RESUMO
INTRODUCTION: The IgV(H) mutational status of B-cell chronic lymphocytic leukemia (B-CLL) is of prognostic value. Expression of ZAP-70 in B-CLL is a surrogate marker for IgV(H) unmutated (UM). As determination of IgV(H) mutational status involves a methodology currently unavailable for most clinical laboratories, it is important to have available a reliable technique for ZAP-70 estimation in B-CLL. Flow cytometry (FC) is a convenient technique for this purpose. However, there is still no adequate way for data analysis, which would prevent the assignment of false positive or negative expression. METHODS: We have modified the currently most accepted technique, which uses the ratio of the mean fluorescent index (MFI) of B-CLL to T cells. The MFI for parallel antibody isotype staining is subtracted from the ZAP-70 MFI of both B-CLL and T cells. We validated this technique comparing the results obtained for ZAP-70 expression by FC with those obtained with quantitative PCR for the same patients. RESULTS: We applied the technique in a series of 53 patients. With this modification, a better correlation between ZAP-70 expression and IgV(H) UM was obtained. CONCLUSIONS: Thus, the MFI ratio B-CLL/T cell corrected by isotype is a reliable analysis technique to estimate ZAP-70 expression in B-CLL.
Assuntos
Citometria de Fluxo , Isotipos de Imunoglobulinas , Leucemia Linfocítica Crônica de Células B/metabolismo , Proteína-Tirosina Quinase ZAP-70/metabolismo , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Humanos , Cadeias Pesadas de Imunoglobulinas/genética , Isotipos de Imunoglobulinas/metabolismo , Região Variável de Imunoglobulina/genética , Leucemia Linfocítica Crônica de Células B/diagnóstico , Leucemia Linfocítica Crônica de Células B/genética , Mutação , Proteína-Tirosina Quinase ZAP-70/genéticaRESUMO
Antecedentes: ha quedado demostrado que la respuesta a la agresión quirúrgica es directamente proporcional al tamaño de la incisión abdominal y al tiempo de exposición de la cavidad abdominal durante la operación. La tendencia actual en el mundo es minimizar el trauma terapéutico de los pacientes, el dolor y recuperación postoperatorios, mejorar resultados estéticos. La colecistectomía por orificios naturales apunta a una cirugía sin dejar cicatrices visibles. La cirugía por un solo puerto está demostrando ser una alternativa posible. Objetivo: presentar nuevo instrumental para cirugía laparoscópica en colecistectomía por un solo puerto. Lugar de aplicación: Hospital Centro de Salud " Zenon J. Santillán", San Miguel de Tucumán, argentina. Diseño: Retrospectivo. Población: 30 pacientes. Método: De diciembre 2008 a mayo 2009, 30 pacientes con litiasis vesicular fueron tratados mediante colecistectomía por un solo puerto con nuevo instrumental para cirugía laparoscópica, evaluando dificultad de técnica quirúrgica y tiempo operatorio. Resultados: 8 varones, 22 mujeres, edad promedio 35,5 años (r:19-55). Tiempo de seguimiento medio postoperatorio: 15 días (r: 1-30). Tiempo operatorio medio: 50 minutos (r:40-85). Hospitalización promedio: 1 día (r:12 h-2 días). Conversiones: 0%. Morbilidad: 1 hematoma de herida (3,3%). Mortalidad:0. Conclusiones: Nuestra experiencia inicial es alentadora y muestra la colecistectomía por un solo puerto con este nuevo instrumental como una alternativa simple y viable de la colecistectomía laparoscópica convencional, con resultados funcionales similares y mejores resultados estéticos.
