Hypoxanthine Effect on Equilibrative and Concentrative Adenosine Transport in Human Lymphocytes. Implications in the Phatogenesis of Lesch-Nyhan Syndrome

We postulated that increased levels of hypoxanthine, a main characteristic of hypoxanthine phosphoribosyltransferase (HPRT) deficiency, may influence adenosine function which could be related to some of the neurological features of the Lesch-Nyhan syndrome. We have examined the effect of hypoxanthine on different adenosine transporters in peripheral blood lymphocytes from control subjects. Increased hypoxanthine concentrations (25 μM) significantly decreased adenosine transport. The equilibrative adenosine transporters (79.6% of the adenosine transport), both NBTI sensitive and NBTI insensitive, were affected significantly. In contrast, the concentrative adenosine transporters were not influenced by hypoxanthine. These results supports the hypothesis that increased hypoxanthine levels influence equilibrative (predominantly NBTI-insensitive type) adenosine transporters.     

Physiological responses to maximal intensity intermittent exercise

Physiological responses to repeated bouts of short duration maximal-intensity exercise were evaluated. Seven male subjects performed three exercise protocols, on separate days, with either 15 (S15), 30 (S30) or 40 (S40) m sprints repeated every 30 s. Plasma hypoxanthine (HX) and uric acid (UA), and blood lactate concentrations were evaluated pre- and postexercise. Oxygen uptake was measured immediately after the last sprint in each protocol. Sprint times were recorded to analyse changes in performance over the trials. Mean plasma concentrations of HX and UA increased during S30 and S40 (P less than 0.05), HX increasing from 2.9 (SEM 1.0) and 4.1 (SEM 0.9), to 25.4 (SEM 7.8) and 42.7 (SEM 7.5) mumol.l-1, and UA from 372.8 (SEM 19) and 382.8 (SEM 26), to 458.7 (SEM 40) and 534.6 (SEM 37) mumol.l-1, respectively. Postexercise blood lactate concentrations were higher than pretest values in all three protocols (P less than 0.05), increasing to 6.8 (SEM 1.5), 13.9 (SEM 1.7) and 16.8 (SEM 1.1) mmol.l-1 in S15, S30 and S40, respectively. There was no significant difference between oxygen uptake immediately after S30 [3.2 (SEM 0.1) l.min-1] and S40 [3.3 (SEM 0.4) l.min-1], but a lower value [2.6 (SEM 0.1) l.min-1] was found after S15 (P less than 0.05). The time of the last sprint [2.63 (SEM 0.04) s] in S15 was not significantly different from that of the first [2.62 (SEM 0.02) s]. However, in S30 and S40 sprint times increased from 4.46 (SEM 0.04) and 5.61 (SEM 0.07) s (first) to 4.66 (SEM 0.05) and 6.19 (SEM 0.09) s (last), respectively (P less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)     

以生产病毒唑的副产物为原料合成6─BA

本文介绍了以抗病毒药病毒唑生产中产生的副产物乙酰次黄嘌呤为原料,经水解、氯代及氨解合成植物生长调节剂６─苄氨基嘌呤的方法     

Vibrational analysis and molecular force field of hypoxanthine as determined from ultraviolet resonance Raman spectra of native and deuterated species

Ultraviolet resonance Raman scattering spectra from aqueous solutions of hypoxanthine and its deuterated species (C8-deuterated, N-deuterated and C8-, N-deuterated derivatives) have been collected and reported in the spectral region between 400 and 1800 cm^–1. The laser excitation wavelengths at 281 nm and 257 nm correspond to preresonance and pure resonance conditions, respectively, with the purine strongly allowed ^* electronic transition: thus the observed experimental Raman features mainly correspond to inplane vibrational modes. The latter were then assigned according to the Wilson GF method by using an empirical harmonic valence force field. Normal mode calculations are based on a non-redundant set of internal coordinates. The calculated vibrational mode wavenumbers and their isotopic shifts upon selective deuterations are in good agreement with the experimental data. The present normal mode analysis rests on the transferability of the guanine and adenine force constants proposed in recent works based on resonance Raman spectroscopy and neutron inelastic scattering data from these major purine bases. Correspondence to: M. Ghomi     

Effects of Hypoxanthine on Adenosine Transport in Human Lymphocytes. Implications in the Phatogenesis of Lesch–Nyhan Syndrome

We have examined the effect of hypoxanthine on adenosine transport and [³H] NBTI binding in peripheral blood lymphocytes (PBL) cultures. Pre‐incubation with hypoxanthine originates a dose dependent decrease of adenosine transport and [³H] NBTI binding sites in PBL.     

Specificity and Catalytic Mechanism in Family 5 Uracil DNA Glycosylase

UDGb belongs to family 5 of the uracil DNA glycosylase (UDG) superfamily. Here, we report that family 5 UDGb from Thermus thermophilus HB8 is not only a uracil DNA glycosyase acting on G/U, T/U, C/U, and A/U base pairs, but also a hypoxanthine DNA glycosylase acting on G/I, T/I, and A/I base pairs and a xanthine DNA glycosylase acting on all double-stranded and single-stranded xanthine-containing DNA. Analysis of potentials of mean force indicates that the tendency of hypoxanthine base flipping follows the order of G/I > T/I, A/I > C/I, matching the trend of hypoxanthine DNA glycosylase activity observed in vitro. Genetic analysis indicates that family 5 UDGb can also act as an enzyme to remove uracil incorporated into DNA through the existence of dUTP in the nucleotide pool. Mutational analysis coupled with molecular modeling and molecular dynamics analysis reveals that although hydrogen bonding to O₂ of uracil underlies the UDG activity in a dissociative fashion, Tth UDGb relies on multiple catalytic residues to facilitate its excision of hypoxanthine and xanthine. This study underscores the structural and functional diversity in the UDG superfamily.     

