Intracellular concentrations of phenylalanine,tyrosine and α-aminobutyric acid in 13 homozybotes and 19 heterozygotes for phenylketonuria (PKU) compared with 26 normals

Summary Intracellular concentrations of phenylalanine, tyrosine, -aminobutyric acid, and seven other aminoacids (glycine, alanine, valine, cystine, methionine, isoleucine, leucine) were measured in lymphocytes of 13 homozygotes and 19 heterozygotes for phenylketonuria and in lymphocytes of 26 normals. Intracellular concentrations for phenylalanine, tyrosine, and -aminobutyric acid were significantly higher in homo- and heterozygotes than in normals (P<0.001; P<0.01). For the other seven aminoacids there were no or only questionable differences. Between homo-and heterozygotes there was no difference in any of the aminoacids. The intracellular phenylalanine: tyrosine ratio was essentially the same in all three groups of individuals. There was no correlation between intracellular phenylalanine above or below 10nmol/10⁶ cells and IQ in heterozygotes. The same is true for phenylalanine: tyrosine ratio greater or smaller than 1. In homozygotes there was no correlation between intracellular phenylalanine and age—to which DQ/IQ is correlated. There was no significant difference in intracellular phenylalanine between homozygotes with blood levels above and below 908 mol/l (15 mg/100 ml) at the time of blood sampling and no correlation between intra- and extracellular phenylalanine concentrations.Among the 26 normals there were only two with intracellular phenylalanine above 10 nmol/10⁶ cells, both showing phenylalanine loading test curves suggestive of heterozygosity.The results are discussed and important functions of the cell wall are proposed. The formation of an abnormal unknown intracellular metabolite being the real noxious agent could explain the incomparably different degrees of brain dysfunction in individuals with equal though elevated intracellular phenylalanine concentrations, i.e., homozygotes and heterozygotes for PKU.     

Intracellular phenylalanine and tyrosine concentrations in 19 heterozygotes for phenylketonuria (PKU) and 26 normals

Summary Intracellular phenylalanine and tyrosine was determined in lymphocytes of 19 heterozygotes (parents) for PKU and in 26 randomly collected apparently normal persons. In cells from the heterozygotes the concentrations of both phenylalanine and tyrosine were higher than in those from the normals, the difference being statistically highly significant. It is argued that this could be responsible for the slight, though statistically significant, intellectual inferiority of heterozygotes for PKU.     

Relationship between phenylalanine tolerance and psychological characteristics of phenylketonuric families

The purpose of this study was to find out how genetic and biochemical limitations influence psycho-social performance and to partially test the validity of justification theory. The ability to convert phenylalanine to tyrosine was compared with intellectual and personality characteristics in PKU family members. Each of the tested persons was given an oral dose of phenylalanine, the Shipley-Hartford Intelligence Test, and the Minnesota Multiphasic Personality Inventory (MMPI). Only those persons with reading ability at the sixth grade level or higher were tested. Eighty-six persons were tested: fifteen PKUs, forty-three siblings, and twenty-eight parents. A comparison was made among parents, PKUs, and the siblings. Siblings with the higher 2/3's of P2/T ratios were contrasted with those with the lowest 1/3 of ratios on measures of intelligence and psychopathology. Statistical analyses of the data reflected a trend in support of the justification theory. PKUs had significantly lower intelligence than their sibs and parents. The PKUs' mean IQ was 95 (homozygotes born of heterozygotes), followed by the upper 2/3's sibling mean IQ of 105 (heterozygotes born of nonheterozygote mothers). The lower 1/3 siblings' mean IQ was 107 (nonheterozygotes born from heterozygote mothers), and finally, the parents' mean IQ was 109 (heterozygotes, among them 50% were born from nonheterozygote mothers). The latter three mean IQs are not significantly different from each other. The personality tests revealed a trend toward more abnormality in PKUs than in their heterozygote siblings. The lowest rate of abnormality occurred in the nonheterozygote sibling group; that rate was significantly lower than in all other groups. The parents had the highest absolute rate of personality abnormality, but statistically so compared to the low-ratio siblings.     

