Post mortem analysis of hepatic volume and lipid content by magnetic resonance imaging and spectroscopy in fixed murine neonates.

Maternal obesity and hyperglycemia are linked to an elevated risk for obesity, diabetes, and steatotic liver disease in the adult offspring. To establish and validate a noninvasive workflow for perinatal metabolic phenotyping, fixed neonates of common mouse strains were analyzed postmortem via magnetic resonance imaging (MRI)/magnetic resonance spectroscopy (MRS) to assess liver volume and hepatic lipid (HL) content. The key advantage of nondestructive MRI/MRS analysis is the possibility of further tissue analyses, such as immunohistochemistry, RNA extraction, and even proteomics, maximizing the data that can be gained per individual and therefore facilitating comprehensive correlation analyses. This study employed an MRI and 1H-MRS workflow to measure liver volume and HL content in 65 paraformaldehyde-fixed murine neonates at 11.7 T. Liver volume was obtained using semiautomatic segmentation of MRI acquired by a RARE sequence with 0.5-mm slice thickness. HL content was measured by a STEAM sequence, applied with and without water suppression. T1 and T2 relaxation times of lipids and water were measured for respective correction of signal intensity. The HL content, given as CH2/(CH2 + H2O), was calculated, and the intrasession repeatability of the method was tested. The established workflow yielded robust results with a variation of ~3% in repeated measurements for HL content determination. HL content measurements were further validated by correlation analysis with biochemically assessed triglyceride contents (R2 = 0.795) that were measured in littermates. In addition, image quality also allowed quantification of subcutaneous adipose tissue and stomach diameter. The highest HL content was measured in C57Bl/6N (4.2%) and the largest liver volume and stomach diameter in CBA (53.1 mm3 and 6.73 mm) and NMRI (51.4 mm3 and 5.96 mm) neonates, which also had the most subcutaneous adipose tissue. The observed effects were independent of sex and litter size. In conclusion, we have successfully tested and validated a robust MRI/MRS workflow that allows assessment of morphology and HL content and further enables paraformaldehyde-fixed tissue-compatible subsequent analyses in murine neonates.

Distinct maternal metabolites are associated with obesity and glucose-insulin axis in the first trimester of pregnancy.

BACKGROUND/OBJECTIVES: Obesity in pregnancy associates with changes in the glucose-insulin axis. We hypothesized that these changes affect the maternal metabolome already in the first trimester of human pregnancy and, thus, aimed to identify these metabolites. PATIENTS/METHODS: We performed untargeted metabolomics (HPLC-MS/MS) on maternal serum (n = 181, gestational weeks 4+0-11+6). For further analysis, we included only non-smoking women as assessed by serum cotinine levels (ELISA) (n = 111). In addition to body mass index (BMI) and leptin as measures of obesity and adiposity, we metabolically phenotyped women by their fasting glucose, C-peptide and insulin sensitivity (ISHOMA index). To identify metabolites (outcome) associated with BMI, leptin, glucose, C-peptide and/or ISHOMA (exposures), we used a combination of univariable and multivariable regression analyses with multiple confounders and machine learning methods (Partial Least Squares Discriminant Analysis, Random Forest and Support Vector Machine). Additional statistical tests confirmed robustness of results. Furthermore, we performed network analyses (MoDentify package) to identify sets of correlating metabolites that are coordinately regulated by the exposures. RESULTS: We detected 2449 serum features of which 277 were annotated. After stringent analysis, 15 metabolites associated with at least one exposure (BMI, leptin, glucose, C-peptide, ISHOMA). Among these, palmitoleoyl ethanolamine (POEA), an endocannabinoid-like lipid endogenously synthesized from palmitoleic acid, and N-acetyl-L-alanine were consistently associated with C-peptide in all the analyses (95% CI: 0.10-0.34; effect size: 21%; p < 0.001; 95% CI: 0.04-0.10; effect size: 7%; p < 0.001). In network analysis, most features correlating with palmitoleoyl ethanolamide and N-acetyl-L-alanine and associated with C-peptide, were amino acids or dipeptides (n = 9, 35%), followed by lipids (n = 7, 27%). CONCLUSIONS: We conclude that the metabolome of pregnant women with overweight/obesity is already altered early in pregnancy because of associated changes of C-peptide. Changes of palmitoleoyl ethanolamide concentration in pregnant women with obesity-associated hyperinsulinemia may reflect dysfunctional endocannabinoid-like signalling.

Isovaleric aciduria identified by newborn screening: Strategies to predict disease severity and stratify treatment.

