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Lesch-Nyhan disease (LND), caused by a deficient salvage purine pathway, is characterized by severe neurological manifestations and uric acid overproduction. However, uric acid is not responsible for brain dysfunction, and it has been suggested that purine nucleotide depletion, or accumulation of other toxic purine intermediates, could be more relevant. Here we show that purine alterations in LND fibroblasts depend on the level of folic acid in the culture media. Thus, physiological levels of folic acid induce accumulation of 5-aminoimidazole-4-carboxamide riboside 5'-monophosphate (ZMP), an intermediary of de novo purine biosynthetic pathway, and depletion of ATP. Additionally, Z-nucleotide derivatives (AICAr, AICA) are detected at high levels in the urine of patients with LND and its variants (hypoxanthine-guanine phosphoribosyltransferase [HGprt]-related neurological dysfunction and HGprt-related hyperuricemia), and the ratio of AICAr/AICA is significantly increased in patients with neurological problems (LND and HGprt-related neurological dysfunction). Moreover, AICAr is present in the cerebrospinal fluid of patients with LND, but not in control individuals. We hypothesize that purine alterations detected in LND fibroblasts may also occur in the brain of patients with LND., Competing Interests: The authors declare no competing interest.

Lesch-Nyhan disease (LND) is a rare X-linked recessive disorder caused by deficiency of the purine salvage enzyme hypoxanthine-guanine phosphoribosyltransferase (HPRT), encoded by the HPRT1. To date, nearly all types of mutations have been reported in the whole gene; however, duplication mutations are rare. We here report the case of a 9-month-old boy with LND. He showed developmental delay, athetosis, and dystonic posture from early infancy, but no self-injurious behaviors. Hyperuricemia was detected, and his HPRT enzyme activity in erythrocytes was completely deficient. A novel duplication mutation (c.372dupT, c.372_374 TTT > c.372_375 TTTT) was identified in exon 4 of the HPRT1, which causes aberrant splicing. This is the third case of a duplication mutation in the HPRT1 that causes splicing error.

Motivation: Mathematical modeling and optimization have been used for detecting enzyme targets in human metabolic disorders. Such optimal drug design methods are generally differentiated as two stages, identification and decision-making, to find optimal targets. We developed a unified method named fuzzy equal metabolic adjustment to formulate an optimal enzyme target design problem for drug discovery. The optimization framework combines the identification of enzyme targets and a decision-making strategy simultaneously. The objectives of this algorithm include evaluations of the therapeutic effect of target enzymes, the adverse effects of drugs and the minimum effective dose (MED)., Results: An existing generalized mass action system model of human uric acid (UA) metabolism was used to formulate the fuzzy optimization method for detecting two types of enzymopathies: hyperuricemia caused by phosphoribosylpyrophosphate synthetase (PRPPS) overactivity and Lesch-Nyhan syndrome. The fuzzy objectives were set so that the concentrations of the metabolites were as close as possible to the healthy levels. The target design included a diet control of ribose-5-phospahate (R5P). The diet control of R5P served as an extra remedy to reduce phosphate uptake entering the purine metabolic pathway, so that we could obtain a more satisfactory treatment than obtained for those without a diet control. Moreover, enhancing UA excretion resulted in an effective treatment of hyperuricemia caused by PRPPS overactivity. This result correlates with using probenecid and benbromazone, which are uricosuric agents present in current clinical medications. By contrast, the Lesch-Nyhan syndrome required at least three enzyme targets to cure hyperuricemia.

Lesch-Nyhan disease (LND) is caused by complete deficiency of the hypoxanthine-guanine phosphoribosyltransferase enzyme. It is characterized by overproduction of uric acid, jointly with severe motor disability and self-injurious behaviour which physiopathology is unknown. These neurological manifestations suggest a dysfunction in the basal ganglia, and three neurotransmitters have been implicated in the pathogenesis of the disease: dopamine, adenosine and serotonin. All of them are implicated in motor function and behaviour, and act by binding to specific G-protein coupled receptors in the synaptic membrane where they seem to be integrated through receptor-receptor interactions. In this work we have confirmed at protein level the previously reported increased expression of DRD5 and the variably aberrant expression of ADORA2A, in LND PBL respect to control PBL. We have also described, for the first time, a decreased expression and protein level of 5-HTR1A in LND PBL respect to control PBL. If these results were confirmed in the Lesch-Nyhan patients basal ganglia cells, this would support the hypothesis that pathogenesis of neurological manifestations of Lesch-Nyhan patients may be related to an imbalance of neurotransmitters, rather than to the isolated disturbance of one of the neurotransmitters, and this fact should be taken into account in the design of pharmacologic treatment for their motor and behavioural disturbances.

