Herman J. Degenhart

The very low birth weight premature infant is capable of synthesizing arachidonic and docosahexaenoic acids from linoleic and linolenic acids.

Infants fed formulas devoid of long-chain polyunsaturated fatty acids (LCP) exhibit low plasma LCP concentrations and have poorer retinal and neurologic development in comparison with their human milk-fed counterparts. It is not known whether the low plasma LCP concentrations result from an impaired biosynthetic capacity, a high need, or a low dietary intake. With stable isotope technology and high sensitivity tracer detection using gas chromatography-isotope ratio mass spectrometry we measured the conversion of[13C]linoleic acid (C18:2n-6) and [13C]linolenic acid (C18:3n-3) into their longer chain derivatives in five 1-mo-old formula-fed preterm infants (birth weight 1.17 ± 0.12 kg and gestational age 28.4 ± 1.3 wk). Carbon-13-labeled linoleic acid and inolenic were mixed with the formula and administered continuously for 48 h. Both tracers were rapidly incorporated in plasma phospholipids, and their metabolic products including arachidonic acid (C20:4n-6) and docosahexaenoic acid (C22:6n-3) became highly enriched. We demonstrate that the preterm infant is capable of synthesizing LCP from their 18-carbon precursors, and our data do not support the hypothesis that a reduced δ6 desaturation is a main factor leading to low arachidonic acid and docosahexaenoic acid levels.

Standards for total body fat and fat-free mass in infants.

Data on body composition in conjunction with reference centiles are helpful in identifying the severity of growth and nutritional disorders in infancy and for evaluating the adequacy of treatment given during this important period of rapid growth. Total body fat (TBF) and fat-free mass (FFM) were estimated from total body electrical conductivity (TBEC) measurements in 423 healthy term Caucasian infants, aged 14-379 days. Cross sectional age, weight, and length related centile standards are presented for TBF and FFM. Centiles were calculated using Altmans method, based on polynomial regression and modelling of the residual variation. The TBF percentage steeply increased during the first half year of life, and slowly declined beyond this age. Various simple TBEC derived anthropometric prediction equations for TBF and FFM are available to be used in conjunction with these standards. Regression equations for the P50 and the residual SD, depending on age, weight, or length, are provided for constructing centile charts and calculating standard deviation scores.

Total Body Electrical Conductivity Measurements: An Evaluation of Current Instrumentation for Infants

ABSTRACT: Quantitation of the bodys fat and lean masses is an important component of nutritional assessment. Such measurements, however, are difficult to conduct routinely in infants due to the numerous limitations of traditional methods. The application of total body electrical conductivity measurements for quantitating fat-free mass (FFM) overcomes many of these limitations. The instruments required to perform these measurements in pediatric patients (HP-2) have recently become commercially available, but their measurement performance has not been evaluated. In these studies, we compared the precision, day-to-day variability, and magnetic field profile of three HP-2 instruments. We also derived a new calibration equation that relates the FFM to the total body electrical conductivity measurement in piglets, and compared it with an equation (provided currently by the manufacturer) derived on a prototype instrument. The performance of the instruments was generally similar, although a significant difference in the magnetic field of one instrument was identified. The coefficient of variation of inanimate phantom measurements varied from ±0.2 to ±0.5%, and the day-to-day variability was generally similar. Such measurement error is significant (±0.035 to ±0.078 kg FFM) for small subjects. The new calibration equation was similar to the original equation; therefore, all the data were pooled to generate a new equation that is linear at least to 10 kg. Thus, the HP-2 total body electrical conductivity instruments, which can be safely and easily used to measure FFM and fat in infants through 1 y of age, proved to be reliable and precise, and results obtained from different instruments can be confidently compared.