Newborn screening (NBS) allows early identification of individuals with rare disease, such as isovaleric aciduria (IVA). Reliable early prediction of disease severity of positively screened individuals with IVA is needed to guide therapeutic decision, prevent life-threatening neonatal disease manifestation in classic IVA and over-medicalization in attenuated IVA that may remain asymptomatic. We analyzed 84 individuals (median age at last study visit 8.5 years) with confirmed IVA identified by NBS between 1998 and 2018 who participated in the national, observational, multicenter study. Screening results, additional metabolic parameters, genotypes, and clinical phenotypic data were included. Individuals with metabolic decompensation showed a higher median isovalerylcarnitine (C5) concentration in the first NBS sample (10.6 vs. 2.7 µmol/L; p < 0.0001) and initial urinary isovalerylglycine concentration (1750 vs. 180 mmol/mol creatinine; p = 0.0003) than those who remained asymptomatic. C5 was in trend inversely correlated with full IQ (R = -0.255; slope = -0.869; p = 0.0870) and was lower for the "attenuated" variants compared to classic genotypes [median (IQR; range): 2.6 µmol/L (2.1-4.0; 0.7-6.4) versus 10.3 µmol/L (7.4-13.1; 4.3-21.7); N = 73]. In-silico prediction scores (M-CAP, MetaSVM, and MetaLR) correlated highly with isovalerylglycine and ratios of C5 to free carnitine and acetylcarnitine, but not sufficiently with clinical endpoints. The results of the first NBS sample and biochemical confirmatory testing are reliable early predictors of the clinical course of IVA, facilitating case definition (attenuated versus classic IVA). Prediction of attenuated IVA is supported by the genotype. On this basis, a reasonable algorithm has been established for neonates with a positive NBS result for IVA, with the aim of providing the necessary treatment immediately, but whenever possible, adjusting the treatment to the individual severity of the disease.

Predicting the earliest deviation in weight gain in the course towards manifest overweight in offspring exposed to obesity in pregnancy: a longitudinal cohort study.

BACKGROUND: Obesity in pregnancy and related early-life factors place the offspring at the highest risk of being overweight. Despite convincing evidence on these associations, there is an unmet public health need to identify "high-risk" offspring by predicting very early deviations in weight gain patterns as a subclinical stage towards overweight. However, data and methods for individual risk prediction are lacking. We aimed to identify those infants exposed to obesity in pregnancy at ages 3 months, 1 year, and 2 years who likely will follow a higher-than-normal body mass index (BMI) growth trajectory towards manifest overweight by developing an early-risk quantification system. METHODS: This study uses data from the prospective mother-child cohort study Programming of Enhanced Adiposity Risk in CHildhood-Early Screening (PEACHES) comprising 1671 mothers with pre-conception obesity and without (controls) and their offspring. Exposures were pre- and postnatal risks documented in patient-held maternal and child health records. The main outcome was a "higher-than-normal BMI growth pattern" preceding overweight, defined as BMI z-score >1 SD (i.e., World Health Organization [WHO] cut-off "at risk of overweight") at least twice during consecutive offspring growth periods between age 6 months and 5 years. The independent cohort PErinatal Prevention of Obesity (PEPO) comprising 11,730 mother-child pairs recruited close to school entry (around age 6 years) was available for data validation. Cluster analysis and sequential prediction modelling were performed. RESULTS: Data of 1557 PEACHES mother-child pairs and the validation cohort were analyzed comprising more than 50,000 offspring BMI measurements. More than 1-in-5 offspring exposed to obesity in pregnancy belonged to an upper BMI z-score cluster as a distinct pattern of BMI development (above the cut-off of 1 SD) from the first months of life onwards resulting in preschool overweight/obesity (age 5 years: odds ratio [OR] 16.13; 95% confidence interval [CI] 9.98-26.05). Contributing early-life factors including excessive weight gain (OR 2.08; 95% CI 1.25-3.45) and smoking (OR 1.94; 95% CI 1.27-2.95) in pregnancy were instrumental in predicting a "higher-than-normal BMI growth pattern" at age 3 months and re-evaluating the risk at ages 1 year and 2 years (area under the receiver operating characteristic [AUROC] 0.69-0.79, sensitivity 70.7-76.0%, specificity 64.7-78.1%). External validation of prediction models demonstrated adequate predictive performances. CONCLUSIONS: We devised a novel sequential strategy of individual prediction and re-evaluation of a higher-than-normal weight gain in "high-risk" infants well before developing overweight to guide decision-making. The strategy holds promise to elaborate interventions in an early preventive manner for integration in systems of well-child care.

[Individualized Somatic Classification of Newborns Using Birth Weight Percentiles Based On Maternal Body Height and Weight (Results of a Validation Study)]. / Individuelle somatische Klassifikation von Neugeborenen auf der Grundlage von Körperhöhe und Körpergewicht der Mutter (Ergebnisse einer Validierungsstudie).

