Mutations in the accessory subunit NDUFB10 result in isolated complex I deficiency and illustrate the critical role of intermembrane space import for complex I holoenzyme assembly.

An infant presented with fatal infantile lactic acidosis and cardiomyopathy, and was found to have profoundly decreased activity of respiratory chain complex I in muscle, heart and liver. Exome sequencing revealed compound heterozygous mutations in NDUFB10, which encodes an accessory subunit located within the PD part of complex I. One mutation resulted in a premature stop codon and absent protein, while the second mutation replaced the highly conserved cysteine 107 with a serine residue. Protein expression of NDUFB10 was decreased in muscle and heart, and less so in the liver and fibroblasts, resulting in the perturbed assembly of the holoenzyme at the 830 kDa stage. NDUFB10 was identified together with three other complex I subunits as a substrate of the intermembrane space oxidoreductase CHCHD4 (also known as Mia40). We found that during its mitochondrial import and maturation NDUFB10 transiently interacts with CHCHD4 and acquires disulfide bonds. The mutation of cysteine residue 107 in NDUFB10 impaired oxidation and efficient mitochondrial accumulation of the protein and resulted in degradation of non-imported precursors. Our findings indicate that mutations in NDUFB10 are a novel cause of complex I deficiency associated with a late stage assembly defect and emphasize the role of intermembrane space proteins for the efficient assembly of complex I.

Neonatal overfeeding alters hypothalamic microglial profiles and central responses to immune challenge long-term.

The early life period is one of significant vulnerability to programming effects from the environment. Given the sensitivity of microglial cells to early life programming and to adult diet, we hypothesized overfeeding during the neonatal period would acutely alter microglial profiles within the developing brain, predisposing the individual to a lasting central pro-inflammatory profile that contributes to overactive immune responses long-term. We tested this idea by manipulating litter sizes in which Wistar rat pups were raised, so the pups were suckled in litters of 4 (neonatally overfed) or 12 (control). This manipulation induces obesity and susceptibility to lipopolysaccharide (LPS) long-term. We then examined microglial and central pro-inflammatory profiles during development and in adulthood as well as susceptibility to neuroimmune challenge with LPS. Neonatally overfed rats have evidence of microgliosis in the paraventricular nucleus of the hypothalamus (PVN) as early as postnatal day 14. They also show changes in hypothalamic gene expression at this time, with suppressed hypothalamic interleukin 1ß mRNA. These effects persist into adulthood, with basal PVN microgliosis and increased hypothalamic toll-like receptor 4, nuclear factor κB, and interleukin 6 gene expression. These neonatally overfed rats also have dramatically exacerbated microglial activation in the PVN 24h after an adult LPS challenge, coupled with changes in inflammatory gene expression. Thus, it appears neonatal overfeeding sensitizes PVN microglia, contributing to a basal pro-inflammatory profile and an altered response to a neuroimmune challenge throughout life. It remains to be seen if these effects can be reversed with early interventions.

Gastrointestinal and nutritional problems occur frequently throughout life in girls and women with Rett syndrome.

OBJECTIVE: We conducted a nationwide survey to determine the prevalence of common gastrointestinal and nutritional disorders in Rett syndrome (RTT) based on parental reporting and related the occurrence of these problems to age and methyl-CpG-binding protein 2 (MECP2) gene status. METHODS: We designed a questionnaire that probed symptoms, diagnoses, diagnostic tests, and treatment interventions related to gastrointestinal and nutritional problems in RTT. The International Rett Syndrome Foundation distributed the questionnaire to 1666 family-based members and forwarded their responses for our review. We interrogated the Rare Disease Clinical Research Network database to supplement findings related to medications used to treat gastrointestinal problems in RTT. RESULTS: Parents of 983 female patients with RTT (59%) responded and identified symptoms and diagnoses associated with gastrointestinal dysmotility (92%), chewing and swallowing difficulties (81%), weight deficits or excess (47%), growth deficits (45%), low bone mineral content or fractures (37%), and biliary tract disorders (3%). Height-for-age, weight-for-age, and body mass index z scores decreased significantly with age; height- and weight-, but not body mass index-for-age z scores were significantly lower in female subjects with MECP2 mutations than in those without. Vomiting, nighttime awakening, gastroesophageal reflux, chewing difficulty, and choking with feeding were significantly less likely to occur with increasing age. Short stature, low bone mineral content, fractures, and gastrostomy placement were significantly more likely to occur with increasing age. Chewing difficulty, choking with feeding, and nighttime awakening were significantly less likely to occur, whereas short stature was significantly more likely to occur, in female subjects with MECP2 mutations than in those without. Diagnostic evaluations and therapeutic interventions were used less frequently than the occurrence of symptoms or diagnoses in the RTT cohort. CONCLUSIONS: Gastrointestinal and nutritional problems perceived by parents are prevalent throughout life in girls and women with RTT and may pose a substantial medical burden for their caregivers. Physician awareness of these features of RTT may improve the health and quality of life of individuals affected with this disorder.

