Current problems in clinical biochemistry最新文献

Three gluconeogenic enzymes, P-pyruvate carboxykinase (PPCK), fructose-1-6 bisphosphatase (FBPase), and glucose-6-phosphatase (G6Pase) were measured in identified structures of rat nephron from 2 days before birth to maturity. In the proximal convoluted tubule, the three enzymes increased from the earliest age assayed to +14 days (PPCK, 7-fold, FBPase, 2-fold and G6Pase, 50-fold). Among the 7 defined structures that were analyzed, highest levels at all ages were in the proximal convoluted tubule with almost no activity in the distal convoluted tubule. All three enzymes had negligible activity in the neogenic zone and mesenchyme. Supported by grants from the Public Health Service (HD 03891 and NS-05221) and the American Cancer Society (P-78).

Single proximal rat kidney tubules and Henle loops were microperfused with labelled S-substituted glutathione derivatives which are physiological mercapturic acid precursors. S-methyl-glutathione as well as the non-permeating bromo-sulfaleinglutathione adduct were degraded in the proximal convolution with a half-life of about 3.5 sec. The findings demonstrate that the brush-border membrane-bound renal gamma-glutamyl transpeptidase acts on luminal substrates and is involved in mercapturic acid synthesis and splitting of extracellular glutathione.

These results establish the existence of a cytoplasmic glutaminase-gamma-glutamyltransferase enzyme as a distinct entity. Its products are ammonia and activated glutamate. Ammonia liberation is obligatory to the utilization of the amide bond energy in forming gamma-glutamyl-PO4. This activated glutamate can then be utilized in the gamma-glutamyl cycle for the synthesis of gamma-glutamylcysteine. The cytoplasmic glutamine utilizing pathway is closely coupled to gamma-glutamyl cycl activity: loading the cycle stimulates increased renal glutamine uptake into this pathway. Consequently, the pathway, stimulated by elevated ADP levels, appears to function as an auxilary source of gamma-glutamyl moieties for the gamma-glutamyl cycle. Although insignificant compared to the mitochondrial pathway's contribution to ammonia production in metabolic acidosis, it is highly significant from the perspective of the Unitary Hypothesis. The existence of this dual system allows the demonstration of a shift in glutamine utilization from predominant cytoplasmic to overwhelming mitochondrial glutamine utilization in metabolic acidosis corresponding to a rise in the NH3/gln ratio from 1.2 to 1.9 and a quantitative recovery of gln carbon as CO2 and glucose. The fact that this shift in pathways is induced by acidosis (through adrenosteroids) and that it represents a 10 to 20 fold activation of the mitochondrial pathway is completely consistent with a glucocorticoid mediated glutamine permeability increase of the inner mitochondrial membrane.

Transport and metabolism of endogenous fatty acids was studied using an isolated rat kidney preparation. Results tended to confirm the inhibitory effects of probenecid on fatty acid uptake by the kidney but, at the concentration used, probenecid was ineffective in changing the distribution of 14C palmitate within tissue lipids. Alpha-bromopalmitate (alphaBP) (0.4 mM), an inhibitor of fatty acid oxidation, caused an initial enhancement of 14C palmitate uptake, a decrease in esterification into tissue phospholipids and an increase in 14C recovered as free fatty acid in the tissue. This increase had no inhibitory effect on entry of fatty acids. alpha BP had no effect on 14C octanoate uptake and appeared not to affect general metabolism. The results are consistent with the possibility that alphaBP interacts with a renal fatty acid binding protein.

Glucose production, pyruvate uptake and lactate production were taken as metabolic viability tests for isolated rat kidney tubules preserved in hypothermia. The results depend on the type of preservation solution used. Species specific serum is the only solution sustaining cellular metabolism at a normal level. Using Collins solution all viability parameters showed the lowest results. Addition of certain substrates to the Krebs-Henseledt solution improves metabolic viability.