Assuntos
Humanos , Masculino , Adulto , Feminino , Pessoa de Meia-Idade , Colecistectomia Laparoscópica/instrumentação , Estudos Retrospectivos , Instrumentos CirúrgicosRESUMO
Antecedentes: ha quedado demostrado que la respuesta a la agresión quirúrgica es directamente proporcional al tamaño de la incisión abdominal y al tiempo de exposición de la cavidad abdominal durante la operación. La tendencia actual en el mundo es minimizar el trauma terapéutico de los pacientes, el dolor y recuperación postoperatorios, mejorar resultados estéticos. La colecistectomía por orificios naturales apunta a una cirugía sin dejar cicatrices visibles. La cirugía por un solo puerto está demostrando ser una alternativa posible. Objetivo: presentar nuevo instrumental para cirugía laparoscópica en colecistectomía por un solo puerto. Lugar de aplicación: Hospital Centro de Salud " Zenon J. Santillán", San Miguel de Tucumán, argentina. Diseño: Retrospectivo. Población: 30 pacientes. Método: De diciembre 2008 a mayo 2009, 30 pacientes con litiasis vesicular fueron tratados mediante colecistectomía por un solo puerto con nuevo instrumental para cirugía laparoscópica, evaluando dificultad de técnica quirúrgica y tiempo operatorio. Resultados: 8 varones, 22 mujeres, edad promedio 35,5 años (r:19-55). Tiempo de seguimiento medio postoperatorio: 15 días (r: 1-30). Tiempo operatorio medio: 50 minutos (r:40-85). Hospitalización promedio: 1 día (r:12 h-2 días). Conversiones: 0%. Morbilidad: 1 hematoma de herida (3,3%). Mortalidad:0. Conclusiones: Nuestra experiencia inicial es alentadora y muestra la colecistectomía por un solo puerto con este nuevo instrumental como una alternativa simple y viable de la colecistectomía laparoscópica convencional, con resultados funcionales similares y mejores resultados estéticos.(AU)
Assuntos
Humanos , Masculino , Adulto , Feminino , Pessoa de Meia-Idade , Colecistectomia Laparoscópica/instrumentação , Instrumentos Cirúrgicos , Estudos RetrospectivosRESUMO
A 30% decrease in osmolarity stimulated 3H-taurine, 3H-GABA and glutamate (followed as 3H-D-aspartate) efflux from rat hippocampal slices. 3H-taurine efflux was activated rapidly but inactivated slowly. It was decreased markedly by 100 microM 5-nitro-(3-phenylpropylamino)benzoic acid (NPPB) and 600 microM niflumic acid and inhibited strongly by tyrphostins AG18, AG879 and AG112 (25-100 microM), suggesting a tyrosine kinase-mediated mechanism. Hyposmolarity activated the mitogen-activated protein kinases (MAPK) extracellular-signal-related kinase-1/2 (ERK1/ERK2) and p38, but blockade of this reaction did not affect 3H-taurine efflux. Hyposmosis also activated phosphatidylinositide 3-kinase (PI3K) and its prevention by wortmannin (100 nM) essentially abolished 3H-taurine efflux. 3H-taurine efflux was insensitive to the protein kinase C (PKC) blocker chelerythrine (2.5 microM) or to cytochalasin E (3 microM). The release of 3H-GABA and 3H-D-aspartate occurred by a different mechanism, characterized by rapid activation and inactivation, insensitivity to NPPB, niflumic acid, tyrphostins or wortmannin. 3H-GABA and 3H-D-aspartate efflux was not due to external [NaCl] decrease, cytosolic Ca2+ increase or depolarization, or to reverse operation of the carrier. This novel mechanism of amino acid release may be mediated by Ca2+-independent exocytosis and modulated by PKC and actin cytoskeleton disruption, as suggested by its inhibition by chelerythrine and potentiation by 100 nM phorbol-12-myristate-13 acetate (PMA) and cytochalasin E. GABA and glutamate osmosensitive efflux may explain the hyposmolarity-elicited increase in amplitude of inhibitory and excitatory postsynaptic potentials in hippocampal slices as well as the hyperexcitability associated with hyponatraemia.