Inherited mutation of hypoxanthine guanine phosphoribosyltransferase (HPRT) gives rise to Lesch-Nyhan syndrome or HPRT-related gout. On the other hand, PRPS1 mutations cause PRPP synthetase superactivity associated with hyperuricemia and gout, sometimes including neurodevelopmental abnormalities. We have identified two mutations in two Lesch-Nyhan families after our last report. One of them, a new single nucleotide substitution (130G>T) resulting in a missense mutation D44Y was detected in exon 2 of HPRT1. RT-PCR amplification showed not only a cDNA fragment with normal size, but also a small amount of shorter fragment skipping exons 2 and 3. The other missense mutation F74L (222C > A) was detected in a Japanese patient but has been reported previously in European families. In four hyperuricemic patients with mild neurological abnormality, no mutations responsible for partial HPRT deficiency were identified in HPRT1. In these four patients, we also performed molecular analysis of PRPS1, but no mutations in PRPP synthetase were found.

The genetic deficiency of hypoxanthine-guanine phosphoribosyltransferase (HPRT), located on the X chromosome, causes a severe neurological disorder in man, known as Lesch-Nyhan disease (LND). The enzyme HPRT is part of the savage pathway of purine biosynthesis and catalyzes the conversion of hypoxanthine and guanine to their respective nucleotides, IMP and GMP. HPRT deficiency is associated with a relatively selective dysfunction of brain dopamine systems. Several metabolites that accumulate in the patients (phosphoribosylpyrophosphate (PRPP), hypoxanthine, guanine, xanthine, and Z-nucleotides) have been proposed as toxic agents in LND. Some authors have pointed that Z-riboside, derived from the accumulation of ZMP, could be the toxic metabolite in LND. However, the available experimental data support a better hypothesis. I suggest that ZMP (and not Z-riboside) is the key toxic metabolite in LND. ZMP is an inhibitor of the bifunctional enzyme adenylosuccinate lyase, and a deficiency of this enzyme causes psychomotor and mental retardation in humans. Moreover, it has been reported that ZMP inhibits mitochondrial oxidative phosphorylation and induces apoptosis in certain cell types. ZMP is also an activator of the AMP-activated protein kinase (AMPK), a homeostatic regulator of energy levels in the cell. The AMPK has been implicated in the regulation of cell viability, catecholamine biosynthesis and cell structure. I propose that accumulation of ZMP will induce a pleiotropic effect in the brain by (1) a direct inhibition of mitochondrial respiration and the bifunctional enzyme adenylosuccinate lyase, and (2) a sustained activation of the AMPK which in turns would reduce cell viability, decrease dopamine synthesis, and alters cell morphology. In addition, a mechanism to explain the accumulation of ZMP in LND is presented. The knowledge of the toxic metabolite, and the way it acts, would help to design a better therapy.

The X-chromosomal gene for hypoxanthine-guanine phosphoribosyltransferase (HPRT), first recognized through its human germinal mutations, quickly became a useful target for studies of somatic mutations in vitro and in vivo in humans and animals. In this role, HPRT serves as a simple reporter gene. The in vivo mutational studies have concentrated on peripheral blood lymphocytes, for obvious reasons. In vivo mutations in T cells are now used to monitor humans exposed to environmental mutagens with analyses of molecular mutational spectra serving as adjuncts for determining causation. Studies of the distributions of HPRT mutants among T cell receptor (TCR) gene-defined T cell clones in vivo have revealed an unexpected clonality, suggesting that HPRT mutations may be probes for fundamental cellular and biological processes. Use of HPRT in this way has allowed the analyses of V(D)J recombinase mediated mutations as markers of a mutational process with carcinogenic potential, the use of somatic mutations as surrogate markers for the in vivo T cell proliferation that underlies immunological processes, and the discovery and study of mutator phenotypes in non-malignant T cells. In this last application, the role of HPRT is related to its function, as well as to its utility as a reporter of mutation. Most recently, HPRT is finding use in studies of in vivo selection for in vivo mutations arising in either somatic or germinal cells.