INTRODUCTION: Birth weight is influenced by maternal anthropometry. The SGA-rate of newborns of short and light mothers (<158 cm,<53 kg) and the LGA-rate of tall and heavy mothers (>177 cm,>79 kg) are overestimated. The LGA-rate of newborns of shorter mothers and the SGA-rate of taller mothers are underestimated. Individualized birth weight percentiles (IBWP) based on 18 maternal groups (6 groups of height combined with 3 groups of weight), sex and weeks of gestation have been published. The aim of this study is to validate IBWP by evaluating SGA-, AGA-, and LGA-rates using perinatal data. METHODS: The validation study compares IBWP (1995 to 2000, n=2.2 million singletons) with percentile values from two German cohorts (i: 1995 to 2000; n=2.3 million and ii: 2007 to 2011, n=3.2 million singletons) using newborns from the Lower Saxony Perinatal Survey (n=0.56 million singleton newborns, 2001 to 2009). SGA-, AGA-, and LGA-rates were calculated using R statistical analysis. RESULTS: Common percentile charts based on the total population 1995-2000 and 2007-2011 yielded SGA-rates among shorter mothers of 21.1 to 21.6% and LGA-rates of 2.0 to 3.1%. In taller mothers, SGA-rates were 3.3 to 3.5% and LGA-rates were 26.6 to 27.1%. IBWP achieved SGA-rates of 9.0% and LGA-rates of 11.4 to 11.6% in shorter mothers and SGA- and LGA-rates of 10% in taller mothers. DISCUSSION: IBWP consider the maternal size for estimation of the fetal growth potential and achieve expected SGA- and LGA-rates of 10%. Consideration of individual growth potential avoids underestimation and overestimation of SGA- and LGA-rates. It aided analyses of birth weight with IBWP simplify the assessment of the nutritional status.

Newborn screening and disease variants predict neurological outcome in isovaleric aciduria.

Isovaleric aciduria (IVA), a metabolic disease with severe (classic IVA) or attenuated phenotype (mild IVA), is included in newborn screening (NBS) programs worldwide. The long-term clinical benefit of screened individuals, however, is still rarely investigated. A national, prospective, observational, multi-center study of individuals with confirmed IVA identified by NBS between 1998 and 2018 was conducted. Long-term clinical outcomes of 94 individuals with IVA were evaluated, representing 73.4% (for classic IVA: 92.3%) of the German NBS cohort. In classic IVA (N = 24), NBS prevented untimely death except in one individual with lethal neonatal sepsis (3.8%) but did not completely prevent single (N = 10) or recurrent (N = 7) metabolic decompensations, 13 of them occurring already neonatally. IQ (mean ± SD, 90.7 ± 10.1) was mostly normal but below the reference population (P = .0022) and was even lower in individuals with severe neonatal decompensations (IQ 78.8 ± 7.1) compared to those without crises (IQ 94.7 ± 7.5; P = .01). Similar results were obtained for school placement. In contrast, individuals with mild IVA had excellent neurocognitive outcomes (IQ 105.5 ± 15.8; normal school placement) and a benign disease course (no metabolic decompensation, normal hospitalization rate), which did not appear to be impacted by metabolic maintenance therapy. In conclusion, NBS reduces mortality in classic IVA, but does not reliably protect against severe neonatal metabolic decompensations, crucial for favorable neurocognitive outcome. In contrast, individuals with mild IVA had excellent clinical outcomes regardless of metabolic maintenance therapy, questioning their benefit from NBS. Harmonized stratified therapeutic concepts are urgently needed.

Impact of interventional and non-interventional variables on anthropometric long-term development in glutaric aciduria type 1: A national prospective multi-centre study.

Glutaric aciduria type 1 (GA1) is a rare neurometabolic disorder, caused by inherited deficiency of glutaryl-CoA dehydrogenase, mostly affecting the brain. Early identification by newborn screening (NBS) significantly improves neurologic outcome. It has remained unclear whether recommended therapy, particular low lysine diet, is safe or negatively affects anthropometric long-term outcome. This national prospective, observational, multi-centre study included 79 patients identified by NBS and investigated effects of interventional and non-interventional parameters on body weight, body length, body mass index (BMI) and head circumference as well as neurological parameters. Adherence to recommended maintenance and emergency treatment (ET) had a positive impact on neurologic outcome and allowed normal anthropometric development until adulthood. In contrast, non-adherence to ET, resulting in increased risk of dystonia, had a negative impact on body weight (mean SDS -1.07; P = .023) and body length (mean SDS -1.34; P = -.016). Consistently, longitudinal analysis showed a negative influence of severe dystonia on weight and length development over time (P < .001). Macrocephaly was more often found in female (mean SDS 0.56) than in male patients (mean SDS -0.20; P = .049), and also in individuals with high excreter phenotype (mean SDS 0.44) compared to low excreter patients (mean SDS -0.68; P = .016). In GA1, recommended long-term treatment is effective and allows for normal anthropometric long-term development up to adolescence, with gender- and excreter type-specific variations. Delayed ET and severe movement disorder result in poor anthropometric outcome.