Assuntos
Ácido Glutâmico/metabolismo , Hipocampo/metabolismo , Concentração Osmolar , Taurina/metabolismo , Ácido gama-Aminobutírico/metabolismo , Ácido 4,4'-Di-Isotiocianoestilbeno-2,2'-Dissulfônico/farmacologia , Androstadienos/farmacologia , Inibidores da Angiogênese/farmacologia , Animais , Ácido Aspártico/química , Ácido Aspártico/metabolismo , Inibidores de Ciclo-Oxigenase/farmacologia , Citocalasinas/farmacologia , Inibidores Enzimáticos/farmacologia , Flavonoides/farmacologia , Ácido Glutâmico/química , Hipocampo/efeitos dos fármacos , Imidazóis/farmacologia , Técnicas In Vitro , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Ácido Niflúmico/farmacologia , Nitrobenzoatos/farmacologia , Fosfatidilinositol 3-Quinases/metabolismo , Inibidores de Fosfoinositídeo-3 Quinase , Proteína Quinase C/antagonistas & inibidores , Proteína Quinase C/metabolismo , Piridinas/farmacologia , Ratos , Taurina/química , Trítio/química , Trítio/metabolismo , Tirfostinas/farmacologia , Wortmanina , Ácido gama-Aminobutírico/químicaRESUMO
Weaning mass in southern elephant seals is highly variable, the heaviest pups being three times as heavy as the lightest ones. After weaning, pups undergo an extensive postweaning period in which they draw on their reserves. To quantify the energy expenditure during the postweaning period, changes in mass, body composition, and postweaning duration were measured in southern elephant seals at King George Island, South Shetland Islands, Antarctica. Overall, mean pup weaning mass was 154 +/- 26 kg (n=117) and did not differ between sexes. Mean minimum postweaning duration was 42.5 +/- 7.5 d. Heavier animals at weaning had lower mass-specific mass loss rates than lighter ones, and a faster depletion of body reserves was associated with a shorter postweaning period. The proportion of body mass represented by fat at weaning was 37% +/- 4% (n=47) and did not differ between sexes. Of these pups, 36 were recaptured after a mean period of 36 d after weaning. On average, total mass loss measured in these animals (39 kg) was composed of 39% water, 47% fat, and 12% protein. The composition of mass loss was not significantly different between sexes and was not related to weaning mass or total body energy reserves. However, fatter animals at weaning lost more fat per kilogram lost than thinner ones. Late in the fast, males and females appeared to be in a similar body condition. Nevertheless, the overall proportion of body mass represented by fat at this time was lower than that presented by the same animals at weaning. We estimated that during the postweaning period pups lost, on average, 30% of their mass at weaning. This comprised approximately 35% of the energy and 32% of the fat in the pup's body.
Assuntos
Composição Corporal/fisiologia , Metabolismo Energético/fisiologia , Focas Verdadeiras/fisiologia , Animais , Regiões Antárticas , Ilhas Atlânticas , Peso Corporal , Feminino , Masculino , Focas Verdadeiras/crescimento & desenvolvimento , Focas Verdadeiras/metabolismo , Fatores Sexuais , DesmameRESUMO
The activity of the pyridine nucleotide-independent lactate dehydrogenase (iLDH) was characterized in mitochondria isolated from the protist Euglena gracilis. The dissociation constants for L- and D-lactate were similar, but the V(max) was higher with the d isomer. A ping-pong kinetic mechanism was displayed with 2,4-dichlorophenol-indolphenol (DCPIP), or coenzyme Q(1), reacting as the second substrate with the modified, reduced enzyme. Oxamate was a competitive inhibitor against both L- and D-lactate. Oxalate exerted a mixed-type inhibition regarding L- or D-lactate and also against DCPIP. The rate of L-lactate uptake was partially inhibited by mersalyl and lower than the rate of dehydrogenation, which was mersalyl-insensitive. These data suggested that the active site of L-iLDH was orientated toward the intermembrane space. The following observations indicated the existence of two stereo-specific iLDH enzymes in the inner membrane of Euglena mitochondria: a greater affinity of the D-iLDH for both inhibitors, D-iLDH thermo-stability at 70 degrees C and denaturation of L-iLDH, opposite signs in the enthalpy change for the association reaction of the isomers to the enzyme, differential solubilization of both activities with detergents, and different molecular mass.