A number of inherited or drug-induced metabolic disorders involving dysfunctions in purines and pyrimidines are strongly associated with neurological dysfunction, e.g., Lesch Nyhan syndrome. Such disorders have been studied extensively using biochemical and molecular techniques in order to examine how such defects occur, sometimes using in vitro models based upon cultured neuroblastoma cell lines. However, these metabolic dysfunctions may manifest their effects in other ways, such as impaired synaptic transmission and gross abnormalities in neuronal growth and differentiation. This review outlines the latter novel facet of purine research. It is proposed that by employing cell imaging techniques and cultured neuroblastoma cell lines, believed to model the nervous system, significant insights into how inherited disorders of purine metabolism affect neuronal development can be obtained. This review provides an example of the application of these techniques to understand the etiology of Lesch Nyhan syndrome, and encourages further study of the role of purines and pyrimidines in the development of the nervous system.

Extracellular purines have essential roles in neuronal development; hence, disruptions in their metabolism as reported in Lesch Nyhan syndrome (LNS) could result in developmental abnormalities. The deficiency of hypoxanthine-guanine phosphoribosyl transferase (HGPRT) in LNS leads to increased hypoxanthine and uric acid production. We have reported that HGPRT-deficient B103-4C neuroblastoma, a neuronal model of LNS, proliferated less and differentiated more than their HGPRT-positive B103 counterparts. Here, we sought to determine whether differences in proliferation and differentiation would occur when these cells were cultured in the presence of hypoxanthine or in a hypoxanthine-/serum-free chemically defined media (NBMN2). In media with 1% serum, hypoxanthine (50 microM) significantly increased the proliferation of both cell lines with a greater effect on B103-4C cells. In 1% serum media, hypoxanthine increased differentiation of B103 but decreased B103-4C differentiation. In NBMN2, B103 proliferated far more than B103-4C, but both cell types differentiated to the same extent. These results are interpreted to suggest that elevated levels of central nervous system (CNS) hypoxanthine as reported in LNS may affect neuronal development, and to implicate hypoxanthine and abnormal neuronal development as causative factors in the etiology of LNS., (Copyright 2001 Wiley-Liss, Inc.)

In hypoxanthine (guanine) phosphoribosyltransferase- (HPRT; EC 2.4.2.8) deficient lymphoblasts, ATP but not nicotinamide-adenine dinucleotide coenzyme concentrations are reduced by limited nutrition. Such reduced ATP concentrations are correlated with reduced poly(ADP-ribose) synthetase (polyADPRT; EC 2.4.2.30) activity; this reduces the breakdown of nicotinamide-adenine dinucleotide coenzymes and thus explains their normal intracellular concentrations. Since reductions in poly(ADP-ribose) synthetase activity reduce DNA repair, alterations in DNA could accumulate even in non-multiplying cells such as neurons, especially in the continuously active 'respiratory centre'. Our Lesch-Nyhan patients suffered respiratory deaths between 15 and 20 years of age.

The distributional and activity changes of hypoxanthine guanine phosphoribosyltransferase (HGPRT) were investigated in the developing mouse brain. The HGPRT activity level was low at birth, increased rapidly during the first 7 days of life, and underwent a gradual increase thereafter to the mature level. Polyclonal antibody against HGPRT purified from mouse brain was prepared for immunohistochemical demonstration of the enzyme during brain development. In the cerebellum, part of the Purkinje cells was consistently immunostained throughout growth, and the presence of HGPRT was observed in the dendrites of mature Purkinje cells. The most dominant change in HGPRT localization was observed in the hippocampus. Little HGPRT was detectable in the newborn mouse hippocampus. At postnatal day 7, cytoplasmic HGPRT appeared sporadically in the granular cells independently of the region of the hippocampus. The number of positive immunoreactive cells increased with growth, and the dendrites of granular cells were also immunostained on postnatal day 28. Further immunostaining was noted in the granule cells of the dentate gyrus on postnatal day 35. The above results suggest that HGPRT may play an important role in the developing hippocampus. Further investigations of the HGPRT in the human hippocampus may help to clarify the mechanism underlying the neurological disorders encountered in the Lesch-Nyhan syndrome.