Sex-specific programming effects of parental obesity in pre-implantation embryonic development.

BACKGROUND: Obesity is a global rising problem with epidemiological dimension. Obese parents can have programming effects on their offspring leading to obesity and associated diseases in later life. This constitutes a vicious circle. Epidemiological data and studies in rodents demonstrated differential programming effects in male and female offspring, but the timing of their developmental origin is not known. METHODS: This study investigated if sex-specific programming effects of parental obesity can already be detected in the pre-implantation period. Diet-induced obese male or female mice were mated with normal-weight partners and blastocysts were recovered. RESULTS: Gene expression profiling revealed sex-specific responses of the blastocyst transcriptome to maternal and paternal obesity. The changes in the transcriptome of male blastocysts were more pronounced than those of female blastocysts, with a stronger impact of paternal than of maternal obesity. The sperm of obese mice revealed an increased abundance of several miRNAs compared with lean mice. CONCLUSIONS: Our study indicates that sex-specific programming effects of parental obesity already start in the pre-implantation period and reveals specific alterations of the sperm miRNA profile as mechanistic link to programming effects of paternal obesity.

Cardiometabolic risk factor clustering in patients with deficient branched-chain amino acid catabolism: A case-control study.

Classical organic acidemias (OAs) result from defective mitochondrial catabolism of branched-chain amino acids (BCAAs). Abnormal mitochondrial function relates to oxidative stress, ectopic lipids and insulin resistance (IR). We investigated whether genetically impaired function of mitochondrial BCAA catabolism associates with cardiometabolic risk factors, altered liver and muscle energy metabolism, and IR. In this case-control study, 31 children and young adults with propionic acidemia (PA), methylmalonic acidemia (MMA) or isovaleric acidemia (IVA) were compared with 30 healthy young humans using comprehensive metabolic phenotyping including in vivo 31 P/1 H magnetic resonance spectroscopy of liver and skeletal muscle. Among all OAs, patients with PA exhibited abdominal adiposity, IR, fasting hyperglycaemia and hypertriglyceridemia as well as increased liver fat accumulation, despite dietary energy intake within recommendations for age and sex. In contrast, patients with MMA more frequently featured higher energy intake than recommended and had a different phenotype including hepatomegaly and mildly lower skeletal muscle ATP content. In skeletal muscle of patients with PA, slightly lower inorganic phosphate levels were found. However, hepatic ATP and inorganic phosphate concentrations were not different between all OA patients and controls. In patients with IVA, no abnormalities were detected. Impaired BCAA catabolism in PA, but not in MMA or IVA, was associated with a previously unrecognised, metabolic syndrome-like phenotype with abdominal adiposity potentially resulting from ectopic lipid storage. These findings suggest the need for early cardiometabolic risk factor screening in PA.

Newborn screening: A disease-changing intervention for glutaric aciduria type 1.

OBJECTIVE: Untreated individuals with glutaric aciduria type 1 (GA1) commonly present with a complex, predominantly dystonic movement disorder (MD) following acute or insidious onset striatal damage. Implementation of GA1 into newborn screening (NBS) programs has improved the short-term outcome. It remains unclear, however, whether NBS changes the long-term outcome and which variables are predictive. METHODS: This prospective, observational, multicenter study includes 87 patients identified by NBS, 4 patients missed by NBS, and 3 women with GA1 identified by positive NBS results of their unaffected children. RESULTS: The study population comprises 98.3% of individuals with GA1 identified by NBS in Germany during 1999-2016. Overall, cumulative sensitivity of NBS is 95.6%, but it is lower (84%) for patients with low excreter phenotype. The neurologic outcome of patients missed by NBS is as poor as in the pre-NBS era, and the clinical phenotype of diagnosed patients depends on the quality of therapeutic interventions rather than noninterventional variables. Presymptomatic start of treatment according to current guideline recommendations clearly improves the neurologic outcome (MD: 7% of patients), whereas delayed emergency treatment results in acute onset MD (100%), and deviations from maintenance treatment increase the risk of insidious onset MD (50%). Independent of the neurologic phenotype, kidney function tends to decline with age, a nonneurologic manifestation not predicted by any variable included in this study. INTERPRETATION: NBS is a beneficial, disease-changing intervention for GA1. However, improved neurologic outcome critically depends on adherence to recommended therapy, whereas kidney dysfunction does not appear to be impacted by recommended therapy. Ann Neurol 2018;83:970-979.