Phenylketonuria heterozygote detection in families with affected children

Improved approaches to the problem of heterozygote detection for phenylketonuria (PKU) were developed in this study. The discrimination was based on 85 obligate heterozygotes and 45 controls who were neither pregnant nor on birth control medication. The best separation between hetrozygotes and normals was achieved with a linear discriminant function involving the logarithms of the serum concentrations of phenylalanine, tyrosine, and tryptophan. The theoretical overlap area between the distributions of heterozygotes and controls based on the above function, was 3.75%. In the 19 obligate hetrozygotes and 13 controls who were either pregnant or on birth control medication, the best separation was achieved with a linear discriminant function involving the logarithms of the serum concentrations of phenylalanine and tyrosine. The theoretical overlap area was 8.23%. The genetic accuracy of the discriminant function was confirmed by testing the results with parental-child exclusions, segregation analysis, and the frequency of heterozygosity in nonrelated collateral spouses. Finally, there was evidence suggesting that the antihypertensive agent, aldomet, alters serum tyrosine and tryptophan levels.     

Assessment of chronic gamma radiosensitivity as an in vitro assay for heterozygote identification of ataxia-telangiectasia

Ataxia-telangiectasia (A-T) is a rare human autosomal recessive disorder characterized by, among other symptoms, catastrophic reaction to conventional radiotherapy. A-T heterozygotes are clinically asymptomatic and their fibroblasts are intermediate in radiosensitivity between homozygotes and normals. We have attempted to identify heterozygotes by assaying for cellular hypersensitivity to chronic gamma irradiation. Cultured dermal fibroblast strains from 13 control subjects and 55 members from a large Amish pedigree segregating for A-T were assayed for loss of colony-forming ability (CFA) in response to 137Cs gamma radiation delivered at a dose rate of 0.8 cGy/min. For each strain, multiple dose-response curves were summarized in a composite D10 value (dose, in cGy, reducing colony survival to 10%). The D10's of the clinically normal controls and of those pedigree members with known A-T genotype formed a trimodal distribution, with the seven obligate heterozygotes displaying an average value (516 cGy) intermediate between that of the 10 healthy controls (797 cGy) and that of the two affected patients (154 cGy). The D10's were modeled statistically using Gaussian penetrance functions. The most parsimonious model yielded a significant difference in D10 means for heterozygotes and normal homozygotes, a significant donor age effect, but no sex effect. We compared probabilistic identification of heterozygotes based on D10 values with identification based on linkage data for two markers, THY1 and D11S144, closely linked to the A-T gene. This comparison revealed that the D10 data were appreciably less informative than the linked markers. Indeed, the extensive overlap between D10 values for heterozygotes and normal homozygotes precludes the use of postirradiation CFA for either accurate identification of heterozygotes or chromosomal mapping of the A-T gene.     

Serum concentrations of vitamin D-binding protein (Group-specific component) in cystic fibrosis

Summary Vitamin D-binding protein (DBP) concentrations were determined in the sera of 90 cystic fibrosis homozygotes, 57 obligate heterozygotes, and 46 normal controls. Very significantly lower mean concentrations were found in the sera of CF homozygotes compared with both heterozygotes and controls (P<0.01, Wilcoxon Rank Sums Test). Subdivision of the samples by Gc phenotype showed that this relationship held true both in the Gc1 and Gc2-1 phenotypes. The small sample size of the Gc2 genotype makes the significance levels of limited usefulness, but the pattern of variation of DBP levels among CF homozygotes, heterozygotes, and controls was consistent with that observed for the Gc1 and Gc2-1 classes. Haptoglobin levels showed high coefficients of variation when compared among CF homozygotes, obligate heterozygotes, and controls, presumably because of nonspecific elevation in the acute-phase response. Alpha₂-macroglobulin levels were, if anything, slightly elevated in CF homozygotes compared with controls, while albumin levels showed no significant mean differences between these groups. Since the DBP concentration does not vary with age nor with levels of vitamin D and its metabolites, we interpret our results to mean that DBP levels are specifically decreased in cystic fibrosis, perhaps as the result of impaired glycosylation of the protein.A preliminary report of this work appeared in the Proceedings of the 8th International Congress on Cystic Fibrosis.     