Assuntos
Euglena gracilis/enzimologia , L-Lactato Desidrogenase/metabolismo , Lactato Desidrogenases , Ácido Láctico/metabolismo , Mitocôndrias/enzimologia , Animais , Ligação Competitiva , Transporte Biológico , Estabilidade Enzimática , Cinética , L-Lactato Desidrogenase (Citocromo) , Proteínas de Membrana/metabolismo , Peso Molecular , Solubilidade , EstereoisomerismoRESUMO
Cardiovascular disease is one of the leading causes of morbidity and mortality in Mexico. We investigated the effects of omega-3 (n-3) and omega-6 (n-6) polyunsaturated fatty acids on the metabolic syndrome associated with cardiovascular disease in a high-sucrose-fed rat model. The metabolic syndrome-induced rats showed a significant increase in systolic blood pressure, serum insulin, nonfasting serum triglyceride and serum cholesterol levels. Experimental high-sucrose-fed animals received either a n-3 or n-6 enriched diet or a control diet during 6 weeks. Animals fed the n-3 enriched diet had a significant reduction in blood pressure and serum insulin and triglyceride levels. Serum triglyceride levels were also significantly reduced in the n-6-rich diet animals.
RESUMO
Brain adaptation to hyposmolarity is accomplished by loss of both electrolytes and organic osmolytes, including amino acids, polyalcohols and methylamines. In brain in vivo, the organic osmolytes account for about 35% of the total solute loss. This review focus on the role of amino acids in cell volume regulation, in conditions of sudden hyposmosis, when cells respond by active regulatory volume decrease (RVD) or after gradual exposure to hyposmotic solutions, a condition where cell volume remains unchanged, named isovolumetric regulation (IVR). The amino acid efflux pathway during RVD is passive and is similar in many respects to the volume-activated anion pathway. The molecular identity of this pathway is still unknown, but the anion exchanger and the phospholemman are good candidates in certain cells. The activation trigger of the osmosensitive amino acid pathway is unclear, but intracellular ionic strength seems to be critically involved. Tyrosine protein kinases markedly influence amino acid efflux during RVD and may play an important role in the transduction signaling cascades for osmosensitive amino acid fluxes. During IVR, amino acids, particularly taurine are promptly released with an efflux threshold markedly lower than that of K(+), emphasizing their contribution (possibly as well as of other organic osmolytes) vs inorganic ions, in the osmolarity range corresponding to physiopathological conditions. Amino acid efflux also occurs in response to isosmotic swelling as that associated with ischemia or trauma. Characterization of the pathway involved in this type of swelling is hampered by the fact that most osmolyte amino acids are also neuroactive amino acids and may be released in response to stimuli concurrent with swelling, such as depolarization or intracellular Ca(++) elevation.
Assuntos
Aminoácidos/metabolismo , Encéfalo/metabolismo , Encéfalo/citologia , Tamanho Celular , Concentração OsmolarRESUMO
The effect of antimycin, myxothiazol, 2-heptyl-4-hydroxyquinoline-N-oxide, stigmatellin and cyanide on respiration, ATP synthesis, cytochrome c reductase, and membrane potential in mitochondria isolated from dark-grown Euglena cells was determined. With L-lactate as substrate, ATP synthesis was partially inhibited by antimycin, but the other four inhibitors completely abolished the process. Cyanide also inhibited the antimycin-resistant ATP synthesis. Membrane potential was collapsed (<60 mV) by cyanide and stigmatellin. However, in the presence of antimycin, a H(+)60 mV) that sufficed to drive ATP synthesis remained. Cytochrome c reductase, with L-lactate as donor, was diminished by antimycin and myxothiazol. Cytochrome bc(1) complex activity was fully inhibited by antimycin, but it was resistant to myxothiazol. Stigmatellin inhibited both L-lactate-dependent cytochrome c reductase and cytochrome bc(1) complex activities. Respiration was partially inhibited by the five inhibitors. The cyanide-resistant respiration was strongly inhibited by diphenylamine, n-propyl-gallate, salicylhydroxamic acid and disulfiram. Based on these results, a model of the respiratory chain of Euglena mitochondria is proposed, in which a quinol-cytochrome c oxidoreductase resistant to antimycin, and a quinol oxidase resistant to antimycin and cyanide are included.