Late-pregnancy dysglycemia in obese pregnancies after negative testing for gestational diabetes and risk of future childhood overweight: An interim analysis from a longitudinal mother-child cohort study.

BACKGROUND: Maternal pre-conception obesity is a strong risk factor for childhood overweight. However, prenatal mechanisms and their effects in susceptible gestational periods that contribute to this risk are not well understood. We aimed to assess the impact of late-pregnancy dysglycemia in obese pregnancies with negative testing for gestational diabetes mellitus (GDM) on long-term mother-child outcomes. METHODS AND FINDINGS: The prospective cohort study Programming of Enhanced Adiposity Risk in Childhood-Early Screening (PEACHES) (n = 1,671) enrolled obese and normal weight mothers from August 2010 to December 2015 with trimester-specific data on glucose metabolism including GDM status at the end of the second trimester and maternal glycated hemoglobin (HbA1c) at delivery as a marker for late-pregnancy dysglycemia (HbA1c ≥ 5.7% [39 mmol/mol]). We assessed offspring short- and long-term outcomes up to 4 years, and maternal glucose metabolism 3.5 years postpartum. Multivariable linear and log-binomial regression with effects presented as mean increments (Δ) or relative risks (RRs) with 95% confidence intervals (CIs) were used to examine the association between late-pregnancy dysglycemia and outcomes. Linear mixed-effects models were used to study the longitudinal development of offspring body mass index (BMI) z-scores. The contribution of late-pregnancy dysglycemia to the association between maternal pre-conception obesity and offspring BMI was estimated using mediation analysis. In all, 898 mother-child pairs were included in this unplanned interim analysis. Among obese mothers with negative testing for GDM (n = 448), those with late-pregnancy dysglycemia (n = 135, 30.1%) had higher proportions of excessive total gestational weight gain (GWG), excessive third-trimester GWG, and offspring with large-for-gestational-age birth weight than those without. Besides higher birth weight (Δ 192 g, 95% CI 100-284) and cord-blood C-peptide concentration (Δ 0.10 ng/ml, 95% CI 0.02-0.17), offspring of these women had greater weight gain during early childhood (Δ BMI z-score per year 0.18, 95% CI 0.06-0.30, n = 262) and higher BMI z-score at 4 years (Δ 0.58, 95% CI 0.18-0.99, n = 43) than offspring of the obese, GDM-negative mothers with normal HbA1c values at delivery. Late-pregnancy dysglycemia in GDM-negative mothers accounted for about one-quarter of the association of maternal obesity with offspring BMI at age 4 years (n = 151). In contrast, childhood BMI z-scores were not affected by a diagnosis of GDM in obese pregnancies (GDM-positive: 0.58, 95% CI 0.36-0.79, versus GDM-negative: 0.62, 95% CI 0.44-0.79). One mechanism triggering late-pregnancy dysglycemia in obese, GDM-negative mothers was related to excessive third-trimester weight gain (RR 1.72, 95% CI 1.12-2.65). Furthermore, in the maternal population, we found a 4-fold (RR 4.01, 95% CI 1.97-8.17) increased risk of future prediabetes or diabetes if obese, GDM-negative women had a high versus normal HbA1c at delivery (absolute risk: 43.2% versus 10.5%). There is a potential for misclassification bias as the predominantly used GDM test procedure changed over the enrollment period. Further studies are required to validate the findings and elucidate the possible third-trimester factors contributing to future mother-child health status. CONCLUSIONS: Findings from this interim analysis suggest that offspring of obese mothers treated because of a diagnosis of GDM appeared to have a better BMI outcome in childhood than those of obese mothers who-following negative GDM testing-remained untreated in the last trimester and developed dysglycemia. Late-pregnancy dysglycemia related to uncontrolled weight gain may contribute to the development of child overweight and maternal diabetes. Our data suggest that negative GDM testing in obese pregnancies is not an "all-clear signal" and should not lead to reduced attention and risk awareness of physicians and obese women. Effective strategies are needed to maintain third-trimester glycemic and weight gain control among otherwise healthy obese pregnant women.

Modification of the fatty acid composition of an obesogenic diet improves the maternal and placental metabolic environment in obese pregnant mice.