Rapid mutation screening of phenylketonuria by polymerase chain reaction-linked restriction enzyme assay and direct sequence of the phenylalanine hydroxylase gene: clinical application in northern Japan and northern China

We describe a simple and technically feasible method for mutation screening of the phenylalanine hydroxylase (PAH) gene and its application to Japanese and Chinese patients with hyperphenylalaninemia. The strategy is based on the identification of a nucleotide substitution by restriction enzyme analysis, coupled with PCR and direct sequencing of exon 7 of the PAH gene. Because the detection of various mutations can proceed simultaneously using the same technique, it is quite rapid and reproducible, making it possible to perform effective molecular diagnosis and carrier screening in most laboratories. Using this procedure, we found that the most common molecular defects were R413P in Hokkaido, Japan (35 %) and R243Q in Heilongjiang, China (50%). R111X, IVS4nt-1, and five mutations in exon 7 (R241C, R243Q, R252W, A259T, and S273P) accounted for 55% of phenylketonuria (PKU) alleles in Hokkaido. In Heilongjiang, the R111X, Y356X, and R408W mutations accounted for 35% of PKU alleles. Clinically, homozygotes or compound heterozygotes of null alleles, which express nonfunctional enzyme activity, were all associated with classic PKU. On the other hand, patients heterozygous for the R241C allele had a benign phenotype of mild hyperphenylalaninemia. The DNA diagnosis in early infancy can predict various PKU phenotypes, and can prove useful in decision-making concerning dietary therapy.     

Large Neutral Amino Acid Supplementation Exerts Its Effect through Three Synergistic Mechanisms: Proof of Principle in Phenylketonuria Mice

Background

Phenylketonuria (PKU) was the first disorder in which severe neurocognitive dysfunction could be prevented by dietary treatment. However, despite this effect, neuropsychological outcome in PKU still remains suboptimal and the phenylalanine-restricted diet is very demanding. To improve neuropsychological outcome and relieve the dietary restrictions for PKU patients, supplementation of large neutral amino acids (LNAA) is suggested as alternative treatment strategy that might correct all brain biochemical disturbances caused by high blood phenylalanine, and thereby improve neurocognitive functioning.

Objective

As a proof-of-principle, this study aimed to investigate all hypothesized biochemical treatment objectives of LNAA supplementation (normalizing brain phenylalanine, non-phenylalanine LNAA, and monoaminergic neurotransmitter concentrations) in PKU mice.

Methods

C57Bl/6 Pah-enu2 (PKU) mice and wild-type mice received a LNAA supplemented diet, an isonitrogenic/isocaloric high-protein control diet, or normal chow. After six weeks of dietary treatment, blood and brain amino acid and monoaminergic neurotransmitter concentrations were assessed.

Results

In PKU mice, the investigated LNAA supplementation regimen significantly reduced blood and brain phenylalanine concentrations by 33% and 26%, respectively, compared to normal chow (p<0.01), while alleviating brain deficiencies of some but not all supplemented LNAA. Moreover, LNAA supplementation in PKU mice significantly increased brain serotonin and norepinephrine concentrations from 35% to 71% and from 57% to 86% of wild-type concentrations (p<0.01), respectively, but not brain dopamine concentrations (p = 0.307).

Conclusions

This study shows that LNAA supplementation without dietary phenylalanine restriction in PKU mice improves brain biochemistry through all three hypothesized biochemical mechanisms. Thereby, these data provide proof-of-concept for LNAA supplementation as a valuable alternative dietary treatment strategy in PKU. Based on these results, LNAA treatment should be further optimized for clinical application with regard to the composition and dose of the LNAA supplement, taking into account all three working mechanisms of LNAA treatment.     

