempo zero de estimulaçäo, condordam com uma diminuiçäo nas atividades das quinases de fosfoinositideos e da fosfolipase C, como consequência de uma hidrólisis näo específica de fosfoinositidos que ocorreria durante o armazenamento

Molecular mechanisms of insulin resistance

1. Insulin stimulates tyrosine phosphorylation of the insulin receptor and of an endogenous substrate of approximately 185 kDa (insulin receptor substrate 1 or IRS-1). IRS-1 fulfills the criteria of a direct substrate of the insulin receptor, and tyrosine phosphorylation of IRS-1 leads to another step in insulin action, i.e., an association of phosphorylated IRS-1 with the enzyme PI3-kinase activating this enzyme. Using antipeptide antibodies to insulin receptor, to IRS-1 and to PI 3-kinase together with anti-phosphotyrosine antibodies it is possible to study insulin-stimulated insulin receptor phosphorylation, IRS-1 phosphorylation and the association/activation of IRS-1/PI 3-kinase. 2. In this review we describe alterations in these three early steps of insulin action after binding in animal models of insulin resistance, i.e., streptozotocin-induced diabetes (STZ diabetes), fasting, spontaneously hypertensive rats, the ob/ob mice, dexamethasone-treated rats, and the chronic effect of insulin on Fao cells in culture. 3. In states of insulin resistance with hypoinsulinemia (STZ diabetes and fasting) there is an increase in these early steps of insulin action. In animal models of insulin resistance with hyperinsulinemia there is a decrease in these steps of insulin action, indicating molecular post-receptor defects. Since we could reproduce the decrease in these three early steps of insulin action in cells in culture by chronic treatment with insulin, we postulate that these defects may be a consequence of the hyperinsulinemia of these animals.