Measurement of fat-free mass in infants

ABSTRACT: Body composition data are important for adequate monitoring of growth and nutritional status in infants. 180 Isotope dilution techniques (ID18-o) are widely used to estimate total body water (TBW) and calculate fat-free mass (FFM). A problem of isotope dilution is an underestimation of TBW by the extrapolation to t = 0 approach and an overestimation of TBW by the plateau approach. Using total body electrical conductivity (TOBEC) as the reference technique we validated the extrapolation approach by 149 measurements (boys, n = 76; girls, n = 73) in 50 healthy infants aged 1–12 mo. TOBEC-derived FFM and fat mass were in excellent agreement with Fomons reference data. Strictly linear relationships with slopes not significantly different from one were found between FFM estimated by TOBEC (FFMTOBEC) and FFM estimated by ID18-O (FFM18-O) (r = 0.98 and residual SD = 0.29 for boys, r = 0.98 and residual SD = 0.32 for girls). FFM18-O was slightly but significantly lower than FFMTOBEC, the difference being on average 0.18 (±0.24) kg for girls and 0.08 (± 0.21) kg for boys (i.e. respectively 4 (±4.5)% (p < 0.0001) and 1.5 (±3.9)% (p = 0.004) of FFMTOBEC). We conclude that ID18-O using the extrapolation to t = 0 approach is suitable for TBW and FFM estimations in groups of infants. Due to the considerable measurement error of ID18-O (estimated at 6%), individual TBW18-O and FFM18-O estimates should be considered with some caution.

Mapping of a de novo unequal crossover causing a deletion of the steroid 21-hydroxylase (CYP21A2) gene and a non-functional hybrid tenascin-X (TNXB) gene.

In the human genome, the major histocompatibility complex class III region on chromosome 6p21.3 stands out as an area of remarkably high gene density.1,2 Within this region, a section of particular complexity centres around the C4 genes, which encode the fourth component of complement.3–7 Centromeric to C4 lies the CYP21A2 gene, which encodes steroid 21-hydroxylase, a key enzyme in the biosynthesis of cortisol and aldosterone.4,8,9 The TNXB gene, which encodes the extracellular matrix protein tenascin-X, lies centromeric to CYP21A2 and is transcribed from the opposite strand.10–12 Telomeric to C4 lies the RP1 gene, encoding a putative serine/threonine kinase.13–15 A typical chromosome 6 carries a duplication of an area of approximately 30 kb encompassing the entire C4 and CYP21 genes4,8 plus small truncated sections of RP and TNX .10–14 This tandem repeat has been named the RCCX module after its four constituent genes.7,14,16,17 In most white populations, about 70% of all haplotypes have a bimodular arrangement similar to the one shown in fig 1. The complex genetics of this region, and the activities and clinical significance of the proteins encoded here, have been the subject of several recent reviews.7,18–21 Figure 1 Overview of a typical C4 / CYP21 area within the MHC class III region showing two RCCX modules as found on most chromosomes. TNXB is the full size 68 kb gene for tenascin-X. TNXA (also known as XA ) is a truncated pseudogene of 5.7 kb that not only lacks most of the coding sequence of TNXB but also has a 120 bp deletion (indicated by the small triangle) spanning an exon-intron boundary.10,12,17 CYP21A2 (also known as CYP21B ) is the active steroid 21-hydroxylase gene; CYP21A1P (also known as …

PCR-based Detection of CYP21 Deletions

We read with interest the Technical Brief by Lee et al. (1), in which the authors describe a novel method to detect C4-CYP21 deletions in patients with steroid 21-hydroxylase deficiency. Such deletions result from an unequal crossover in the RCCX module ( RP-C4-CYP21-TNX ) on chromosome 6. In most cases, chromosome 6 carries two RCCX modules, one with a CYP21P ( CYP21A1P ) pseudogene and a truncated XA pseudogene, and one with a functional CYP21 ( CYP21A2 ) gene (encoding steroid 21-hydroxylase) and a functional TNXB gene (encoding tenascin-X). Meiotic misalignment and recombination may occur at several locations and create a chromosome with a single chimeric RCCX module. The PCR described by Lee et al. uses one primer in the 5′ flanking sequence of CYP21 and CYP21P (2), whereas the other primer is positioned in a 120-bp sequence of TNXB that is not present in the XA pseudogene (3). Although this PCR is indeed suitable for the detection of chimeric CYP21P / CYP21 genes, it would fail to detect any RCCX chimera in which the pseudogene-like region includes the 120-bp deletion of XA (4), as …

The effect of alterations in physical and chemical characteristics on TOBEC-derived body composition estimates: validation with non-human models