Assuntos
Euglena/metabolismo , Mitocôndrias/metabolismo , Trifosfato de Adenosina/biossíntese , Animais , Antimicina A/análogos & derivados , Antimicina A/farmacologia , Respiração Celular/efeitos dos fármacos , Ativação Enzimática/efeitos dos fármacos , Ácido Láctico/metabolismo , Metacrilatos , NADH Desidrogenase/metabolismo , Fosforilação Oxidativa/efeitos dos fármacos , Polienos/farmacologia , Cianeto de Sódio/farmacologia , Tiazóis/farmacologiaRESUMO
The oxidation of several metabolites in AS-30D tumor cells was determined. Glucose and glycogen consumption and lactic acid production showed high rates, indicating a high glycolytic activity. The utilization of ketone bodies, oxidation of endogenous glutamate, and oxidative phosphorylation were also very active: tumor cells showed a high respiration rate (100 ng atoms oxygen (min x 10(7) cells)(-1)), which was 90% oligomycin-sensitive. AS-30D tumor cells underwent significant intracellular volume changes, which preserved high concentrations of several metabolites. A high O(2) concentration, but a low glucose concentration were found in the cell-free ascites liquid. Glutamine was the oxidizable substrate found at the highest concentration in the ascites liquid. We estimated that cellular ATP was mainly provided by oxidative phosphorylation. These data indicated that AS-30D hepatoma cells had a predominantly oxidative and not a glycolytic type of metabolism. The NADH-ubiquinol oxido reductase and the enzyme block for ATP utilization were the sites that exerted most of the control of oxidative phosphorylation (flux control coefficient = 0.3-0.42).
Assuntos
Trifosfato de Adenosina/metabolismo , Carcinoma Hepatocelular/metabolismo , Neoplasias Hepáticas Experimentais/metabolismo , Ácido 3-Hidroxibutírico/metabolismo , Acetoacetatos/metabolismo , Animais , Divisão Celular/fisiologia , Respiração Celular/fisiologia , Citosol/metabolismo , Feminino , Glucose/metabolismo , Ácido Glutâmico/metabolismo , Glutamina/metabolismo , Glicogênio/metabolismo , Glicólise/fisiologia , Mitocôndrias/metabolismo , Oxirredução , Fosforilação Oxidativa , Ácido Pirúvico/metabolismo , Ratos , Ratos Wistar , Especificidade por Substrato , Células Tumorais CultivadasRESUMO
This paper reports Immunoglobulin M (IgM) levels in serum samples from eight female-pup pairs of southern elephant seals (Mirounga leonina), at King George Island, Antarctica. IgM levels were determined on sera obtained from sequential sampling throughout the suckling period (approximately 23 days). The IgM concentration in southern elephant seal serum was measured by single radial immunodiffusion on agarose plates. Female IgM levels (123.5-613.0 mg/dL, n = 8) were significantly higher than pup levels (5.9-123.6 mg/dL, n = 8). Both groups showed an increasing trend throughout the entire suckling period, with significant differences in relation to stages of lactation. Pup IgM levels on the first day of life (mean +/- SD, 7.6 +/- 2.9 mg/dL, n = 3) suggest that endogenous synthesis takes place before birth.
Assuntos
Imunoglobulina M/sangue , Focas Verdadeiras/imunologia , Animais , Animais Lactentes/imunologia , Regiões Antárticas , Feminino , Humanos , Imunidade Materno-Adquirida , Lactação/imunologia , Especificidade da EspécieRESUMO
Immunoglobulin A (IgA) levels in milk samples from southern elephant seals at King George Island, Antarctica are reported. IgA levels were determined throughout the suckling period (approximately 23 days). The IgA concentration in southern elephant seal milk was lower than in other mammals and, unlike most mammalian milk, was not high during early lactation. There was not a definite pattern in IgA levels, which fluctuated within narrow limits throughout the suckling period (mean +/- SD, 30.81 +/- 6.38 mg IgA/100 g milk). If IgG was present, its level was too low to be detected by the method used. This is the first evidence in Southern elephant seal of the possibility of transmission of passive immunity after birth involving secretion of IgA in the milk.