Compound heterozygosity in nonphenylketonuria hyperphenylalanemia: the contribution of mutations for classical phenylketonuria

Hyperphenylalaninemia (HPA) results from defective hydroxylation of phenylalanine in the liver, in most cases because of defective phenylalanine hydroxylase. HPA is highly variable, ranging from moderate elevation of plasma phenylalanine with no clinical consequences to a severe disease, classical phenylketonuria (PKU). Non-PKU HPA was found in excess of PKU in Israel, while the opposite is true in Europe. To study the genetic basis of non-PKU HPA, we performed haplotype analysis at the phenylalanine hydroxylase locus in 27 families with non-PKU HPA. All individuals with this condition were compound heterozygotes. In six of these families, in which both PKU and non-PKU HPA were segregating, haplotype analysis showed that non-PKU HPA resulted from compound heterozygosity for a PKU mutation and a second mutation, with milder effect, which is probably expressed only when it interacts with the severe mutation. The involvement of PKU mutations in non-PKU HPA was further demonstrated in Jewish Yemenite families with non-PKU HPA, in which the individuals with this condition were carriers of the single PKU allele which exists in this community. In addition, two previously known PKU point mutations (R261Q and R408W) were found in individuals with non-PKU HPA. These mutations are associated, in our population, with the same haplotypes as those with which it is associated in Europe. Based on the above-mentioned genetic model for non-PKU HPA, successful prenatal diagnosis of this condition was performed in one family.(ABSTRACT TRUNCATED AT 250 WORDS)     

Growth induction in cystic fibrosis fibroblasts with low dexamethasone concentrations. Experience with application to genotyping

Summary Dexamethasone (DM) resistance was evaluated in fibroblasts from a pool of five patients with cystic fibrosis (CF) homozygotes, ten of their parental obligate heterozygotes, and seventeen age-matched controls of both sexes. The CF heterozygotes showed a mean DM resistance greater than homozygotes and both groups exhibited a higher mean DM resistance at every DM concentration than controls. However, substantial interassay variability rendered these differences in the total pooled data to non-significance. One control showed a consistently increased resistance and was possibly a covert heterozygote. It was concluded that the phenomenon of DM resistance was exhibited by CF heterozygotes and homozygotes but was not discrete enough for genotyping in the prenatal diagnosis of CF.     

In vivo chromosomal instability in ataxia-telangiectasia homozygotes and heterozygotes

Summary The exfoliated cell micronucleus test was used to monitor in vivo chromosomal instability in a population comprised of five ataxia-telangiectasia (A-T) homozygotes and seven obligate heterozygotes (parents of A-T patients). This assay was previously validated as a procedure for quantifying non-invasively carcinogen-induced chromosomal aberrations occurring in vivo in epithelial tissues of both the oral cavity and the urinary bladder. The procedure involved taking airdried smears of three sites in the oral cavity of each examined individual. Desquamated urinary bladder cells were collected by centrifugation of freshly voided urine samples. Frequencies of exfoliated cells in these preparations were determined and compared with control values (individuals with no genetic chromosomal instability and no known carcinogene exposure) for these sites. Exforliated cell micronucleus (MEC) frequencies were elevated 5- to 14-fold in samples from the A-T homozygotes. This elevation in MEC frequency occurred for both the oral cavity and urinary bladder. Five out of the seven obligate A-T heterozygotes had an elevated MEC frequency in samples from the oral cavity. In addition, all examined urine samples from A-T heterozygotes contained an elevated percentage of micronucleated cells. These data suggest that this assay is suitable for in vivo monitoring of groups of individuals in which genetically produced chromosomal damage occurs. The possibility of A-T heterozygote detection with this simple procedure is of particular significance, since such individuals are believed to comprise up to 1% of the general population, and have been identified as being at elevated risk for cancer.     