The measurement of total-body electrical conductivity (TOBEC) has become one of the standard methods for the estimation of body composition in infants. We investigated, using non-human models, the effect on the accuracy of TOBEC-derived body composition estimates of alterations in physical and chemical characteristics of the fat-free mass (FFM). The effect of electrolyte type, concentration and volume on TOBEC was determined using 2, 3 and 51 solutions of six different chlorides and sodium bicarbonate. Equimolar concentrations yielded TOBEC values in accordance with known ion conductivities: H+ >> Ca2+ > Mg2+ > K+ > Na+ > Li+ and Cl- > HCO3-. The behaviour of these solutions was described very accurately over a wide range of concentrations (1-200 mM) by a simple exponential law. Dissolved egg-white protein, glycine and L-glutamine elicited no TOBEC signal. In vitro, using polyethylene bottles filled with physiologic saline, in the interval of 2-45 degrees C a linear relation was observed between temperature and TOBEC. Below the freezing point no TOBEC signal was elicited. The effect of tissue autolysis and body temperature on TOBEC was examined by repeated measurements of TOBEC and temperature in seven fresh infant minipig cadavers. Five minipigs were allowed to cool. Shortly after death TOBEC decreased by 2.5% per degrees C. Two animals were kept at constant temperature. The TOBEC signal showed a gradual increase of 9% after 7 h due to autolysis. We conclude that in vivo TOBEC measurements are affected by ion concentration (e.g. non-isotonic hydration changes), geometry (e.g. deviations in body shape), temperature (e.g. fever, skin cooling) and tissue autolysis (measurements after death). Proteins, molecules with strong dipole moments and ions trapped in crystalline structures do not significantly affect the TOBEC reading.

A rare TaqI polymorphism in a human complement C4 gene is caused by an additional restriction site in the first intron

We studied the configuration of the complement C4/CYP21 (steroid 21-hydroxylase) region of the human major histocompatibility complex in patients suffering from congenital adrenal hyperplasia (CAH) and in the general population in The Netherlands, using C4 and CYP21 probes and the restriction enzymes TaqI and Bg/II. We found a rare TaqI 3.9-kb restriction fragment in the mother of a CAH patient, and present evidence that this polymorphism is caused by an additional restriction site in the first intron of a complement C4 gene.

Body composition in the first year of life

The two-compartment model is still the most widely used model for the study of body composition in infants. The model assumes that the body consists of fat and fat-free mass (FFM). Body composition data are important for adequate follow-up and monitoring of nutritional status and quality of growth, the nutritional adequacy of specific preterm and term infant diets, severity of malnutrition, and the progress of recovery from malnutrition, particularly in fast-growing preterm and term young infants.

56 A RECOMBINATION EVENT CAUSING A DE NOVO DELETION OF THE STEROID 21-HYDROXYLASE GENE CYP21

Steroid 21-hydroxylase deficiency is the most prominent cause of congenital adrenal hyperplasia (CAH). Patients suffer from virilization, and in severe cases, from salt loss caused by lack of aldosterone. The CYP21 gene, encoding steroid 21-hydroxylase, and the highly homologous pseudogene CYP21P are located within the human MHC on chromosome 6p21.3, the usual arrangement being centromere - HLA-DP, -DQ, -DR - CYP21-complement C4B-CYP21P-complement C4A - tumour necrosis factor (TNF) genes - HLA-B. -C. -A. We investigated a CAH family (father, mother, patient, three healthy sibs) using 21-hydroxylase and complement C4 probes and allele specific oligonucleotides to detect specific mutations. Oligonucleotide hybridization showed that the patient inherited the Ile172->Asn mutation. characteristic of “simple virilizing” CAH. from the mother. On the other chromosome, the CYP21 gene was deleted: however, this defect was not found in DNA from the father. Paternity was confirmed using VNTR probes. Establishment of HLA-B. TNF and HLA-DQα markers showed that the patient had the HLA-B and TNF genes from one paternal chromosome and the HLA-DQα gene from the other. Apparently, a paternal meiotic recombination event has eliminated the CYP21 gene (as well as the adjacent C4B gene), contributing to steroid 21-hydroxylase deficiency in the patient.