Peri-conceptional exposure to maternal obesogenic nutrition is associated with in utero programming of later-life overweight and metabolic disease in the offspring. We aimed to investigate whether dietary intervention with a modified fatty acid quality in an obesogenic high-calorie (HC) diet during the preconception and gestational phases can improve unfavourable effects of an adipogenic maternal environment. In NMRI mice, peri-conceptional and gestational obesity was induced by feeding a HC diet (controls), and they were compared with dams on a fat-modified (Fat-mod) HC diet of the same energy content but enriched with medium-chain fatty acids (MCFAs) and adjusted to a decreased ratio of n-6 to n-3 long-chain polyunsaturated fatty acids (LC-PUFAs). Effects on maternal and placental outcomes at delivery (day 17.5 post coitum) were investigated. Despite comparable energy assimilation between the two groups of dams, the altered fatty acid composition of the Fat-mod HC diet induced lower maternal body weight, weights of fat depots, adipocyte size, and hepatic fat accumulation compared to the unmodified HC diet group. Further, there was a trend towards lower fasting glucose, insulin and leptin concentrations in dams fed the Fat-mod HC diet. Phenotypic changes were accompanied by inhibition of transcript and protein expression of genes involved in hepatic de novo lipogenesis comprising PPARG2 and its target genes Fasn, Acaca, and Fabp4, whereas regulation of other lipogenic factors (Srebf1, Nr1h3, Abca1) appeared to be more complex. The modified diet led to a sex-specific placental response by upregulating PPARG-dependent fatty acid transport gene expression in female versus male placentae. Qualitative modification of the fatty acid spectrum of a high-energy maternal diet, using a combination of both MCFAs and n-3 LC-PUFAs, seems to be a promising interventional approach to ameliorate the adipogenic milieu of mice before and during gestation.

Disorders of fatty acid oxidation and autosomal recessive polycystic kidney disease-different clinical entities and comparable perinatal renal abnormalities.

BACKGROUND: Differential diagnosis of prenatally detected hyperechogenic and enlarged kidneys can be challenging as there is a broad phenotypic overlap between several rare genetic and non-genetic disorders. Metabolic diseases are among the rarest underlying disorders, but they demand particular attention as their prognosis and postnatal management differ from those of other diseases. METHODS: We report two cases of cystic, hyperechogenic and enlarged kidneys detected on prenatal ultrasound images, resulting in the suspected diagnosis of autosomal recessive polycystic kidney disease (ARPKD). Postnatal clinical course and work-up, however, revealed early, neonatal forms of disorders of fatty acid oxidation (DFAO) in both cases, namely, glutaric acidemia type II, based on identification of the novel, homozygous splice-site mutation c.1117-2A > G in the ETFDH gene, in one case and carnitine palmitoyltransferase II deficiency in the other case. RESULTS: Review of pre- and postnatal sonographic findings resulted in the identification of some important differences that might help to differentiate DFAO from ARPKD. In DFAO, kidneys are enlarged to a milder degree than in ARPKD, and the cysts are located ubiquitously, including also in the cortex and the subcapsular area. Interestingly, recent studies have pointed to a switch in metabolic homeostasis, referred to as the Warburg effect (aerobic glycolysis), as one of the underlying mechanisms of cell proliferation and cyst formation in cystic kidney disease. DFAO are characterized by the inhibition of oxidative phosphorylation, resulting in aerobic glycolysis, and thus they do resemble the Warburg effect. We therefore speculate that this inhibition might be one of the pathomechanisms of renal hyperproliferation and cyst formation in DFAO analogous to the reported findings in ARPKD. CONCLUSIONS: Neonatal forms of DFAO can be differentially diagnosed in neonates with cystic or hyperechogenic kidneys and necessitate immediate biochemical work-up to provide early metabolic management.

Chitinase activation in patients with fungus-associated cystic fibrosis lung disease.

BACKGROUND: Chitinases have recently gained attention in the field of pulmonary diseases, particularly in asthma and chronic obstructive pulmonary disease, but their potential role in patients with cystic fibrosis (CF)-associated lung disease remains unclear. OBJECTIVE: The aim of this study was to assess chitinase activity systemically and in the airways of patients with CF and asthma compared with healthy subjects. Additionally, we assessed factors that regulate chitinase activity within the lungs of patients with CF. METHODS: Chitinase activities were quantified in serum and bronchoalveolar lavage fluid from patients with CF, asthmatic patients, and healthy control subjects. Mechanistically, the role of CF airway proteases and genetic chitinase deficiency was assessed. RESULTS: Chitinase activity was systemically increased in patients with CF compared with that in healthy control subjects and asthmatic patients. Further stratification showed that chitinase activity was enhanced in patients with CF colonized with Candida albicans compared with that in noncolonized patients. CF proteases degraded chitinases in the airway microenvironment of patients with CF. Genetic chitinase deficiency was associated with C albicans colonization in patients with CF. CONCLUSION: Patients with CF have enhanced chitinase activation associated with C albicans colonization. Therefore chitinases might represent a novel biomarker and therapeutic target for CF-associated fungal disease.

