Hypouricemia and tubular transport of uric acid.

Hypouricemia is defined when a serum urate concentration is less than or equal 2.0mg/dl. Differential diagnosis is made by fractional uric acid excretion with the identification of urate transporters and intracellular proteins involved in the tubular transport of uric acid. This review examines current knowledge on uric acid tubular transport and the various clinical situations of hypouricemia.

Functional characterization of human xanthine oxidase allelic variants.

OBJECTIVE: Xanthine oxidase (XO) catalyzes the oxidation of endogenous and exogenous purines and pyrimidines. In this study, we speculated that individual variations in XO activity are caused by genetic variations in the XO gene. METHODS: To investigate the genetic variations in XO in 96 Japanese participants, denaturing high-performance liquid chromatography was used. To assess the effects of these variations on enzymatic activity, wild-type XO and 21 types of variant XO--including those in the database and those just discovered--were transiently expressed in COS-7 cells. RESULTS: Three nonsynonymous single nucleotide polymorphisms, including 514G>A (Gly172Arg), 3326A>C (Asp1109Thr), and 3662A>G (His1221Arg) were identified in Japanese participants. Functional characterization of 21 XO variants showed a deficiency in enzyme activity in two variants (Arg149Cys and Thr910Lys); low activity (intrinsic clearance, CLint: 22-69% compared with the wild-type) in six variants (Pro555Ser, Arg607Gln, Thr623Ile, Asn909Lys, Pro1150Arg, and Cys1318Tyr); and high activity (CLint: approximately two-fold higher than that in the wild-type) in two variants (Ile703Val and His1221Arg). CONCLUSION: These results suggest that several single nucleotide polymorphisms in the XO gene are involved in individual variations in XO activity. In addition, such findings will be useful to identify xanthinuria patients.

Inhibitory or scavenging action of ketoconazole and ciclopiroxolamine against reactive oxygen species released by primed inflammatory cells.

BACKGROUND: Reactive oxygen species (ROS) released from inflammatory cells constitute one of the critical causative factors in inflammatory skin diseases such as seborrhoeic dermatitis and atopic dermatitis. OBJECTIVES: To investigate inhibitory effects of ketoconazole (KCZ) and ciclopiroxolamine (CPO), both of which have been used for the treatment of seborrhoeic dermatitis, on ROS released from inflammatory cells. METHODS: The methyl-Cypridina-luciferin analogue-dependent chemiluminescence method was employed for the detection of ROS production by phorbol 12-myristate 13-acetate (PMA)-stimulated inflammatory cells. Moreover, the radical scavenging activities of both agents were examined by using a hypoxanthine-xanthine oxidase system and the stable radical 1,1-diphenyl-2-picrylhydrazyl (DPPH). NADPH oxidase activity was determined in particulate (membrane) fractions prepared from PMA-stimulated RAW 264 x 7 cells, a macrophage-like cell line. RESULTS: Both of these antifungal agents inhibited PMA-stimulated ROS production. However, only CPO significantly scavenged both ROS generated by the hypoxanthine-xanthine oxidase system and DPPH, and the scavenging activity of CPO seemed to act on ROS other than superoxide anions. Although KCZ inhibited PMA-stimulated ROS production, it did not show radical-scavenging activities. The inhibition of ROS production by KCZ is probably attributable to the inhibition of NADPH oxidase activity. CONCLUSIONS: The mechanism of the inhibitory action of KCZ against PMA-stimulated ROS production is distinct from that of CPO. Knowledge of the inhibitory or scavenging effects of both antifungal agents on ROS released from inflammatory cells may be useful in developing a therapeutic strategy for dermatitis.

Xanthine oxidoreductase is an endogenous regulator of cyclooxygenase-2.

Xanthine oxidoreductase (XOR) is the enzyme responsible for the final step in purine degradation resulting in the generation of uric acid. Here we have generated mice deficient in XOR. As expected, these animals lack tissue XOR activity and have low to undetectable serum levels of uric acid. Although normal at birth, XOR-/- mice fail to thrive after 10 to 14 days, and most die within the first month. The cause of death appears to be a form of severe renal dysplasia, a phenotype that closely resembles what has been observed previously in cyclooxygenase-2 (COX-2)-deficient mice. We further demonstrate that in the first month of life, a period in which the mouse kidney is undergoing rapid maturation and remodeling, wild-type mice exhibit an approximately 30-fold increase in renal XOR activity, with a corresponding induction of COX-2 expression. In contrast, during this same period, XOR-/- animals fail to augment renal COX-2 expression. Finally, we show that in vitro and in vivo, uric acid can stimulate basal COX-2 expression. These results demonstrate that XOR activity is an endogenous physiological regulator of COX-2 expression and thereby provide insight into previous epidemiological evidence linking elevated serum uric levels with systemic hypertension and increased mortality from cardiovascular diseases. In addition, these results suggest a novel molecular link between cellular injury and the inflammatory response.

