O. Sperling

Metabolic and calcium kinetic studies in idiopathic hypercalciuria

Calcium balances and calcium kinetic studies using (47)Ca were performed in nine male patients with idiopathic hypercalciuria and in three normal male subjects. A sharp reduction in calcium intake in eight patients with idiopathic hypercalciuria caused a decrease in urinary calcium excretion, the latter remaining elevated above that reported for normal subjects on a low calcium diet. The hypercalciuric patients had an enlarged miscible calcium pool size, an increased calcium turnover rate, increased bone formation and bone resorption rates, and an elevated true intestinal calcium absorption rate, the increase of the latter three parameters being proportional to the increase of the turnover rate. The fraction of the calcium turnover rate excreted in the urine was elevated whereas that constituted by the endogenous fecal calcium excretion was decreased. Arguments are presented for the concept that the primary abnormality in idiopathic hypercalciuria is neither renal calcium hyperexcretion nor intestinal calcium hyperreabsorption, but a more fundamental disturbance in calcium metabolism of as yet unknown cause, leading to a high calcium turnover.

De novo synthesis of purine nucleotides in human peripheral blood leukocytes. Excessive activity of the pathway in hypoxanthine-guanine phosphoribosyltransferase deficiency.

Human peripheral blood leukocytes were studied for the presence and the regulatory properties of the pathway of de novo synthesis of purine nucleotides. The cells were found to incorporate the labeled precursors formate and glycine into purines. The rate of [14C]-formate incorporation was decreased by several compounds known to inhibit purine synthesis by affecting the activity by glutamine phosphoribosylpyrophosphate (PRPP) amidotransferase, the first committed enzyme in the pathway, either through decreasing the availability of PRPP, a substrate for this enzyme, or through exerting inhibition on this enzyme. PRPP availability in the leukocyte was found to be limiting for purine synthesis. Increased PRPP availability resulting from activation of PRPP synthetase by increasing inorganic phosphate (Pi) concentration caused acceleration of purine synthesis. On the other hand, no clear-cut evidence was obtained for the availability of ribose-5-phosphate in the leukocyte being rate limiting at physiological extracellular Pi concentration for PRPP generation, and thus for purine synthesis. However, the addition of methylene blue, which accelerates the oxidative pentose shunt that produces ribose-5-phosphate, resulted in acceleration of PRPP generation and of purine synthesis only when PRPP synthetase was largely activated at high Pi concentration. These results may be taken to suggest that ribose-5-phosphate availability is indeed not limiting for PRPP generation under physiological conditions. Purine synthesis de novo was accelerated more than 13-fold in the leukocytes of two gouty patients affected with partial deficiency of hypoxanthine-guanine phosphoribosyltransferase, but was normal in the leukocytes of an obligate heterozygote for this enzyme abnormality. The results domonstrate in peripheral human leukocytes the presence of the complete pathway of de novo synthesis of purine nucleotides and the manifestation in these cells of the biochemical consequences of hypoxanthine-guanine phosphoribosyltransferase deficiency, i.e., increased availability of PRPP and acceleration of purine synthesis de novo. The results indicate the usefulness of leukocytes as a model tissue for the study of purine metabolism in man.

Metabolic cooperation between human fibroblasts with normal and with mutant superactive phosphoribosylpyrophosphate synthetase.

METABOLIC cooperation is a form of intercellular communication by which cells in contact exchange molecules, a process providing multicellular organisms with an important mechanism for control of metabolic activity1. Subak-Sharpe et al.2 observed contact-dependent transfer of purine nucleotides from normal cells to cells mutationally incapable of producing inosinic acid due to deficiency in hypoxanthine–guanine phosphoribosyl-transferase3. Metabolic cooperation of this type was later also demonstrated with other enzymic markers, such as adenine phosphoribosyltransferase and thymidine kinase4,5. Such transfers are characterised, by the normal cell being the donor and the mutant cell being the recipient, the former transferring to the latter a mutationally lacking metabolite. We report here on a new form of contact-dependent metabolic cooperation, unique in that the transfer of a metabolite occurs from a mutant donor cell to a normal recipient cell.

Urine xanthine oxidase activity in urinary tract infection.

Xanthine oxidase (XO) activity was found to be negligible in sterile human urines (less than 480 units, as presently defined, per litre). Significant XO activity was found in all urines containing more than 10(5) bacteria/ml, except for urines infected with Staphylococcus aureus, in which XO activity ranged from 347 to 714 units per litre. Plasma XO is not transferred to the urine, as demonstrated by the negligible XO activity found in sterile urines from patients with raised plasma XO activity. Determination of urinary XO activity is a suitable procedure for the detection of urinary tract infection.

A kinetic model for the intramolecular distribution of 15N in uric acid in patients with primary gout fed 15N-glycine

Abstract The concept of an abnormality of glutamine metabolism in primary gout was first proposed on the basis of isotope data: when 15 N-glycine was administered to gouty subjects, there was disproportionately great enrichment of N-(3+9) of uric acid, which derive from the amide-N of glutamine. An unduly high concentration of 15 N in glutamine was postulated, and attributed to a hypothetical defect in the catabolism of glutamine. Excess glutamine was proposed as the driving force of uric acid overproduction. We have reexamined this proposition in four gouty and three control subjects. In three of the gouty subjects the driving force of excessive purine biosynthesis was a known surplus of α-5-phosphoribosyl-1-pyrophosphate. The precursor glycine and glutamine pools were sampled by isolation of urinary hippurate and phenylacetylglutamine. Enrichment values of hippurate and of phenacetylglutamine were normal in all of the gouty subjects studied. However, the time course of 15 N enrichment of hippurate differed from that of the amide-N of glutamine. A kenetic model was constructed, and experimentally derived data were used to drive the model. It was found that disproportionate labeling of N-(3+9) could be produced from the model when appropriate constraints were used. Thus, preferential enrichment of N-(3+9) in gouty overproducers given 15 N-glycine does not necessarily reflect a specific abnormality of glutamine metabolism, but rather appears to be a kinetic phenomenon associated with accelerated purine biosynthesis per se.

The Kinetics of Intramolecular Distribution of 15N in Uric Acid Following Administration of 15N-Glycine: Preferential Labeling of N−(3+9) of Uric Acid in Primary Gout and a Reappraisal of the “Glutamine Hypothesis”

Patients with primary gout and excessive uric acid excretion fed a test dose of 15N-glycine incorporate increased quantities of 15N into urinary urate [1–4]. Although enrichment of all 4 nitrogen atoms of uric acid is excessive [4,5] that of N−(3+9) is disproportionately great, especially in flamboyant overexcretors of uric acid [5]. Since N−3 and N−9 of uric acid are derived from the amide-N of glutamine [6,7], Gutman and Yu [5,8] proposed a defect in glutamine metabolism in primary gout. Since urinary ammonium, which arises principally from glutamine [9,10], is reduced in many gouty subjects [11,12], they further postulated a reduction of glutaminase activity [5,8]. The hyperglutamatemia of gout has now suggested a defect of glutamate metabolism, with diversion of glutamic acid toward glutamine and purine biosynthesis [13,14].