Oxidative Stress in Phenylketonuria: What is the Evidence?

Phenylketonuria (PKU) is an inborn error of amino acid metabolism caused by severe deficiency of phenylalanine hydroxylase activity, leading to the accumulation of phenylalanine and its metabolites in blood and tissues of affected patients. Phenylketonuric patients present as the major clinical feature mental retardation, whose pathomechanisms are poorly understood. In recent years, mounting evidence has emerged indicating that oxidative stress is possibly involved in the pathology of PKU. This article addresses some of the recent developments obtained from animal studies and from phenylketonuric patients indicating that oxidative stress may represent an important element in the pathophysiology of PKU. Several studies have shown that enzymatic and non-enzymatic antioxidant defenses are decreased in plasma and erythrocytes of PKU patients, which may be due to an increased free radical generation or secondary to the deprivation of micronutrients which are essential for these defenses. Indeed, markers of lipid, protein, and DNA oxidative damage have been reported in PKU patients, implying that reactive species production is increased in this disorder. A considerable set of data from in vitro and in vivo animal studies have shown that phenylalanine and/or its metabolites elicit reactive species in brain rodent. These findings point to a disruption of pro-oxidant/antioxidant balance in PKU. Considering that the brain is particularly vulnerable to oxidative attack, it is presumed that the administration of appropriate antioxidants as adjuvant agents, in addition to the usual treatment based on restricted diets or supplementation of tetrahydrobiopterin, may represent another step in the prevention of the neurological damage in PKU.     

Intellectual level (IQ) in heterozygotes for phenylketonuria (PKU)

Summary There is a statistically significant difference in the IQ's of PKU and histidinemia parents. The difference is due entirely to the verbal part of the Hamburg-Wechsler test. There is no significant difference in performance. The heterozygous state of histidinemia does not seem to bear an intellectual (evolutionary) advantage, since the IQ's of histidinemia parents show the same distribution as a normal population. In early and mostly well-treated PKU patients, the same slight deficit in verbal IQ appears with increasing age (changing test methods). These patients, simultaneously tested at 4 years of age with the Bühler-Hetzer and Kramer tests, exhibit a statistically significant difference between the results in favor of the less verbal Bühler-Hetzer. Since heterozygots, for PKU never have elevated phenylalanine blood levels, and because tryosine deficiency as argued by others seems highly improbable, we believe that the PKU gene has a more direct action on (or in) at least certain ganglion cells, lowering the verbal IQ slightly, but significantly. This action is not reflected by phenylalanine increase in the extracellular space in heterozygots and is not abolished by dietary treatment in homozygous PKU patients. The major damage in PKU patients must be due to chronic phenylalanine poisoning, which deteriorates cells and/or functions on a much larger scale, because it can be easily prevented by decreasing the phenylalanine blood level with correct dietary treatment.     

Maternal phenylketonuria. Review with emphasis on pathogenesis

Maternal phenylketonuria (PKU) refers to fetal damage from PKU in the pregnant woman. The progeny from such pregnancies are almost always microcephalic and mentally subnormal and have an increased frequency of congenital heart disease and low birth weight. Treatment with a phenylalanine-restricted diet, if begun before conception, seems to protect the fetus. The degree of protection is much less if dietary treatment is delayed until the pregnancy is in progress. The origin of fetal damage in maternal PKU is not known. Due to placental concentration of amino acids, the fetus is exposed to a higher concentration of phenylalanine than that in the mother, but it is not certain that phenylalanine is the toxic agent. Animal models made hyperphenylalaninemic by the administration of phenylalanine, often accompanied by a phenylalanine hydroxylase inhibitor, do not reproduce the full maternal PKU syndrome; but fetuses and newborns from these models have had reduced growth of the body and brain, and offspring later may show evidence of impaired learning ability.     

