Ahch, the mouse homologue of DAX1: cloning, characterization and synteny with GyK, the glycerol kinase locus.

We cloned the murine full-length cDNA encoding Ahch, the mouse homologue of DAX1 (DSS-AHC Region on Human X Chromosome, Gene1) which is the gene responsible for human X-linked adrenal hypoplasia congenita (AHC) and hypogonadotropic hypogonadism (HH). Sequence analysis revealed that the murine and human cDNAs have 65% aa identity and 75% aa similarity overall. The cysteine residues in the putative DNA binding domain, which may interact with Zn2+ ions to form zinc fingers, are 100% conserved between the two species, indicating that the novel zinc-finger structures in DAX1 may be functional. In addition, mouse interspecific backcrosses show that the Ahch gene is closely linked to the glycerol kinase locus, GyK, on the mouse X chromosome, indicating that the order of the loci is conserved in this syntenic region between mouse and human.

Expression of DAX-1, the gene responsible for X-linked adrenal hypoplasia congenita and hypogonadotropic hypogonadism, in the hypothalamic-pituitary-adrenal/gonadal axis.

DAX-1, an orphan member of the nuclear hormone receptor superfamily, is responsible for X-linked adrenal hypoplasia congenita (AHC) and the frequently associated hypogonadotropic hypogonadism (HH). The entire DAX-1 genomic region has been sequenced and a putative steroidogenic factor-1 response element has been identified in the promoter region of the gene. The purpose of these investigations was to determine if DAX-1 was expressed in the central nervous system, particularly the hypothalamus and pituitary, in order to better understand the relationship of mutations in this gene to HH associated with AHC. We used Northern blot analysis and reverse transcription PCR to demonstrate that DAX-1 was expressed in the hypothalamus and the pituitary, and to confirm its expression in adrenal cortex and gonads. The expression of DAX-1 in these tissues indicates the involvement of DAX-1 in the development of the reproductive system at multiple levels within the hypothalamic-pituitary-adrenal/gonadal axis. We also observed the expression of DAX-1 in a human adrenocortical carcinoma cell line, NCI-H295, that has features characteristic of the fetal adrenal cortex. Therefore, NCI-H295 cells will be a useful cellular model for investigating the involvement of DAX-1 in the regulation of steroidogenesis.

An unusual member of the nuclear hormone receptor superfamily responsible for X-linked adrenal hypoplasia congenita.

X-linked adrenal hypoplasia congenita is a developmental disorder of the human adrenal gland that results in profound hormonal deficiencies and is lethal if untreated. We have isolated the gene responsible for the disease, DAX-1, which is deleted or mutated in X-linked adrenal hypoplasia patients. DAX-1 encodes a new member of the nuclear hormone receptor superfamily displaying a novel DNA-binding domain. The DAX-1 product acts as a dominant negative regulator of transcription mediated by the retinoic acid receptor.

The management of breast feeding among infants with phenylketonuria.

Treatment for phenylketonuria (PKU) involves using low phenylalanine-free or phenylalanine-free formulas and supplementation with sufficient phenylalanine for normal growth and development. Eighteen infants with phenylketonuria who received breast milk as their primary phenylalanine source were compared with ten other infants with PKU who received their phenylalanine primarily from infant formulas. There were no significant differences between breast-fed and formula-fed infants for serum phenylalanine, serum tyrosine, length, weight, head circumference, haematocrit, haemoglobin, serum iron, total iron binding capacity, percentage iron saturation, ferritin, plasma zinc and total calorie intake. Breast-fed infants did show lower mean corpuscular volume at 3 months and 6 months of age. Breast-fed infants had lower phenylalanine intake at 2, 4, 5 and 6 months of age. Breast-fed infants at 1, 2, 3, 4, 5 and 6 months of age had lower protein intake. Breast feeding may be continued in the newly diagnosed phenylketonuric infant without any apparent adverse nutritional consequences.

Measurement of adenosine triphosphate and 2,3-diphosphoglycerate in stored blood with 31P nuclear magnetic resonance spectroscopy.

Conditions for blood storage are chosen to assure adequate levels of adenosine triphosphate (ATP) and 2,3-diphosphoglycerate (2,3-DPG). Because of the invasive nature of the techniques, biochemical assays are not routinely used to measure levels of these compounds in stored blood. However, 31P NMR spectroscopy measures phosphorylated intermediates in intact cells and could be used without disruption of the storage pack. We compared levels of ATP and 2,3-DPG measured by 31P spectroscopy and standard enzyme-linked biochemical assays in whole blood (WB) and packed red blood cells (PRBCs) at weekly intervals during a 35-day storage period. NMR demonstrated a marked decrease in 2,3-DPG and an increase in inorganic phosphate after the first week of storage. No significant differences in ATP concentrations were seen in WB during the storage period, but a significant decrease in ATP in PRBCs was documented. There was good agreement in levels of ATP and 2,3-DPG measured by NMR and biochemical techniques. 31P NMR spectroscopy is a noninvasive technique for measuring ATP and 2,3-DPG which has a potential use in quality assurance of stored blood.

Issues in the dietary management of phenylketonuria: breast-feeding and trace-metal nutriture.

We have reviewed our experience with supplemental breast-feeding of the infant with PKU. Our results indicate no harmful nutritional effects of breast-feeding the child with PKU, in comparison with the traditional approach using formula or cow's milk for supplementation. In addition, breast-feeding may provide a source of emotional support for the mother during this difficult period of initial diagnosis and management. It is hoped that this may improve the family's adjustment to this chronic illness. Our work with breast-feeding led us to a consideration of trace-metal nutriture in children treated with these synthetic and semi-synthetic formulas. The results of these investigations suggest that there is a biochemically significant decrease in the bioavailability of zinc when these artificial formulas are used. While no clinical trace-metal deficiency has been described in treated PKU patients, we suggest that these nutritional deficits may relate to subtle abnormalities exhibited by these patients.

Zinc status and growth of children undergoing treatment for phenylketonuria.

Children with phenylketonuria (PKU) are treated with semi-synthetic diets restricted in phenylalanine. Low-phenylalanine or phenylalanine-free formulae provide the majority of protein and energy in the diet while phenylalanine requirements are met by low-protein natural foods. Because of the restriction of natural protein sources in this diet, the study assessed the zinc nutrition of 22 treated children with PKU (aged from 1 month to 8 1/2 years) and correlated linear growth with zinc status. The mean (+/- SE) plasma zinc concentration of the PKU population was 66.6 +/- 3.3 micrograms/dl. The mean (+/- SE) hair zinc concentration was 70.2 +/- 11.5 micrograms/g. The mean plasma and hair zinc concentrations of the PKU population were significantly different (p less than 0.05) when compared with mean (+/- SE) normal values of 84.2 +/- 2.9 micrograms/dl and 130.7 +/- 8.3 micrograms/g, respectively. The mean (+/- SD) dietary zinc intake of 10 PKU patients was 8.56 +/- 2.68 mg/day. No significant differences (p less than 0.123) were found when the mean zinc intake was compared with National Academy of Sciences Recommended Dietary Allowance for age of 10 mg/day. No significant correlations were found when plasma and hair zinc concentrations were plotted with height percentiles. Further studies are required to assess the effects of zinc supplementation and the bioavailability of zinc from low-phenylalanine diets.