S. Brosh

Serum xanthine oxidase in jaundice.

Serum xanthine oxidase activity was measured by a radiochemical method in 137 consecutive patients with jaundice of varying etiology and in 40 non-jaundiced patients with liver or other disease. Serum xanthine oxidase was markedly increased, up to 50 times the upper normal limit (mean + 2 S.D.), in 32 out of 34 patients with infectious hepatitis. A slight elevation of serum xanthine oxidase, up to twice the upper normal limit, was found in 2 out of 49 patients with extrahepatic obstructive jaundice and in 4 out of 20 patients with chronic renal failure. In comparison to serum glutamic-oxaloacetic transaminase and lactate dehydrogenase serum xanthine oxidase appeared to be the more sensitive and specific indicator of acute hepatocellular damage.

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.

Developmental Changes in the Activity of Enzymes of Purine Metabolism in Rat Neuronal Cells in Culture and in Whole Brain

Abstract: The activities (Vmax) of several enzymes of purine nucleotide metabolism were assayed in premature and mature primary rat neuronal cultures and in whole rat brains. In the neuronal cultures, representing 90% pure neurons, maturation (up to 14 days in culture) resulted in an increase in the activities of guanine deaminase (guanase), purine‐nucleoside phosphorylase (PNP), IMP 5′‐nucleotidase, adenine phosphoribosyltransferase (APRT), and AMP deaminase, but in no change in the activities of hypoxanthine‐guanine phosphoribosyltransferase (HGPRT), adenosine deaminase, adenosine kinase, and AMP 5′‐nucleotidase. In whole brains in vivo, maturation (from 18 days of gestation to 14 days post partum) was associated with an increase in the activities of guanase, PNP, IMP 5′‐nucleotidase, AMP deaminase, and HGPRT, a decrease in the activities of adenosine deaminase and IMP dehydrogenase, and no change in the activities of APRT, AMP 5′‐nucleotidase, and adenosine kinase. The profound changes in purine metabolism, which occur with maturation of the neuronal cells in primary cultures in vitro and in whole brains in vivo, create an advantage for AMP degradation by deamination, rather than by dephosphorylation, and for guanine degradation to xanthine over its reutilization for synthesis of GMP. The physiological meaning of the maturational increase in these two ammonia‐producing enzymes in the brain is not yet clear. The striking similarity in the alterations of enzyme activities in the two systems indicates that the primary culture system may serve as an appropriate model for the study of purine metabolism in brain.

De novo purine synthesis in skeletal muscle

Myoblast and primary muscle cultures from rat were found to contain the complete pathway of de novo purine nucleotide synthesis. Quantitative assessment of the pathway in skeletal muscle in mice in vivo, revealed a more intensive purine production in muscle than in liver. Skeletal muscle is thus a major site of de novo purine production in the mammalian body.

Guanine Nucleotide Metabolism in Red Blood Cells: The Metabolic Basis for GTP Depletion in HGPRT and PNP Deficiency

The salvage cycles of purine bases include the activity of 5′-nucleotidase on IMP and GMP to form the respective nucleosides inosine (Ino) and guanosine (Guo); further degradation of the nucleosides by purine nucleoside phosphorylase (PNP) to Hx and Gu, and the ribophosphorylation of these bases by HGPRT to IMP and GMP, respectively.

Effects of fructose on synthesis and degradation of purine nucleotides in isolated rat hepatocytes

The effects of fructose on purine nucleotide synthesis and degradation were studied in isolated rat hepatocytes. Incubation of the hepatocytes with fructose resulted in deceleration of the rate of de novo purine synthesis, gauged by the rate of incorporation of precusor [14C]formate into total purines produced, and in acceleration of purine nucleotide degradation, as measured by the rate of conversion of prelabelled purine nucleotides into end-product allantoin. These effects were found to be associated with decreases in cellular content of ATP and Pi and in the metabolic availability of 5-phosphoribosyl 1-pyrophosphate. The results support the suggestion that the fructose-induced acceleration of purine degradation is mediated through activation of AMP deaminase. However, the results also suggest that decreased reutilization of hypoxanthine for IMP synthesis, due to the decreased PP-Rib-P availability, is an additional mechanism for the acceleration of purine degradation. The decreased PP-Rib-P availability is also suggested to be the main mechanism for the fructose-induced deceleration of purine synthesis.

Hormone-Induced Stimulation of Phosphoribosylpyrophosphate and of Purine Synthesis in Mouse Liver in Vivo

A possible role for glycogenolytic hormones in the regulation of purine metabolism was indicated by the finding in normal mice that glucagon and epinephrine increased liver content of PRPP (1) and that epinephrine stimulated purine synthesis (2). Furthermore, increased glucagon level was suggested recently to underly the purine overproduction common in glycogen storage disease type I (3).

Effects of Fructose on Purine Nucleotide Metabolism in Isolated Rat Hepatocytes

In recent years it became apparent that fructose metabolism in the liver tissue has considerable effects on purine nucleotide metabolism. There is ample evidence that fructose metabolism is associated with acceleration of nucleotide degradation (1–5) and evidence is accumulating (6–8), indicating an acceleration effect of fructose also on purine synthesis.

Guanine Metabolism in Primary Rat Neuronal Cells

Guanine ribonucleotides (GuRN) play a crucial role in many functions related to normal cellular and neuronal activities. Depletion of intracellular GuRN pools was reported in red blood cells from patients affected with the hereditary Lesch-Nyhan syndrome (1,2) and suggested as the possible metabolic basis for the neurological manifestations associated with this syndrome. Despite the apparent connection between guanine and GuRN metabolism and neuronal function, knowledge regarding the specific metabolism of guanine and GuRN in neuronal cells is rather limited.

Hormone-induced stimulation of purine synthesis

Urid acid overproduction is an etiological factor in uric acid lithiasis. It is well documented to reflect primary metabolic defects in several rare inborn errors of purine metabolism (deficiency of hypoxanthine-guanine-phosphoribosyltransferase (HGPRT) and superactivity of phosphoribosylpyrophosphate (PRPP) synthetase), and as a secondary manifestation in several myeloproliferative and other hematological disorders. In addition to the above, purine overproduction patients have been observed, many with stone disease, in whom the etiology of urate overproduction is not known. In the framework of our investigation of the mechanisms of purine overproduction in man, this study was done in order to clarify the possible role of the glycogenolytic hormones in the regulation of purine synthesis. Such a role was indicated by the finding in normal mice that glucagon and epinephrine increased liver content of PRPP (1), a rate-limiting substrate for de novo purine synthesis, and that epinephrine stimulated purine synthesis (2). Furthermore, increased glucagon level was suggested recently to underly the purine overproduction common in glycogen storage disease type I (3). In the present communication we report that several glycogenolytic hormones exert a stimulatory effect on the rate of PRPP and purine synthesis in mouse liver in vivo (4, 5). Our studies concerning the mechanism of this phenomenon are discribed.