Epigenetic malprogramming of the insulin receptor promoter due to developmental overfeeding.

AIM: Prenatal and neonatal overfeeding programs a permanent obesity and diabetes disposition, e.g., due to induction of hypothalamic insulin resistance. We investigated acquired alterations of the DNA methylation pattern of the hypothalamic insulin receptor promoter (IRP) which might be an underlying molecular mechanism. METHODS: Neonatal overfeeding was induced by rearing Wistar rats in small litters (SL). Methylation of CpG-dinucleotides of the hypothalamic IRP was mapped using bisulfite sequencing. RESULTS: Neonatal overfeeding led to rapid early weight gain, resulting in a metabolic syndrome phenotype, i.e., obesity, hyperleptinemia, hyperglycemia, hyperinsulinemia, and increased insulin/glucose-ratio. The proportion of animals carrying any methylated CpG residue in the 322 bp CpG island of the IRP was increased in neonatally overfed SL rats (n=8), as compared to controls (n=8; P=0.04). Moreover, the mean percentage of methylated CpG positions was also higher in SL rats (P=0.01). Over both groups, neonatal blood glucose levels were positively correlated to the extent of promoter methylation (r=0.52; P=0.04). CONCLUSIONS: This study characterizes for the first time the IRP epigenomically in any species and tissue. Our data reveal that the IRP is vulnerable to hypermethylation due to overnutrition, probably especially glucose-dependent in a dose-response manner. This paradigmatically indicates the impact of nutrient-dependent epigenetic malprogramming, leading to a "diabesity" disposition which may become pathogenic throughout life.

Outpatient transition of an infant with permanent neonatal diabetes due to a KCNJ11 activating mutation from subcutaneous insulin to oral glyburide.

Neonatal diabetes mellitus is rare, may either be transient or permanent, and may be caused by mutations in any of the several different genes. Until recently, most forms of permanent neonatal diabetes required lifelong subcutaneous insulin for management; however, permanent neonatal diabetes due to activating mutations in the KCNJ11 gene, which encodes the Kir6.2 protein subunit of the ATP-sensitive K+ (K(ATP)) channel, may be amenable to oral sulfonylurea therapy. We describe a case of an 18-month-old infant with permanent neonatal diabetes due to an activating KCNJ11 mutation successfully transitioned from subcutaneous insulin therapy to oral sulfonylurea therapy in the outpatient setting.

The inheritance of obesity.

Syndromic adiposity appears to have a predisposition to run in families suggesting a hereditary element in its transmission. Purely genetic defects and DNA sequence variants have been directly associated with the development of adiposity; however, these account for a very small proportion of cases. A stronger association has been made between the intrauterine and early childhood nutritional environment of the foetus and young child and the predisposition of childhood and subsequent adulthood obesity. The nutritional environments include both a situation of nutritional deprivation or excess working through the interplay of epigenetic changes, and pancreatic and hypothalamic development. This is further compounded by the nutritional and lifestyle attitudes of the particular at-risk family. Adiposity prevention measures must include reenforced intervention strategies stating with lifestyle education schemes during pregnancy followed through until infancy and early childhood especially in those families/individuals identified as being at a risk of developing significant adiposity.

The effects of infantile malnutrition on behavioral development: a follow-up study.

Thirty-one children and young adults who had been severely malnourished in infancy due to intestinal disease, were compared with sibling controls for psychometric intelligence, academic performance as judged by teacher ratings, and developmental history as judged by parental questionnaires. Older members of the sample were also examined on the Lincoln-Oseretzky Motor Development Scale and by a brief psychiatric interview. There were no significant differences between patients and controls on any outcome measure. Furthermore, previously malnourished individuals were performing at or above age equivalent norms on all psychological tests. The findings suggest that the adverse behavioral effects of severe infantile malnutrition observed in children below the age of 5 yr are, to a large extent, compensated during development when the children are raised in supportive home and school environments.

Hydrogen peroxide-induced DNA damage and DNA repair in lymphocytes from malnourished children.

