In vitro OKT3-induced mitogenesis in selenium-deficient patients on a diet for phenylketonuria

Patients with phenylketonuria (PKU) are frequently deficient in the essential trace element selenium (Se), because of their very low protein diet. Using two approaches to investigate T-cell response to proliferative signaling, viz, mitogenesis caused by the monoclonal antibody OKT3 and the plant lectin phytohaemagglutinin (PHA), we demonstrated significantly reduced responses to optimal concentrations of OKT3 in a group of PKU patients with reduced serum Se compared with a normal group (p = 0.0005) and with a group of PKU patients whose serum Se was normal (p = 0.0023). The response of the Se-deficient group to optimal levels of PHA did not differ from that of the normal controls or from that of Se-normal PKU patients. A dose-dependent relationship between serum Se levels and mitogenic response was evident for OKT3 (r = 0.34, p = 0.0154), but not for PHA (r = -0.02, p = 0.9086). We suggest that the reduced response to OKT3 mitogenesis in Se-deficient PKU patients is possibly the consequence of impaired Se-dependent metabolic activity, which affects mitogenic signaling via the T cell antigen receptor (TCR/CD3) complex.     

Plasma molybdenum concentrations in children with and without phenylketonuria

Plasma molybdenum concentrations were determined in children, ages two to 12 yr, with and without phenylketonuria (PKU). Mean plasma molybdenum concentrations did not differ significantly between the children with PKU (1.33±0.5 μg/L) and without PKU (1.75±0.8 μg/L). Plasma molybdenum concentrations in both groups of children ranged from <1 to 3 μg/L. When data from all children were combined and then separated based on gender, mean plasma molybdenum levels did not differ significantly between 9 females (1.56±0.68 μg/L) and 12 males (1.58±0.76 μg/L). Data were also combined and mean (±SD) plasma molybdenum concentrations calculated for age groups. Two children aged 1 to <4 yr had plasma molybdenum concentrations of 1.0 μg/L, and six children aged 4 to <7 yr had mean (±SD) plasma molybdenum concentrations of 1.5±0.8 μg/L. Eleven children aged 7 to <11 yr had a mean plasma molybdenum concentration of 1.7±0.7 μg/L, and two children 11 to <14 yr had plasma molybdenum concentrations of 1 μg/L and 2 μg/L. Plasma molybdenum concentrations did not differ significantly among children in the age groups.     

Plasma micronutrient concentrations in infants undergoing therapy for phenylketonuria

Twenty-seven infants with classical phenylketonuria were evaluated longitudinally for 6 mo while ingesting Phenex^TM -1 Amino Acid Modified Medical Food With Iron as their primary protein source. Intake of selected nutrients and biochemical indices of trace and ultratrace mineral status and plasma retinol and α-tocopherol concentrations were evaluated. The means of iron status indices (complete blood count, plasma ferritin, iron, transferrin saturation, total iron binding capacity) and the plasma concentrations of trace and ultratrace minerals (copper, manganese, molybdenum, selenium, zinc) and plasma retinol and α-tocopherol were in the reference ranges. Vitamin A intakes (r = 0.49,p < 0.05) and plasma retinol-binding protein concentrations (r = 0.42,p < 0.05) were positively correlated with plasma retinol concentrations at 3 mo of study. At 6 mo, concentrations of plasma transthyretin (r = 0.72,p < 0.01) and retinolbinding protein (r = 0.48,p < 0.05) were positively correlated with plasma retinol concentrations. At 6 mo, concentrations of plasma transthyretin (r = 0.52,p < 0.05) were positively correlated with retinol-binding protein concentrations. Phenex-1 supports normal mean iron status indices and mean concentrations of trace and ultratrace minerals, retinol, and α-tocopherol when fed in adequate amounts.     