Experimental Evidence that Phenylalanine Provokes Oxidative Stress in Hippocampus and Cerebral Cortex of Developing Rats

High levels of phenylalanine (Phe) are the biochemical hallmark of phenylketonuria (PKU), a neurometabolic disorder clinically characterized by severe mental retardation and other brain abnormalities, including cortical atrophy and microcephaly. Considering that the pathomechanisms leading to brain damage and particularly the marked cognitive impairment in this disease are poorly understood, in the present study we investigated the in vitro effect of Phe, at similar concentrations as to those found in brain of PKU patients, on important parameters of oxidative stress in the hippocampus and cerebral cortex of developing rats. We found that Phe induced in vitro lipid peroxidation (increase of TBA-RS values) and protein oxidative damage (sulfhydryl oxidation) in both cerebral structures. Furthermore, these effects were probably mediated by reactive oxygen species, since the lipid oxidative damage was totally prevented by the free radical scavengers α-tocopherol and melatonin, but not by L-NAME, a potent inhibitor of nitric oxide synthase. Accordingly, Phe did not induce nitric oxide synthesis, but significantly decreased the levels of reduced glutathione (GSH), the major brain antioxidant defense, in hippocampus and cerebral cortex supernatants. Phe also reduced the thiol groups of a commercial GSH solution in a cell-free medium. We also found that the major metabolites of Phe catabolism, phenylpyruvate, phenyllactate and phenylacetate also increased TBA-RS levels in cerebral cortex, but to a lesser degree. The data indicate that Phe elicits oxidative stress in the hippocampus, a structure mainly involved with learning/memory, and also in the cerebral cortex, which is severely damaged in PKU patients. It is therefore presumed that this pathomechanism may be involved at least in part in the severe cognitive deficit and in the characteristic cortical atrophy associated with dysmyelination and leukodystrophy observed in this disorder.     

Dehydrogenase based reagentless biosensor for monitoring phenylketonuria

Phenylketonuria (PKU) is a disease characterized by an inability to metabolize the amino acid l-phenylalanine. The resulting buildup leads to brain damage and ultimately mental retardation in children if their phenylalanine intake is not carefully controlled. The National Institutes of Health recently suggested that people with PKU monitor their phenylalanine levels throughout their life and be put on a low phenylalanine diet. As an alternative approach to analysis using blood, this paper describes the first reagentless dehydrogenase based sensor for the determination of phenylalanine in human urine. The clinical range of phenylalanine in human urine is 20-60mM for people with PKU. Although most clinical analysis is performed using blood, urine was chosen due to its high concentrations of phenylalanine in phenylketonurics, as well as its simple, safe, and painless collection. The sensor is comprised of a carbon paste electrode with nicotinamide adenine dinucleotide (NAD(+)), phenylalanine dehydrogenase (PDH), uricase, and an electron mediator, 3,4-dihydroxybenzaldehyde (3,4-DHB), all mixed into the paste. The electron mediator reacts with the electrode surface to produce two redox species, which catalytically oxidize NADH. The behavior of the electron mediator mixed into a carbon paste electrode has not been previously investigated. Cyclic voltammetry was used to characterize the sensor's response to NADH, and with the addition of PDH and NAD(+) to the paste, its response to phenylalanine in human urine. The limit of detection for phenylalanine is 0.5mM (S/N=3).     

The effects of n-3 and n-6 polyunsaturated fatty acids on plasma lipids and fatty acids of treated phenylketonuric children

Dietary-treated phenylketonuric patients (PKUs) display low levels of long-chain polyunsaturated fatty acids (PUFA) in plasma lipids. In a 6-month clinical trial we observed a decrease of triglycerides and an increase of n-3 long-chain PUFA in plasma of PKUs supplemented with fish oil, while no major differences in respect to the baseline values were found in a group supplemented with blackcurrant oil. A more complete source of long-chain PUFA of both the n-6 and n-3 series should be investigated for dietary supplementation of PKU patients.     