In dealing with a child with a structural defect, an overall diagnosis must be formulated. Such a diagnosis makes it possible to provide genetic counseling for the parents and an accurate prediction relative to such a child's future development. Because there are a great many abnormalities involving the head and neck, it is hoped that the approach set forth in this article will allow for a systematic narrowing of the diagnostic possibilities. Nomenclature is established. Prenatal-onset defects are described, including both single primary defects (malformations and deformations) and multiple malformation syndromes (chromosomal abnormalities, genetic disorders, defects resulting from teratogenic factors, and disorders of unknown etiology). Genetic and environmental factors of postnatal developmental problems are also discussed.

Neonatal-6-OHDA treated rats given L-DOPA after a decarboxylase inhibitor showed a high incidence of self-mutilation behavior (SMB) and self-biting. These behaviors were not observed in adult-6-OHDA-treated rats or in controls. Since inhibition of dopamine-beta-hydroxylase did not prevent or inhibit the SMB exhibited in neonatal-6-OHDA-treated rats after L-DOPA, norepinephrine is not likely to be contributing to this response. The age dependent effects observed are consistent with the hypothesis that neonatal reduction of dopamine-containing fibers is responsible for the SMB susceptibility observed in Lesch-Nyhan disease, making the neonatal-6-OHDA-treated rat a model of this neurological syndrome.

Administration of L-dopa or apomorphine to neonatal and adult 6-hydroxydopamine (6-OHDA)-treated rats resulted in different behavioral responses depending on the age at which dopaminergic fibers were destroyed. When neonatal 6-OHDA-treated rats were tested as adults, they exhibited marked stereotypies, self-biting and self-mutilation behavior (SMB) when given these dopamine agonists. Self-biting as well as the incidence of SMB in neonatal 6-OHDA-treated rats showed dose-related changes between 10 and 100 mg/kg of L-dopa. This SMB and self-biting after L-dopa was observed as early as 22 to 24 days of age. Adult 6-OHDA-treated rats did not exhibit SMB or self-biting to L-dopa (100 mg/kg) or apomorphine (10 mg/kg), but did display paw treading and head nodding--behaviors not observed in neonatal 6-OHDA-treated rats. In addition, the locomotor response to apomorphine (1 mg/kg) was significantly greater in adult 6-OHDA-treated rats than in neonatal 6-OHDA-treated rats. Brain dopamine was reduced markedly in striatum, nucleus accumbens and olfactory tubercles in both 6-OHDA treatment groups with the reduction being slightly greater in rats treated with 6-OHDA neonatally. Serotonin content was elevated in striatum of rats treated neonatally with 6-OHDA, but not in adult 6-OHDA-treated rats. SMB and behaviors observed after L-dopa in rats treated neonatally with 6-OHDA were not apparent after L-dopa in rats with brain serotonin or norepinephrine reduced. Rats with brain dopaminergic fibers destroyed neonatally exhibited self-biting and SMB after L-dopa, suggesting that neonatal reduction of this amine is responsible for the SMB and self-biting in neonatal 6-OHDA-treated rats. 5-Hydroxytryptophan administration to neonatal 6-OHDA-treated rats did not induce SMB, indicating that release of serotonin by L-dopa is not responsible for this behavior. Because inhibition of dopamine-beta-hydroxylase did not alter the SMB response to L-dopa observed in neonatal 6-OHDA-treated rats, norepinephrine synthesized from L-dopa does not appear to contribute to the response. High doses of a decarboxylase inhibitor sufficient to inhibit conversion of dopa to dopamine in brain did not reduce the incidence of SMB. Administration of haloperidol (1 mg/kg) reduced the incidence of SMB, but did not antagonize the self-biting or the taffy pulling exhibited by L-dopa. In contrast, cisflupentixol completely blocked the SMB and self-biting induced by L-dopa.(ABSTRACT TRUNCATED AT 400 WORDS)