Simultaneous diagnostic method for phenylketonuria and galactosemia from dried blood spots using high-performance liquid chromatography-pulsed amperometric detection

We developed a simultaneous diagnostic method for phenylketonuria (PKU) and galactosemia through simultaneous determination of phenylalanine (Phe) and galactose (Gal) by high-performance liquid chromatography (HPLC) with pulsed amperometric detection (PAD). The intra- and inter-day precisions were <5.8%, with satisfactory mean recoveries (98.2–105%). For all PKU-positive samples, Phe levels were above the cut-off value (>30.0 mg/L), but Gal levels were nearly zero. For 77% of galactosemia-positive samples, Phe levels were above the cut-off value, but Gal levels were above the cut-off value (>80.0 mg/L) for all samples. Our HPLC-PAD method can reduce the false-positive rate of misdiagnosis for PKU and galactosemia.     

Variations in genotype–phenotype correlations in phenylalanine hydroxylase deficiency in Chinese Han population

Background

The value of genotyping to predict variant phenotypes in patients with phenylalanine hydroxylase (Pah) deficiency is a matter of debate. However, there exists no comprehensive population relationship study focused on the Han Chinese.

Methods

We analyzed genotype–phenotype correlation for 186 different genotypes in 338 unrelated Chinese patients harboring 109 different Pah mutations. Two systems were used in this process. The first was a phenotype prediction system based on arbitrary values (AV) attributed to each mutation. The second was a pair-wise correlation analysis. The observed phenotype for AV analysis was the corresponding metabolic phenotype stratified according to the pretreatment phenylalanine (Phe) value.

Results

We found that the observed phenotype matched the predicted phenotype in 54.41% of 272 patients for whom AV information was available; the highest degree of concordance (61.83%) was found in patients with null/null genotypes, whereas the lowest “concordance rate” (32.69%) was observed for patients with expected mild-PKU phenotype. There are repeated inconsistencies for such mutations as R241C, R243Q, R261Q, V388M, V399V, R408Q, A434D and EX6-96A>G which are associated with variable phenotypes in patients with identical genotype. Significant correlations were disclosed between pretreatment Phe values and predicted residual activity (r = − 0.45643, P < 0.0001) or AV sum (r = − 0.59523, P < 0.0001).

Conclusion

Our study supports the notion that the Pah mutation genotype is the main determinant of metabolic phenotype in most patients in a particular population, and provided novel insights into the values that underpin the subsequent treatment and the prognosis of PKU in Chinese.     

The essential fatty acid status in phenylketonuria patients under treatment

Phenylketonuric patients are on a special diet that lacks certain essential fatty acids. This study evaluates the essential fatty acid status of a group of phenylketonuric patients in the Netherlands undergoing dietary treatment. To this end, the essential fatty acid status of nine phenylketonuria patients was studied. On the basis of age and gender, two control subjects were selected for each patient. The essential fatty acid composition of duplicate food portions and the essential fatty acid status of plasma and erythrocytes were analyzed. Phenylketonuria subjects had a different essential fatty acid profile from their peers, especially concerning the n-3 fatty acids. N-6 and n-3 fatty long-chain polyenes were hardly consumed by phenylketonuria subjects, in contrast to the control subjects. Linoleic acid, on the other hand, was consumed in significantly higher amounts by phenylketonuria subjects and made up about 40% of their daily fat consumption. The essential fatty acid consumption pattern of the phenylketonuria subjects is mirrored by the essential fatty acid concentrations in blood. The essential fatty acid status of the phenylketonuric diet should be improved in order to prevent deficiency in n-3 fatty acids.     

Molecular phylogenetic study of the Iranians based on polymorphic markers

The application of polymorphic markers in construction of phylogenetic trees has been documented. Five polymorphic markers located in the PAH gene region including PAH-BglII, PAH-PvuII(A), PAH-EcoRI, PAH-MspI and PAH-STR were selected for analysis of phylogenetic relationships of the Iranians with 15 other populations of the world. The lowest genetic distance was observed between the Iranians and populations residing in Adygei (an ethnic group of the Russian Caucasus), Russia and Druze (a Middle Eastern group). However, East Asian populations including Han, Japanese and Cambodians, Khmer or the Oceanians (Melanesian, Nasioi) showed high genetic distance with the Iranians. The data suggested that the Iranians might have relatively close evolutionary history with the populations residing in Russia rather than East Asian populations. This study provided the first new molecular insight into the evolutionary history of the Iranian population.     