The aim of this study was to assess DNA repair capacity in lymphocytes of children with protein calorie malnutrition using the single-cell gel electrophoresis (comet) assay. Repair capacity was assessed by estimating the relative decrease of DNA migration length 5, 15, 30, and 60 min after hydrogen peroxide treatment, in three groups of children: well-nourished (WN), well-nourished infected (WN-I), and malnourished infected (MN-I). In addition, the DNA migration length was evaluated in all groups before and after peroxide treatment. Comparison of mean migration lengths observed in WN and WN-I children showed significant differences at all times tested; between WN-I and MN-I differences were also observed, except after hydrogen peroxide exposure. This implies that lymphocytes of WN-I and MN-I children were equally sensitive to hydrogen peroxide. Nevertheless, the MN-I group clearly shows the greatest overall percentage of damaged cells at all times tested. In relation to repair capacity, at 5 min it was approximately 30% in both groups of well-nourished children, but only 20% in MN-I; 15 min after exposure, repair capacity increased to 51% in well-nourished children but only to 31% in MN-I; and at 60 min this capacity increased to 82% in well-nourished but only to 55% in MN-I. These data indicate that lymphocytes of malnourished children show a decreased capacity to repair hydrogen peroxide-induced DNA damage compared to that of well-nourished controls. This reflects that only malnutrition is associated with decreased DNA repair capacity. Additionally, the data confirm that severe infection and malnutrition are two factors clearly associated with increased DNA damage.

An evaluation of nutrition centre effectiveness by measurement of younger siblings.

Thirty children were measured when they entered nutrition centres. Their younger siblings were later measured at the same age. Over the same period, a matched group of control children and their siblings were measured. The younger siblings of Centre participants showed the same growth patterns as the younger siblings of control children. Centre teaching had evidently not affected the way mothers fed their younger children, and thus it had no effect on their growth.

Sister-chromatid exchange (SCE) and cell proliferation in lymphocytes from infected and non-infected children with severe protein calorie malnutrition (PCM).

The frequency of sister-chromatid exchanges (SCE) and the rate of cell proliferation were evaluated through differential staining of sister chromatids in mitogen-stimulated cultured lymphocytes sampled from five well-nourished children, from seven severely malnourished children infected with bacterium, and from 10 severely malnourished children following treatment for infection with antimicrobial drugs 2 weeks before blood sampling. The replication indices at 48 h of culture were higher in both groups of malnourished children than in the well-nourished children, indicating either a faster response to PHA and/or a shorter cell cycle in lymphocytes of these patients. The average frequency of SCE per mitosis was also significantly higher than in the control group. The mitotic index was similar in the three groups of children. The lack of significant difference in response between the two groups of malnourished children suggests that the effects observed at the cytogenetic level are caused by severe malnutrition per se, and not by any associated infection.

Animal models of epigenetic inheritance.

Although genomic DNA is the template of our heredity, it is the coordination and regulation of its expression that results in the wide complexity and diversity seen among organisms. In recent years, an emerging body of evidence has focused on the role of epigenetics as one mechanism by which gene expression can be maintained and modulated throughout the lifetime of an individual. Epigenetics refers to heritable alterations in gene expression that are not mediated by changes in primary DNA sequence and includes mitotic and/or meiotic events. In essence, epigenetic modulation results in functional adaptations of the genomic response to the environment and is believed to play a fundamental role in early developmental plasticity. This article focuses on several animal models that have been developed over the past decade to study epigenetic inheritance, many of which have arisen from the developmental origins of adult health and disease fields.

Developmental origins of health and disease: brief history of the approach and current focus on epigenetic mechanisms.

"Barker's hypothesis" emerged almost 25 years ago from epidemiological studies of birth and death records that revealed a high geographic correlation between rates of infant mortality and certain classes of later adult deaths as well as an association between birthweight and rates of adult death from ischemic heart disease. These observations led to a theory that undernutrition during gestation was an important early origin of adult cardiac and metabolic disorders due to fetal programming that permanently shaped the body's structure, function, and metabolism and contributed to adult disease. This theory stimulated interest in the fetal origins of adult disorders, which expanded and coalesced approximately 5 years ago with the formation of an international society for developmental origins of health and disease (DOHaD). Here we review a few examples of the many emergent themes of the DOHaD approach, including theoretical advances related to predictive adaptive responses of the fetus to a broad range of environmental cues, empirical observations of effects of overnutrition and stress during pregnancy on outcomes in childhood and adulthood, and potential epigenetic mechanisms that may underlie these observations and theory. Next, we discuss the relevance of the DOHaD approach to reproductive medicine. Finally, we consider the next steps that might be taken to apply, evaluate, and extend the DOHaD approach.

Newborn screening of metabolic disorders: recent progress and future developments.