Three patients with the Lesch-Nyhan syndrome were found to have normal delayed hypersensitivity, peripheral-blood T-lymphocyte counts, lymphocyte responses to P.H.A., and serum IgM, IgA, and IgE levels. However, the percentages of B-lymphocytes, IgG levels, serum-isohaemagglutinin titres, and lymphocyte responses to pokeweed mitogen (P.W.M.) were subnormal. These observations suggest that activity of the salvage pathway of purine synthesis catalysed by hypoxanthine-guanine phosphoribosyl transferase (H.G.P.R.T.) is not required for the responses of T-lymphocytes to mitogenic or antigenic stimulation, but may contribute to the proliferation and function of B lymphocytes. The major role of the de-novo pathway of purine synthesis in human lymphocyte responses to mitogenic or antigenic stimulation is shown by the effects of inhibitors of this pathway, including immunosuppressive agents, and by the effects of congenital deficiency or inhibition of adenosine deaminase.

A phylism is a unit of existence, part of an individual's species heritage. It is neither nature nor nurture, but a phyletically dictated product of the interaction of both at a crucial developmental period which may be either prenatal, postnatal, or both. In the syndrome of autism, the phylism of troopbonding is specifically impaired, and other phylisms may be hypertrophic. The phylism of limerent (lover-lover) pairbonding is not impaired, and it may be the source of discordance with other members of the troop. Shyness is an attenuated manifestation of autism. The etiology of impaired troopbonding may be heritable, and almost certainly will prove to be neurochemical also, probably as an analogue of the Lesch-Nyhan syndrome. The lives of autistic children are subject to secondary shaping by the environment of living.

The results of clinical, experimental psychological and biochemical studies of patients with a subclinical form of Lesch-Nyhan syndrome suggested a reduction of the inhibitory influence of the caudate nucleus due to incomplete genetic defect of Hg-PRTase and related disorders of other types of metabolism in the area of basal ganglia as one of the links of pathogenesis. The pathogenetic therapy is based on maintaining high activity of Hg-PRTase by dietary treatment with limited intake of exogenous purines.

We report the identification of a female patient with the X-linked recessive Lesch-Nyhan syndrome (hypoxanthine phosphoribosyltransferase [HPRT] deficiency). Cytogenetic and carrier studies revealed structurally normal chromosomes for this patient and her parents and demonstrated that this mutation arose through a de novo gametic event. Comparison of this patient's DNA with the DNA of her parents revealed that a microdeletion, which occurred within a maternal gamete and involved the entire HPRT gene, was partially responsible for the disease in this patient. Somatic cell hybrids, generated to separate maternal and paternal X chromosomes, showed that expression of two additional X-linked enzymes, phosphoglycerate kinase and glucose-6-phosphate dehydrogenase, were expressed only in cells that contained the maternal X chromosome, suggesting the presence of a functionally inactive paternal X chromosome. Furthermore, comparison of methylation patterns within a region of the HPRT gene known to be important in gene regulation revealed differences between DNA from the father and the patient, in keeping with an active HPRT locus in the father and an inactive HPRT locus in the patient. Together these data indicate that nonrandom inactivation of the cytogenetically normal paternal X chromosome and a microdeletion of the HPRT gene on an active maternal X chromosome were responsible for the absence of HPRT in this patient.

We have analyzed for purine compounds entering and leaving the liver in lightly anesthetized rabbits and rats and for the export of utilizable purine from liver perfused with oxypurine. The in vivo results indicate that roughly 80% of hypoxanthine, xanthine, and urate is removed in a single passage of blood through liver. Conversely, the adenosine concentration of hepatic venous blood is increased 10-fold over portal or arterial levels. When the liver is isolated and perfused with hypoxanthine there is significant release of adenosine, whether measured quantitatively by microbiological assay or qualitatively by analysis of the radioactive purines released from liver that has been prelabeled with [14C]hypoxanthine. These results provide direct evidence for the clearance of hydroxylated purines and the release of utilizable adenine derivatives by liver.