The peroxisome proliferator-activated receptors under epigenetic control in placental metabolism and fetal development.

The placental metabolism can adapt to the environment throughout pregnancy to both the demands of the fetus and the signals from the mother. Such adaption processes include epigenetic mechanisms, which alter gene expression and may influence the offspring's health. These mechanisms are linked to the diversity of prenatal environmental exposures, including maternal under- or overnutrition or gestational diabetes. The peroxisome proliferator-activated receptors (PPARs) are nuclear receptors that contribute to the developmental plasticity of the placenta by regulating lipid and glucose metabolism pathways, including lipogenesis, steroidogenesis, glucose transporters, and placental signaling pathways, thus representing a link between energy metabolism and reproduction. Among the PPAR isoforms, PPARγ appears to be the main modulator of mammalian placentation. Certain fatty acids and lipid-derived moieties are the natural activating PPAR ligands. By controlling the amounts of maternal nutrients that go across to the fetus, the PPARs play an important regulatory role in placenta metabolism, thereby adapting to the maternal nutritional status. As demonstrated in animal studies, maternal nutrition during gestation can exert long-term influences on the PPAR methylation pattern in offspring organs. This review underlines the current state of knowledge on the relationship between environmental factors and the epigenetic regulation of the PPARs in placenta metabolism and offspring development.

Obese Nondiabetic Pregnancies and High Maternal Glycated Hemoglobin at Delivery as an Indicator of Offspring and Maternal Postpartum Risks: The Prospective PEACHES Mother-Child Cohort.

BACKGROUND: We investigated whether obese pregnant women negative for gestational diabetes (GDM) still experience dysglycemia, as indicated by high glycated hemoglobin (Hb A1c) at delivery, and whether this impacts offspring and long-term maternal outcomes. METHODS: Data of 462 mother-child pairs of our prospective Programming of Enhanced Adiposity Risk in Childhood - Early Screening (PEACHES) cohort study were analyzed. Of 885 obese and normal-weight pregnancies prospectively enrolled after GDM testing according to the International Association of Diabetes and Pregnancy Study Groups criteria, 462 GDM-negative mothers and their offspring were investigated. We assessed associations of maternal Hb A1c at delivery with large-for-gestational-age (LGA) birth weights, cord-blood C-peptide, and biomarkers of glucose metabolism and inflammation in obese mothers followed for 2.9 years (median) postpartum (n = 42). RESULTS: Cumulative distribution analysis in GDM-negative normal-weight women (n = 155) revealed that 12% had Hb A1c ≥5.7% at delivery (high Hb A1c). Among obese GDM-negative women (n = 307), 31.9% (95% CI, 26.7%-37.4%) equaled or exceeded this cutoff. In obese GDM-negative women with Hb A1c ≥5.7% (n = 98) vs <5.7% (n = 209) at delivery, newborns were more likely to be born LGA [adjusted odds ratio 3.56 (95% CI, 1.64-8.02)], and mean cordblood serum C-peptide was increased by 0.09 ng/mL (95% CI, 0.01-0.17 ng/mL). In the mothers at follow-up, mean postpartum Hb A1c, fasting glucose, high-sensitivity C-reactive protein, and fibrinogen concentrations were higher by 0.3% (95% CI, 0.1%-0.5%), 6.0 mg/dL (95% CI, 2.4-9.5 mg/dL), 6.8 mg/L (95% CI, 1.4-12.3 mg/L), and 74.9 mg/dL (95% CI, 13.6-136.2 mg/dL), respectively. CONCLUSIONS: Increased Hb A1c in obese GDM-negative women at delivery indicates gestational dysglycemia, potentially conferring offspring and long-term maternal health risks. These findings should raise awareness as to careful monitoring of obese pregnancies. Measurement of Hb A1c at delivery could help select women who may need closer postpartum health checks.

Digital image analysis approach for lipid droplet size quantitation of Oil Red O-stained cultured cells.

A simple approach was developed for the quantification of lipid droplet size and frequency distribution in images acquired by standard light microscopy. Oil Red O-stained cell images were thresholded for the lipid droplet signal using the freely available imaging software ImageJ. Watershed algorithms allowed analyzing the area of each individual lipid droplet. The method was validated by the decrease in lipid droplet size of 3T3-L1 adipocytes on lowered glucose availability associated with reduced glycerol-3-phosphate dehydrogenase activity and reduced transcription of lipid droplet size markers. This approach can be easily applied using standard laboratory equipment without requiring expensive and complex instrumentation.