The spectrum of mutations and methods for their detection in the phenylalanine hydroxylase gene in phenylketonuria patients from the novosibirsk region

Phenylketonuria (PKU) is a widespread autosome recessive hereditary disease caused by a deficiency of the liver enzyme phenylalanine hydroxylase, which results in distortion of metabolism of phenylalanine and accumulation of toxic metabolites. The knowledge of molecular bases of PKU is of a high social importance as it enables phenotypic correction of the disease in the case of its early diagnostics. This disease is known to be associated with mutations in the phenylalanine hydroxylase gene, the distribution and mutation spectrum having pronounced ethnic and regional features. We studied the spectrum of mutations in the phenylalanine hydroxylase gene in a group of patients with PKU from the Novosibirsk region to reveal 10 missense point mutations, 1 mutation in the splice donor site, and 1 microdeletion. For these mutations, most widely distributed in the region, we used straightforward detection methods basing on the restriction fragment length polymorphism (RFLP), artificial constructed restriction sites (ACRS) PCR, and denaturing gradient gel electrophoresis (DGGE).     

Altered brain protein expression profiles are associated with molecular neurological dysfunction in the PKU mouse model

Phenylketonuria (PKU), if not detected and treated in newborns, causes severe neurological dysfunction and cognitive and behavioral deficiencies. Despite the biochemical characterization of PKU, the molecular mechanisms underlying PKU‐associated brain dysfunction remain poorly understood. The aim of this study was to gain insights into the pathogenesis of this neurological damage by analyzing protein expression profiles in brain tissue of Black and Tan BRachyury‐PahEnu2 mice (a mouse model of PKU). We compared the cerebral protein expression of homozygous PKU mice with that of their heterozygous counterparts using two‐dimensional difference gel electrophoresis analysis, and identified 21 differentially expressed proteins, four of which were over‐expressed and 17 under‐expressed. An in silico bioinformatic approach indicated that protein under‐expression was related to neuronal differentiation and dendritic growth, and to such neurological disorders as progressive motor neuropathy and movement disorders. Moreover, functional annotation analyses showed that some identified proteins were involved in oxidative metabolism. To further investigate the proteins involved in the neurological damage, we validated two of the proteins that were most strikingly under‐expressed, namely, Syn2 and Dpysl2, which are involved in synaptic function and neurotransmission. We found that Glu2/3 and NR1 receptor subunits were over‐expressed in PKU mouse brain. Our results indicate that differential expression of these proteins may be associated with the processes underlying PKU brain dysfunction, namely, decreased synaptic plasticity and impaired neurotransmission.

     

苯丙酮尿症分子遗传学研究进展

苯丙酮尿症是由于苯丙氨酸羟化酶基因突变引起的常染色体隐性遗传病。文章综述了苯丙酮尿症中的苯丙氨酸羟化酶基因的定位、结构、突变、调控以及突变基因的体外表达和苯丙氨酸羟化酶的三维结构特点等分子遗传学进展,阐述了苯丙氨酸羟化酶基因的突变对苯丙氨酸羟化酶的体外表达及其三维结构的影响, 以及部分基因型与表型相关的分子机制。 Abstract: Phenylketonuria(PKU) is one kinds of autusomal recessive disease caused by phenylalanine hydroxylase(PAH) gene mutation. This article reviews the recent molecular heredity progress on the phenylalanine hydroxylase gene’s orientation、structureand gene mutation and gene regulation. At same time, mutation gene in vitro expression and the character of 3D structure of PAH in PKU are involved. In this paper, also discussed the inflence of vitro expression and 3D protein structure by gene mutations and the molecular mechanism of the relationship between genotype and phenotype in PKU patient.