Capillary electrophoresis for detection of inherited disorders of purine and pyrimidine metabolism.

BACKGROUND: Measurement of purine and pyrimidine metabolites presents complex problems for separations currently performed by HPLC and thin-layer chromatography in clinical practice. We developed a novel capillary electrophoresis method for this purpose. METHODS: Separations were performed in 60 mmol/L borate-2-amino-2-methyl-1-propanol-80 mmol/L sodium dodecyl sulfate (pH 9.6) at 35 degrees C. RESULTS: The conditions reported allowed separation of all diagnostic metabolites from major urinary constituents in an analysis time of 3 min and with a separation efficiency of 220 000 theoretical plates/m. The clinically important metabolites were detectable at concentrations of 0.85-4.28 micromol/L. The method was linear over the range 5-500 micromol/L (r >0.99). The within-run and intra- and interday imprecision (CV) was <5%. Characteristic abnormalities were detected in the electropherograms of urine samples from patients with purine and pyrimidine enzyme deficiencies. We provide the electrophoretic and spectral characteristics of many intermediates in purine and pyrimidine metabolism and describe common artifacts from medication and ultraviolet-absorbing compounds. CONCLUSION: Capillary electrophoresis is a valuable screening tool in the detection of inborn errors of purine and pyrimidine metabolism.

[Hereditary xanthinuria, rare cause of hypo-uric acidemia. 2 cases]. / La xanthinurie héréditaire, cause rare d'hypo-uricémie. 2 observations.

BACKGROUND: Hypouricemia can be observed in uncommon situations as in our two patients with hereditary xanthinuria. CASE REPORTS: In the first case, hereditary xanthinuria was discovered in a 36-year-old man when routine tests revealed hypouricemia. In the second case, a 76-year-old woman, hypouricemia was also a fortuitous discovery. She had major xanthinuria and a radiotranslucid lithiasis in the right kidney. DISCUSSION: Hereditary xanthinuria is characterized by hypouricemia, low urinary urate excretion and increased concentration of xanthine and to a lesser extent hypoxanthine. The disease results from a defect in xanthine oxidase and is considered to be transmitted by autosomal recessive heredity. This rare metabolic disorder is more often asymptomatic and detected by routine chemistry. Development of xanthine lithiasis is directly related to the low solubility of xanthine and is the main complication of the disease, occurring in 30-40% of patients. There is no effective treatment and the only useful measure is to prevent xanthine urolithiasis by maintaining urinary output above 2 l/day.

Neutrophil mediated microvascular injury in acute, experimental compartment syndrome.

The purpose of this study was to determine the contribution of neutrophils and tissue xanthine oxidase to the skeletal muscle microvascular dysfunction in an ex vivo model of acute compartment syndrome. Adult dogs were rendered neutropenic or depleted of tissue xanthine oxidase before gracilis muscle isolation. Compared with continuously perfused, nonischemic muscles, acute, experimental compartment syndrome resulted in a dramatic increase in microvascular permeability, muscle neutrophil content, and muscle vascular resistance. Neutropenia prevented, whereas xanthine oxidase depletion had no effect on, the microvascular dysfunction and muscle neutrophil infiltration elicited by experimental compartment syndrome. These results suggest that neutrophils contribute to the microvascular dysfunction and blood flow distribution abnormalities elicited by acute, experimental compartment syndrome.

[Inherited disorders of uric acid metabolism--classification, enzymatic- and DNA-diagnosis].

Uric acid is the end product of purine metabolism in human. Then, the enzymatic abnormalities, concerning purine metabolism, cause disorders of uric acid metabolism including hyperuricemia and hypouricemia. The superactivity of 5-phosphoribosyl-pyrophosphate (PRPP) synthetase and deficiency of hypoxanthine-guanine phosphoribosyltransferase (HGPRT) caused hyperuricemia. In glycogen storage diseases of type I, III, V, and VII, decreased energy supply induces hyperuricemia by accelerating ATP degradation. Deficiencies of xanthine oxidase (XO), purine nucleoside phosphorylase (PNP), and PRPP were reported causing hypouricemia. Many methods for DNA-diagnosis were developed including Southern blot, Northern blot, PCR-SSCP (polymerase chain reaction-single strand conformation polymorphism), PCR-RFLP (restriction fragment length polymorphism), and allele specific oligonucleotide hybridization etc.