Effects of trimester-specific and total gestational weight gain on children's anthropometrics.

BACKGROUND: Gestational weight gain (GWG) has been shown to be a risk factor for overweight in offspring.Aim of this study was to quantify the contributions of trimester-specific and total GWG on offspring's BMI and waist circumference (WC). This is of interest for the design of interventions targeted at women showing a high GWG in early pregnancy. METHODS: In a retrospective cohort study data on GWG (total and by trimester, exposure), a number of potential confounders, and children's BMI z-scores and WC (outcomes) were analyzed using structural equation models to disentangle the trimester-specific direct effects of GWG and indirect effects mediated via total GWG. RESULTS: 7313 mother child pairs with a children's mean age of 5.81 years were analyzed. Total effects (indirect + direct) of GWG (kg/week) on children's BMI z-score and WC (cm) were observed in all trimesters, most prominently in the second. The longitudinal effect of GWG is a composite of trimester-specific direct effects (on BMI: 0.105, 0.255, 0.002, on WC: 0.538, 1.64, 0.308) and total GWG (on BMI 0.608, on WC: 1.03) at the end of pregnancy. CONCLUSIONS: Both trimester-specific priming and total GWG explained offspring's anthropometrics. The results indicate, that reversal from additional weight gain attained early in pregnancy resulting in normal total GWG at the end of pregnancy might still contribute to a substantial reduction of offspring's BMI and WC.

Low lysine diet in glutaric aciduria type I--effect on anthropometric and biochemical follow-up parameters.

BACKGROUND: Metabolic treatment in glutaric aciduria type I (GA-I) including a low lysine diet with lysine-free, tryptophan-reduced amino acid supplements (AAS), carnitine supplementation and early start of emergency treatment during putatively threatening episodes of intermittent febrile illness dramatically improves the outcome and thus has been recommended by an international guideline group (Kölker et al, J Inherit Metab Dis 30:5-22, 2007). However, possible affection of linear growth, weight gain and biochemical follow-up monitoring has not been studied systematically. METHODS: Thirty-three patients (n = 29 asymptomatic, n = 4 dystonic) with GA-I who have been identified by newborn screening in Germany from 1999 to 2009 were followed prospectively during the first six years of life. Dietary treatment protocols, anthropometrical and biochemical parameters were longitudinally evaluated. RESULTS: Mean daily intake as percentage of guideline recommendations was excellent for lysine (asymptomatic patients: 101 %; dystonic patients: 103 %), lysine-free, tryptophan-reduced AAS (108 %; 104 %), energy (106 %; 110 %), and carnitine (92 %; 102 %). Low lysine diet did not affect weight gain (mean SDS 0.05) but mildly impaired linear growth in asymptomatic patients (mean SDS -0.38), while dystonic patients showed significantly reduced weight gain (mean SDS -1.32) and a tendency towards linear growth retardation (mean SDS -1.03). Patients treated in accordance with recent recommendations did not show relevant abnormalities of routine biochemical follow-up parameters. INTERPRETATION: Low lysine diet promotes sufficient intake of essential nutrients and anthropometric development in asymptomatic children up to age 6 year, whereas individualized nutritional concepts are required for dystonic patients. Revised recommendations for biochemical monitoring might be required for asymptomatic patients.

High-Throughput Analysis of Underivatized Amino Acids and Acylcarnitines in Infant Serum: A Micromethod Based on Stable Isotope Dilution Targeted HILIC-ESI-MS/MS.

Amino acids and acylcarnitines are important biomarkers of the body's energy state and can be used as diagnostic markers of certain inborn errors of metabolism. Few multianalyte methods for high-throughput analysis in serum exist for these compounds, but micromethods suitable for use in young children and infants are lacking. Therefore, we developed a quantitative high-throughput multianalyte hydrophilic interaction liquid chromatography-tandem mass spectrometry method preceded by a derivatization-free sample preparation using minimum amounts of serum (25 µL). Isotopically labeled standards were utilized for quantification. Forty amino acids and amino acid derivatives and 22 acylcarnitines were detected by applying a multiple reaction monitoring mode within a 20 min run. The method was comprehensively validated, comprising linearity, accuracy, (intraday/interday) precision, and quantitation limits, of which the latter ranged from 0.25 to 50 nM for acylcarnitines and from 0.005 to 1 µM for amino acids and their derivatives. Application of the method to 145 serum samples of three- to four-month-old healthy infants showed excellent reproducibility for multiday analyses and enabled simultaneous amino acid and acylcarnitine profiling in this age group.