RESUMO
The cell separator A 201 was a new platelet pheresis prototype designed to carry out a discontinuous apheresis while platelet-rich plasma (PRP) is sampled continuously. After donation, a platelet concentrate (PC) and a fresh frozen plasma was collected from the PRP by the plasma cell-C device. The PC contained 2.6 +/- 0.7 x 10(11) platelets with a leucocyte contamination of 3.9 +/- 3.9 x 10(8) and an erythrocyte contamination of 3.0 +/- 3.0 x 10(8). The fresh frozen plasma was nearly cell-free. Prior, during and after apheresis we analysed hemolysis and coagulation parameters. There was no evidence for hemolysis. The analysis of the coagulation factors and of the Thrombin-Antithrombin III-complex, however, gave a hint of an activation of coagulation.
Assuntos
Materiais Biocompatíveis , Plaquetoferese/instrumentação , Testes de Coagulação Sanguínea , Falha de Equipamento , Hemólise/fisiologia , Humanos , Contagem de Leucócitos , Teste de Materiais , Contagem de PlaquetasRESUMO
The ability of coronary endothelial cells in 14 day confluent cultures to metabolize glucose, palmitate, lactate and various amino acids was investigated. Under aerobic conditions, 99% of glucose, (5 mM) was degraded to lactate and only 0.04% was oxidized in the Krebs cycle. One percent of the glucose catabolized was directed into the hexose monophosphate pathway, but this fraction could be increased by 81% by 0.4 mM methylene blue. Glucose oxidation in the Krebs cycle was increased at glucose concentrations lower than 1 mM, or by the uncoupler 2,4-dinitrophenol. Oxidation to CO2 of palmitate (300 microM), lactate (1 mM), and glutamine (0.5 mM) was diminished in the presence of glucose (5 mM) by 80, 66, and 48%, respectively. These results demonstrate that coronary endothelial cells utilize exogenous glucose, at physiological concentration, predominantly for glycolytic energy production. The metabolic pattern is characteristic of the Crabtree effect. In these cells, glucose not only effectively suppresses the oxidation of the substrates lactate and palmitate, i.e. of substrates preferred by the whole heart, but also of glutamine, which is a major oxidative substrate for coronary endothelial cells. Absolute rates of substrate catabolism are low as compared to those of the beating heart indicating a low energy demand of coronary endothelial cells.
Assuntos
Vasos Coronários/metabolismo , Endotélio Vascular/metabolismo , Nucleotídeos de Adenina/análise , Aminoácidos/metabolismo , Animais , Células Cultivadas , DNA/análise , Glucose/metabolismo , Lactatos/metabolismo , Ácido Láctico , Masculino , Ácido Palmítico , Ácidos Palmíticos/metabolismo , Proteínas/análise , Ratos , Ratos EndogâmicosRESUMO
Energy metabolism of bovine cultured lens epithelial cells (CLEC) was compared to that of fresh bovine lens. CLEC contained high levels of ATP (44 nmol mg protein-1) and creatine phosphate (13 nmol mg protein-1). An ATP/ADP ratio of ten and a creatine phosphate/creatine ratio of two indicated the cells were in a well-energized state. ATP concentration in fresh epithelium was comparable to that of CLEC; however in the anterior cortex it was tenfold lower. In contrast to fresh lenses, CLEC were able to oxidize glucose, lactate and palmitic acid. Lactate was oxidized at the highest rate. In CLEC, 42% of the ATP generated by catabolizing glucose resulted from oxidative phosphorylation. Glucose (5 mM) was degraded to lactate and CO2 at a 2:1 ratio. The hexose monophosphate pathway accounted for two thirds of the CO2 produced. In the fresh whole bovine lens palmitate was not oxidized and lactate was oxidized to a lesser degree than in CLEC. Only one-tenth of the ATP generated by glucose catabolism in the fresh whole lens was derived from oxidative phosphorylation. This was also the case for a preparation of fresh epithelium, maintained in air and 100% oxygen, demonstrating that the preferential glycolytic catabolism of glucose in lens is not caused by limited oxygen diffusion. In the fresh bovine lens the epithelium accounted for one third of the glucose catabolism of the whole lens, even though it had only about 0.1% of its protein mass. In fresh human lenses, conversion of glucose into lactate was even more pronounced--the lactate/CO2 ratio was 73:1.(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Metabolismo Energético , Cápsula do Cristalino/metabolismo , Cristalino/metabolismo , Trifosfato de Adenosina/metabolismo , Idoso , Idoso de 80 Anos ou mais , Animais , Dióxido de Carbono/metabolismo , Bovinos , Células Cultivadas , Epitélio/metabolismo , Glucose/metabolismo , Humanos , Lactatos/metabolismoRESUMO