Measurement of phenyllactate, phenylacetate, and phenylpyruvate by negative ion chemical ionization-gas chromatography/mass spectrometry in brain of mouse genetic models of phenylketonuria and non-phenylketonuria hyperphenylalaninemia

Phenylketonuria (PKU) (OMIM 261600) is the first Mendelian disease to have an identified chemical cause of impaired cognitive development. The disease is accompanied by hyperphenylalaninemia (HPA) and elevated levels of phenylalanine metabolites (phenylacetate (PAA), phenyllactate (PLA), and phenylpyruvate (PPA)) in body fluids. Here we describe a method to determine the concentrations of PAA, PPA, and PLA in the brain of normal and mutant orthologous mice, the latter being models of human PKU and non-PKU HPA. Stable isotope dilution techniques are employed with the use of [(2)H(5)]-phenylacetic acid and [2,3, 3-(2)H(3)]-3-phenyllactic acid as internal standards. Negative ion chemical ionization (NICI)-GC/MS analyses are performed on the pentafluorobenzyl ester derivatives formed in situ in brain homogenates. Unstable PPA in the homogenate is reduced by NaB(2)H(4) to stable PLA, which is labeled with a single deuterium and discriminated from endogenous PLA in the mass spectrometer on that basis. The method demonstrates that these metabolites are easily measured in normal mouse brain and are elevated moderately in HPA mice and greatly in PKU mice. However, their concentrations are not sufficient in PKU to be "toxic"; phenylalanine itself remains the chemical candidate causing impaired cognitive development.     

Survival curves of glutathione synthetase deficient human fibroblasts: correlation between radiosensitivity in hypoxia and glutathione synthetase activity

The role of intracellular non-protein bound sulphydryl compounds (NPSH), and in particular that of glutathione (GSH), in the response of cells to ionizing radiation under different O2 concentrations has been assessed using cell strains deficient in glutathione synthetase and exhibiting different NPSH levels. The cell strains used originated from patients with 5-oxoprolinuria and from their relatives (heterozygotes and proficient homozygotes). No correlation has been found between NPSH and GSH concentrations and radiosensitivity under oxic, aerobic and hypoxic conditions. However, a highly significant correlation has been observed between radiosensitivity under hypoxic conditions (and therefore the oxygen enhancement ratio) and the glutathione synthetase activity, suggesting that synthesis of GSH is required after irradiation. In order to explain our results we postulated, beside radical processes, the existence of a GSH-dependent enzymatic repair mechanism for N2 type damage. Hypoxic radio-sensitivity measured with survival curves would result from the interaction of both competition and biochemical repair processes.     

Role of Catalase and Superoxide Dismutase Activities on Oxidative Stress in the Brain of a Phenylketonuria Animal Model and the Effect of Lipoic Acid

Phenylketonuria (PKU) is an inherited metabolic disorder caused by deficiency of phenylalanine hydroxylase which leads to accumulation of phenylalanine and its metabolites in tissues of patients with severe neurological involvement. Recently, many studies in animal models or patients have reported the role of oxidative stress in PKU. In the present work we studied the effect of lipoic acid against oxidative stress in rat brain provoked by an animal model of hyperphenylalaninemia (HPA), induced by repetitive injections of phenylalanine and α-methylphenylalanine (a phenylalanine hydroxylase inhibitor) for 7 days, on some oxidative stress parameters. Lipoic acid prevented alterations on catalase (CAT) and superoxide dismutase (SOD), and the oxidative damage of lipids, proteins, and DNA observed in HPA rats. In addition, lipoic acid diminished reactive species generation compared to HPA group which was positively correlated to SOD/CAT ratio. We also observed that in vitro Phe inhibited CAT activity while phenyllactic and phenylacetic acids stimulated superoxide dismutase activity. These results demonstrate the efficacy of lipoic acid to prevent oxidative stress induced by HPA model in rats. The possible benefits of lipoic acid administration to PKU patients should be considered.