Tandem mass spectrometry has been the main driver behind a significant expansion in newborn screening programs. The ability to detect more than 40 conditions by a single test underscores the need to better understand the clinical and laboratory characteristics of the conditions being tested, and the complexity of pattern recognition and differential diagnoses of one or more elevated markers. The panel of conditions recommended by the American College of Medical Genetics, including 20 primary conditions and 22 secondary targets that are detectable by tandem mass spectrometry has been adopted as the standard of care in the vast majority of US states. The evolution of newborn screening is far from being idle as a large number of infectious, genetic, and metabolic conditions are currently under investigation at variable stages of test development and clinical validation. In the US, a formal process with oversight by the Advisory Committee on Heritable Disorders and Genetic Diseases in Newborns and Children has been established for nomination and evidence-based review of new candidate conditions. If approved, these conditions could be added to the uniform panel and consequently pave the way to large scale implementation.

Metabolic profiling.

The concept of chemical individuality was introduced by Garrod in 1908. Inheritance of Mendelian traits including disease states has finally reached a new level of understanding based on the modern principles of gene expression coupled with new insight into the metabolism of RNA species and protein. Over 300 different perturbations in metabolite profiles with their identifying alteration(s) in protein and/or gene structure and/or function have been identified in the past 100 years. With the realization in 1953 that the sentinel disease, phenylketonuria, can be effectively treated by nutritional manipulation tailored to the needs of each individual, we have essentially entered a new phase in metabolic medicine, namely that of nutritional therapeutics. The infant destined for a lifetime of cognitive and motoric handicaps may be rescued by the implementation of a nutritional prescription in early development. Patients with inherited defects that impact on intermediary metabolism need to receive nutritional therapy on an individualized basis. Metabolic profiling, i.e., the array of small molecules or analytes, as well as large macromolecules, measured with precision in body fluids or tissues, can be used to devise a nutritional therapeutic plan, as well as serve as endpoints to evaluate the biochemical efficacy of intervention.

Do we need personalized recommendations for infants at risk of developing disease?

Current nutrition recommendations, directed towards populations, are based on estimated average nutrient requirements for a target population and intend to meet the needs of most individuals within that population. They also aim at preventing common diseases such as obesity, diabetes and cardiovascular disease. For infants with specific genetic polymorphisms, e.g. some inborn errors of metabolism, adherence to current recommendations will cause disease symptoms and they need personalized nutrition recommendations. Some other monogenic polymorphisms, e.g. adult hypolactasia, are common but with varying prevalence between ethnic groups and within populations. Ages at onset as well as the degree of the resulting lactose intolerance also vary, making population-based as well as personalized recommendations difficult. The tolerable intake is best set by each individual based on symptoms. For polygenetic diseases such as celiac disease, type-1 diabetes and allergic disease, current knowledge is insufficient to suggest personalized recommendations aiming at primary prevention for all high-risk infants, although it may be justified to provide such recommendations on an individual level should the parents ask for them. New technologies such as nutrigenetics and nutrigenomics are promising tools with which current nutrition recommendations can possibly be refined and the potential of individualized nutrition be explored. It seems likely that in the future it will be possible to offer more subgroups within a population personalized recommendations.

Expression of cytokine mRNA in lymphocytes of malnourished children.

INTRODUCTION: Protein-calorie malnutrition represents a significant worldwide health problem and is associated with an increased risk for infections. The purpose of this study was to evaluate possible changes in type 1/type 2 responses balance in malnourished children. RESULTS: The data obtained in the present study showed that the expression levels of tumor necrosis factor-alpha, interleukin (IL)-4, and IL-10 were more highly, in contrast IL-2, gamma interferon, and IL-6 genes were expressed less in all groups of malnourished children compared with the well-nourished infected children. It is important to indicate that the data collected in the present work agree with the results obtained by different authors, who showed differences in the production of cytokines in malnourished children. CONCLUSION: In conclusion, the results suggest that alterations in the balance of type 1/type 2 immune responses exist in malnourished children, and this could be the reason that the immunological system of the malnourished children is incapable of eradicating infections.

Unexplained diarrhoea and failure to thrive in 2 siblings with unusual facies and abnormal scalp hair shafts: a new syndrome.

A family is described in which 2 siblings born to healthy parents presented with abnormal facies, persistent diarrhoea, and early death. Exhaustive pathological and biochemical investigations failed to find a cause. The scalp hair of both babies had an abnormal amino-acid composition, and presented an appearance that was unique on scanning electron microscopical examination; this fact and the clinical picture probably represents a new syndrome.