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A system for the study in situ of the pentose phosphate pathway in rabbit liver.

ABSTRACT: A surgical procedure for the isolation of the liver from the systemic circulation of the anaesthetized rabbit is described. The technique allowed the metabolism in situ of intraportally infused substrates to be followed for periods up to 5min, free from the contaminating influences of metabolism by other body tissues. Details of the procedures necessary to achieve the uniform infusion, homogeneous distribution and containment of (14)C-labelled glucose substrates in the liver by haemostasis are described. Changes in pO(2), pCO(2), pH and the concentrations of NADP(+), NADPH and glucose during each minute interval of the total 5min period of metabolism are given. Reactant ratios of the lactate dehydrogenase system and the adenine nucleotide system have been calculated from the concentrations of the pertinent metabolites for the same period of metabolism. Glucose production by rabbit liver in situ proceeded at the rate of 1.08mumol/min per g wet wt. of liver during the 5min metabolic interval. The presence of the oxidative reactions of the pentose phosphate pathway of glucose metabolism was inferred from the quotient oxidation of [1-(14)C]glucose/oxidation of [6-(14)C]glucose=1.8.

SUBMITTER: Williams JF

PROVIDER: S-EPMC1177093 | biostudies-other | 1971 Aug

REPOSITORIES: biostudies-other

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Project description:AimsInflammation and possibly hypoxia largely affect glucose utilization in atherosclerotic arteries, which could alter many metabolic systems. However, metabolic changes in atherosclerotic plaques remain unknown. The present study aims to identify changes in metabolic systems relative to glucose uptake and hypoxia in rabbit atherosclerotic arteries and cultured macrophages.MethodsMacrophage-rich or smooth muscle cell (SMC)-rich neointima was created by balloon injury in the iliac-femoral arteries of rabbits fed with a 0.5% cholesterol diet or a conventional diet. THP-1 macrophages stimulated with lipopolysaccharides (LPS) and interferon-γ (INFγ) were cultured under normoxic and hypoxic conditions. We evaluated comprehensive arterial and macrophage metabolism by performing metabolomic analyses using capillary electrophoresis-time of flight mass spectrometry. We evaluated glucose uptake and its relationship to vascular hypoxia using (18)F-fluorodeoxyglucose ((18)F-FDG) and pimonidazole, a marker of hypoxia.ResultsThe levels of many metabolites increased in the iliac-femoral arteries with macrophage-rich neointima, compared with those that were not injured and those with SMC-rich neointima (glycolysis, 4 of 9; pentose phosphate pathway, 4 of 6; tricarboxylic acid cycle, 4 of 6; nucleotides, 10 of 20). The uptake of (18)F-FDG in arterial walls measured by autoradiography positively correlated with macrophage- and pimonidazole-immunopositive areas (r = 0.76, and r = 0.59 respectively; n = 69 for both; p<0.0001). Pimonidazole immunoreactivity was closely localized with the nuclear translocation of hypoxia inducible factor-1α and hexokinase II expression in macrophage-rich neointima. The levels of glycolytic (8 of 8) and pentose phosphate pathway (4 of 6) metabolites increased in LPS and INFγ stimulated macrophages under hypoxic but not normoxic condition. Plasminogen activator inhibitor-1 protein levels in the supernatant were closely associated with metabolic pathways in the macrophages.ConclusionInfiltrative macrophages in atherosclerotic arteries might affect metabolic systems, and hypoxia but not classical activation might augment glycolytic and pentose phosphate pathways in macrophages.

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A system for the study in situ of the pentose phosphate pathway in rabbit liver.

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