肌苷产生菌中降低核苷水解酶的研究

本研究以枯草杆菌肌苷产生菌ＳＤ９４０１（Ａｄｅˉ＋Ｔｈｉˉ＋Ｈｉｓˉ＋８—ＡＧｒ＋６－ＭＰｒ）为出发菌株,经紫外线和硫酸二乙酯诱变,在以肌苷为唯一碳源的补充培养基上筛选到一株核苷水解酶缺失菌株。实验结果表明这一突变株积累比亲株高３０％左右的肌苷,平均达到２６．８９ｍｇ／ｍｌ,最高到２８．０３ｍｇ／ｍｌ。且发酵液中未测出次黄嘌呤。     

Kainate-Evoked Release of Adenosine from the Hippocampus of the Anaesthetised Rat: Possible Involvement of Free Radicals

Abstract: Using microdialysis in the hippocampus of anaesthetised rats, the concentration of extracellular adenosine was estimated to be 0.8 µ M . Kainic acid (0.1–25 m M ) in the perfusate evoked a concentration-dependent release of adenosine with an EC₅₀ of 940 µ M . Two 5-min pulses of 1 m M kainic acid in the perfusate increased the dialysate levels with an S2/S1 ratio of 0.52 ± 0.03. Kainate-evoked release of adenosine was reduced significantly by 10 µ M tetrodotoxin and by a κ-receptor agonist, U50,488H (100 µ M ). The S2/S1 ratio was reduced by 4.5 µ M 6-cyano-7-nitroquinoxaline-2,3-dione, a non-NMDA receptor antagonist, but not by the NMDA receptor blockers (+)-MK-801 (dizocilpine; 100 µ M ) or (±)-2-amino-5-phosphonopentanoic acid (1 m M ), indicating a non-NMDA receptor-mediated process. The S2/S1 ratio was also reduced significantly by 10 m M ascorbic acid, 10 m M glutathione (a scavenger of hydroperoxides), and 1 m M oxypurinol (a xanthine oxidase inhibitor), indicating the possible involvement of free radicals. Neither the adenosine A1 receptor antagonist 8-cyclopentyl-1,3-dimethylxanthine (100 µ M ) nor the A1 adenosine receptor agonist R (−)- N ⁶-(2-phenylisopropyl)adenosine (100 µ M ) affected release. Adenosine release evoked by kainic acid is therefore mediated by activation of non-NMDA receptors and may involve the propagation of action potentials and the production of free radicals.     

Abnormal purine and pyrimidine nucleotide content in primary astroglia cultures from hypoxanthine-guanine phosphoribosyltransferase-deficient transgenic mice

Lesch-Nyhan syndrome is a pediatric metabolic-neurological syndrome caused by the X-linked deficiency of the purine salvage enzyme hypoxanthine-guanine phosphoribosyltransferase (HGPRT). The cause of the metabolic consequences of HGPRT deficiency has been clarified, but the connection between the enzyme deficiency and the neurological manifestations is still unknown. In search for this connection, in the present study, we characterized purine nucleotide metabolism in primary astroglia cultures from HGPRT-deficient transgenic mice. The HGPRT-deficient astroglia exhibited the basic abnormalities in purine metabolism reported before in neurons and various other HGPRT-deficient cells. The following abnormalities were found: absence of detectable uptake of guanine and of hypoxanthine into intact cell nucleotides; 27.8% increase in the availability of 5-phosphoribosyl-1-pyrophosphate; 9.4-fold acceleration of the rate of de novo nucleotide synthesis; manyfold increase in the excretion into the culture media of hypoxanthine (but normal excretion of xanthine); enhanced loss of label from prelabeled adenine nucleotides (loss of 71% in 24 h, in comparison with 52.7% in the normal cells), due to 4.2-fold greater excretion into the media of labeled hypoxanthine. In addition, the HGPRT-deficient astroglia were shown to contain lower cellular levels of ADP, ATP, and GTP, indicating that the accelerated de novo purine synthesis does not compensate adequately for the deficiency of salvage nucleotide synthesis, and higher level of UTP, probably due to enhanced de novo synthesis of pyrimidine nucleotides. Altered nucleotide content in the brain may have a role in the pathogenesis of the neurological deficit in Lesch-Nyhan syndrome.     

Posttranslational ruling of xanthine oxidase activity in bovine milk by its substrates

The aims of this study were to test the hypothesis that the substrates of xanthine oxidase (XO), xanthine and hypoxanthine, are consumed while the milk is stored in the gland between milkings, and to explore how XO activity responds to bacteria commonly associated with subclinical infections in the mammary gland. Freshly secreted milk was obtained following complete evacuation of the gland and induction of milk ejection with oxytocin. In bacteria-free fresh milk xanthine and hypoxanthine were converted to uric acid within 30 min (T1/2 approximately 10 min), which in turn provides electrons for formation of hydrogen peroxide and endows the alveolar lumen with passive protection against invading bacteria. On the other hand, the longer residence time of milk in the cistern compartment was not associated with oxidative stress as a result of XO idleness caused by exhaustion of its physiological fuels. The specific response of XO to bacteria species and the resulting bacteria-dependent nitrosative stress further demonstrates that it is part of the gland immune system.