[Xanthine oxidase deficiency (hereditary xanthinuria), molybdenum cofactor deficiency].

Hereditary xanthinuria is a rare autosomal recessive disorder, with xanthine oxidase deficiency. Patients often display renal symptoms because they excrete a large amounts of xanthine in urine. An high-fluid-intake, alow-purine-food, and alkalinization of urine are effective in the patients. Molybdenum cofactor is essential for xanthine oxidase, sulfite oxidase and aldehyde oxidase. Patients with molybdenum cofactor deficiency display severe neurological symptoms, such as severe convulsions. The patients increase urinary excretions of xanthine and sulfite. Treatments are ineffective for neurological symptoms.

[Hematotoxicity caused by azathioprine genetically determined and aggravated by xanthine oxidase deficiency in a patient following renal transplantation]. / Hématotoxicité de l'azathioprine à déterminisme génétique aggravée par un déficit en xanthine oxydase chez une transplantée rénale.

Azathioprine is an immunosuppressor used with ciclosporin and corticosteroids after organ transplantation. Azathioprine is rapidly transformed into 6-mercaptopurine which in turn is metabolized by three competitive pathways: a) intracellular hypoxanthine guanine phosphoribosyl transferase leads to 6-thioguanine nucleotides which can damage chromosome DNA; b) thiopurine methyltransferase produces inactive methylated derivatives; c) xanthine oxidase produces thiouric acid. Due to inter-individual variations in the later two pathways, azathioprine dose must be adapted to each patient. A 48-year-old female patient underwent renal transplantation in 1994 and was given immunosuppressive therapy combining thymoglobulins, azathioprine and ciclosporin. Severe leukopenia (< 3000/mm3) occurred on day 5 requiring withdrawal of azathioprine. Known hypouricaemia (< 50 mumol/l) suggested xanthine oxidase deficiency. Laboratory results confirmed xanthine oxidase deficiency and also revealed reduced thiopurine methyltransferase activity (14.9 pmol/h/mg Hb). Azathioprine toxicity was confirmed by regression of the leukopenia after withdrawal and recurrence at rechallenge. Xanthine oxidase deficiency occurs in 2% of the general population. Reduced thiopurine methyltransferase activity affects 11% of the population. The combined presence of these two genetic anomalies led to early and sudden intolerance to azathioprine and emphasize the need to develop new immunosuppressor agents degraded by other metabolic pathways.

Effects of verapamil on myocardial stunning in xanthine-oxidase deficient hearts: pre-treatment vs. post-ischemic treatment.

The role of free radicals and the protective action of calcium antagonists have been established in myocardial stunning in canine hearts, which contain a considerable level of xanthine oxidase, a free radical producing enzyme. However, myocardial stunning in hearts which lack xanthine oxidase and its modification by calcium antagonists in vivo remain uncharacterized. The present study examined this issue using open-chest anesthetized rabbits. Myocardial stunning was induced by a 10-min coronary occlusion and reperfusion. Regional systolic thickening fraction (TF) was determined using an epicardial Doppler sensor, together with other hemodynamic parameters. In untreated control rabbits, recovery of TF from the 10 min transient ischemia was 43 +/- 3% of the baseline at 30 min after reperfusion. Administration of verapamil (200 micrograms/kg bolus plus 40 micrograms/kg/min), which was started before the onset of ischemia and continued until 20 min after reperfusion, significantly improved the recovery of TF to 74 +/- 6% (p < 0.05). A similar improvement in post-ischemic contractile function (TF = 77 +/- 10%) was observed when verapamil was injected at the same rate, but the infusion was discontinued 1 min after the coronary occlusion. Myocardial ATP depletion after the 10 min ischemia was significantly less in the verapamil-pretreated rabbits compared with untreated controls (10.1 +/- 1.0 vs. 6.2 +/- 0.7 mumol/g dry wt., p < 0.05). The difference in TF between the rabbit with and without verapamil treatment could not be explained by afterload reduction. When verapamil (100 micrograms/kg bolus plus 20 micrograms/kg/min) was given during the reperfusion period alone, TF recovery was poorer (TF = 22 +/- 8%) than the control value. Thus, it was concluded that verapamil attenuates myocardial stunning in the hearts with trace levels of xanthine oxidase, and that the beneficial effect is achieved only by pretreatment, not by post-ischemic treatment with verapamil.