The response of endothelial energy metabolism to oxygen supply was studied in cultured coronary endothelial cells from the rat at defined PO2 levels between 0.1 and 100 Torr. In the presence of glucose (5 mM), endothelial respiration (4 nmol O2.min-1.mg protein-1) was independent of the exterior PO2 greater than 3 Torr; oxygen consumption was half maximal at 0.8 Torr. At 100 Torr, lactate production was 26 nmol.min-1.mg protein-1; the decrease of the PO2 to 0.1 Torr resulted in a 2.2-fold increase in lactate production. The contents of ATP, ADP, and AMP were 21, 4, and 2 nmol/mg protein, respectively; they remained constant for 2.5-h incubations at PO2 levels between 0.1 and 100 Torr. In the presence of palmitate (100 microM) plus glutamine (0.5 mM), oxygen consumption was 8 nmol.min-1.mg protein-1 at PO2 levels greater than 3 Torr, and the half-maximal rate was again observed at 0.8 Torr. Lactate production was negligible. At PO2 levels greater than 3 Torr, the cells remained well energized. Below 3 Torr, however, the adenine nucleotide contents rapidly declined. These results demonstrate that the oxygen demand of coronary endothelial cells is low compared with the beating myocardium. In the presence of glucose, aerobic glycolysis is pronounced and the Pasteur effect small. In severe hypoxia (PO2 less than 0.1 Torr) the energetic state remained stable. In the absence of glucose, the energetic state of coronary endothelial cells is sensitive to the exterior PO2 less than 3 Torr, declining concomitantly with the decrease in respiration.
Assuntos
Vasos Coronários/metabolismo , Endotélio Vascular/metabolismo , Metabolismo Energético , Hipóxia/metabolismo , Animais , Células Cultivadas , Metabolismo Energético/efeitos dos fármacos , Glucose/farmacologia , Glutamina/farmacologia , Consumo de Oxigênio/efeitos dos fármacos , Ácido Palmítico , Ácidos Palmíticos/farmacologiaRESUMO
The release of cytosolic enzymes from myocardial and endothelial cells in the anoxic-reoxygenated guinea pig heart was investigated. Isolated hearts were perfused with Tyrode solution in the Langendorff mode. Sixty-minute anoxic perfusion with or without glucose (5 mM) was followed by 15-min normoxic perfusion with glucose. The losses of purine-nucleoside phosphorylase (PNP) from endothelial cells and of lactate dehydrogenase (LDH) and creatine kinase (CK) from the mass of myocardial cells were determined. After 30-min anoxia, the release of LDH and CK but not of PNP increased. Reoxygenation after 60-min anoxia with glucose caused a partial recovery of tissue ATP but also an increase in leakage of LDH (11% of total in 15 min) and CK (10%) and a sudden rise in coronary resistance, indicating contracture development ("oxygen paradox"). PNP release remained low (0.5%). In hearts subjected to glucose-free anoxia, ATP levels did not rise during 15-min reoxygenation, contracture development was delayed, and the release of LDH and CK was diminished (3.1 and 2.7%, respectively). Leakage of PNP was again low (0.5%). The results indicate that cardiomyocytes are more severely injured by anoxia-reoxygenation than the coronary endothelium. The rapidly developing reoxygenation-induced injury of cardiomyocytes seems to be an energy-dependent phenomenon, since it was attenuated in hearts deprived of substrate in anoxia.
Assuntos
Endotélio Vascular/fisiologia , Nucleotídeos de Adenina/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Células Cultivadas , Circulação Coronária , Creatina Quinase/metabolismo , Hipóxia/fisiopatologia , L-Lactato Desidrogenase/metabolismo , Lactatos/metabolismo , Masculino , Microcirculação/fisiologia , Perfusão , Purina-Núcleosídeo Fosforilase/metabolismo , Ratos , Ratos Endogâmicos , Resistência VascularRESUMO
The metabolism by coronary microvascular endothelial cells (CMEC) of the heart typical substrates palmitate and lactate was compared to that of glucose and glutamine. Confluent cultures of CMEC were used. Palmitate oxidation was saturable and independent of the exogenous albumin concentration. Palmitate, 300 microM, lactate, 1 mM, and glutamine, 0.5 mM, were oxidized to 35, 46, and 56 nmol CO2/h x mg protein. These oxidation rates were decreased by 80, 66, and 48% in presence of 5 mM glucose. The largest energy yield was obtained by glycolytic breakdown of glucose. Glucose, 5 mM, was degraded to lactate by 99%, and oxidized in the Krebs cycle by only 0.04%. 1% was catabolized via the hexose monophosphate pathway. The rate of glucose oxidation in the Krebs cycle could be 30-fold increased by the uncoupler 2,4-dinitrophenol, 30 microM. At concentrations lower than 1 mM the amount of glucose oxidized in the Krebs cycle also grew, indicating existence of the Crabtree effect. The energy demand of CMEC seems to be of the same order as that of the arrested heart.