Assuntos
Animais Lactentes , Imunoglobulina A/análise , Leite/imunologia , Focas Verdadeiras/imunologia , Animais , Feminino , Imunodifusão , Imunoglobulina G/análise , LactaçãoAssuntos
Coração/fisiologia , Receptores Adrenérgicos alfa 1/fisiologia , Animais , Catecolaminas/fisiologia , Proteínas de Ligação ao GTP/efeitos dos fármacos , Proteínas de Ligação ao GTP/fisiologia , Coração/efeitos dos fármacos , Receptores Adrenérgicos alfa 1/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologiaRESUMO
The making and sealing of a tight junction (TJ) requires cell-cell contacts and Ca2+, and can be gauged through the development of transepithelial electrical resistance (TER) and the accumulation of ZO-1 peptide at the cell borders. We observe that pertussis toxin increases TER, while AIF3 and carbamil choline (carbachol) inhibit it, and 5-guanylylimidodiphosphate (GTPTs) blocks the development of a cell border pattern of ZO-1, suggesting that G-proteins are involved. Phospholipase C (PLC) and protein kinase C (PKC) probably participate in these processes since (i) activation of PLC by thyrotropin-1 releasing hormone increases TER, and its inhibition by neomycin blocks the development of this resistance; (ii) 1,2-dioctanoylglycerol, an activator of PKC, stimulates TER development, while polymyxin B and 1-(5-isoquinoline sulfonyl)-2-methyl-piperazine dihydrochloride (H7), which inhibit this enzyme, abolish TER. Addition of 3-isobutyl-1-methyl-xanthine, dB-cAMP or forskolin do not enhance the value of TER, but have just the opposite effect. Trifluoperazine and calmidazoline inhibit TER development, suggesting that calmodulin (CaM) also plays a role in junction formation. These results indicate that junction formation may be controlled by a network of reactions where G-proteins, phospholipase C, adenylate cyclase, protein kinase C and CaM are involved.
Assuntos
Calmodulina/fisiologia , Proteínas de Ligação ao GTP/fisiologia , Junções Intercelulares/fisiologia , Proteína Quinase C/fisiologia , Fosfolipases Tipo C/fisiologia , 1-(5-Isoquinolinasulfonil)-2-Metilpiperazina , Toxina Adenilato Ciclase , Animais , Cálcio/fisiologia , Carbacol/farmacologia , Células Cultivadas , Condutividade Elétrica/efeitos dos fármacos , Condutividade Elétrica/fisiologia , Células Epiteliais , Epitélio/metabolismo , Epitélio/fisiologia , Junções Intercelulares/metabolismo , Isoquinolinas/farmacologia , Rim/citologia , Rim/metabolismo , Rim/fisiologia , Potenciais da Membrana/efeitos dos fármacos , Potenciais da Membrana/fisiologia , Neomicina/farmacologia , Toxina Pertussis , Piperazinas/farmacologia , Polimixina B/farmacologia , Proteína Quinase C/antagonistas & inibidores , Trifluoperazina/farmacologia , Fatores de Virulência de Bordetella/farmacologiaRESUMO
A simple procedure that allowed the extraction and partial purification of a component of 65,000 mol.wt. from the surface of Leishmania braziliensis promastigotes is described. Iodinated cell surface membrane fractions were solubilized using Triton X-100 followed by Nonidet P-40. The macromolecular components were then freed of the detergents by passage of the extracts through a column of DE52 cellulose. The component of 65,000 mol.wt. was eluted from the column with 1 m NaCl. This component in whole parasites was immunoprecipitated by sera from patients with cutaneous, mucocutaneous, and diffuse cutaneous leishmaniasis and kala-azar. None of the major surface determinants reacted with sera from normal individuals with antibovine rabbit serum. The relevance and the possible applications on the immunoprophylaxis of the disease are briefly discussed.