Integration of CTCF loops, methylome, and transcriptome in differentiating LUHMES as a model for imprinting dynamics of the 15q11-q13 locus in human neurons.

Human cell line models, including the neuronal precursor line LUHMES, are important for investigating developmental transcriptional dynamics within imprinted regions, particularly the 15q11-q13 Angelman (AS) and Prader-Willi (PWS) syndrome locus. AS results from loss of maternal UBE3A in neurons, where the paternal allele is silenced by a convergent antisense transcript UBE3A-ATS, a lncRNA that terminates at PWAR1 in non-neurons. qRT-PCR analysis confirmed the exclusive and progressive increase in UBE3A-ATS in differentiating LUHMES neurons, validating their use for studying UBE3A silencing. Genome-wide transcriptome analyses revealed changes to 11 834 genes during neuronal differentiation, including the upregulation of most genes within the 15q11-q13 locus. To identify dynamic changes in chromatin loops linked to transcriptional activity, we performed a HiChIP validated by 4C, which identified two neuron-specific CTCF loops between MAGEL2-SNRPN and PWAR1-UBE3A. To determine if allele-specific differentially methylated regions (DMR) may be associated with CTCF loop anchors, whole genome long-read nanopore sequencing was performed. We identified a paternally hypomethylated DMR near the SNRPN upstream loop anchor exclusive to neurons and a paternally hypermethylated DMR near the PWAR1 CTCF anchor exclusive to undifferentiated cells, consistent with increases in neuronal transcription. Additionally, DMRs near CTCF loop anchors were observed in both cell types, indicative of allele-specific differences in chromatin loops regulating imprinted transcription. These results provide an integrated view of the 15q11-q13 epigenetic landscape during LUHMES neuronal differentiation, underscoring the complex interplay of transcription, chromatin looping, and DNA methylation. They also provide insights for future therapeutic approaches for AS and PWS.

Phosphoserine inhibits neighboring arginine methylation in the RKS motif of histone H3.

The effects of phosphorylation of histone H3 at serine 10 have been studied in the context of other posttranslational modifications such as lysine methylation. We set out to investigate the impact of phosphoserine-10 on arginine-8 methylation. We performed methylation reactions using peptides based on histone H3 that contain a phosphorylated serine and compared the extent of arginine methylation with unmodified peptides. Results obtained via fluorography indicate that peptides containing a phosphorylated serine-10 inhibit deposition of methyl groups to arginine-8 residues. To further explore the effects of phosphoserine on neighboring arginine residues, we physically characterized the non-covalent interactions between histone H3 phosphoserine-10 and arginine-8 using 31P NMR spectroscopy. A salt bridge was detected between the negatively charged phosphoserine-10 and the positively charged unmodified arginine-8 residue. This salt bridge was not detected when arginine-8 was symmetrically dimethylated. Finally, molecular simulations not only confirm the presence of a salt bridge but also identify a subset of electrostatic interactions present when arginine is replaced with alanine. Taken together, our work suggests that the negatively charged phosphoserine maximizes its interactions. By limiting its exposure and creating new contacts with neighboring residues, it will inhibit deposition of neighboring methyl groups, not through steric hindrance, but by forming intrapeptide interactions that may mask substrate recognition. Our work provides a mechanistic framework for understanding the role of phosphoserine on nearby amino acid residues and arginine methylation.

Integration of CTCF Loops, Methylome, and Transcriptome in Differentiating LUHMES as a Model for Imprinting Dynamics of the 15q11-q13 Locus in Human Neurons.

Human cell line models, including the neuronal precursor line LUHMES, are important for investigating developmental transcriptional dynamics within imprinted regions, particularly the 15q11-q13 Angelman (AS) and Prader-Willi (PWS) syndrome locus. AS results from loss of maternal UBE3A in neurons, where the paternal allele is silenced by a convergent antisense transcript UBE3A-ATS, a lncRNA that normally terminates at PWAR1 in non-neurons. qRTPCR analysis confirmed the exclusive and progressive increase in UBE3A-ATS in differentiating LUHMES neurons, validating their use for studying UBE3A silencing. Genome-wide transcriptome analyses revealed changes to 11,834 genes during neuronal differentiation, including the upregulation of most genes within the 15q11-q13 locus. To identify dynamic changes in chromatin loops linked to transcriptional activity, we performed a HiChIP validated by 4C, which identified two neuron-specific CTCF loops between MAGEL2-SNRPN and PWAR1-UBE3A. To determine if allele-specific differentially methylated regions (DMR) may be associated with CTCF loop anchors, whole genome long-read nanopore sequencing was performed. We identified a paternally hypomethylated DMR near the SNRPN upstream loop anchor exclusive to neurons and a paternally hypermethylated DMR near the PWAR1 CTCF anchor exclusive to undifferentiated cells, consistent with increases in neuronal transcription. Additionally, DMRs near CTCF loop anchors were observed in both cell types, indicative of allele-specific differences in chromatin loops regulating imprinted transcription. These results provide an integrated view of the 15q11-q13 epigenetic landscape during LUHMES neuronal differentiation, underscoring the complex interplay of transcription, chromatin looping, and DNA methylation. They also provide insights for future therapeutic approaches for AS and PWS.

Validation of the Children's International Mucositis Evaluation Scale (ChIMES) in paediatric cancer and SCT.

BACKGROUND: Objectives were to describe the reliability and validity of a new paediatric-specific mucositis scale, the Children's International Mucositis Evaluation Scale (ChIMES). METHODS: In a multi-centre prospective study, children aged 0 to ≤18 years were eligible if they were receiving any of the following: myeloablative stem cell transplantation (SCT), ≥60 mg m(-2) course(-1) doxorubicin or ≥12 g m(-2) methotrexate. Multiple measures of mucositis were included along with ChIMES. Respondents were parent proxy report for children aged <12 years, and child self-report for children aged 12-18 years and 8 to <12 years. Mucositis diaries were completed at baseline and on Days 7-17 following chemotherapy/conditioning. On Day 14, the respondent reported presence of mucositis and change since the previous day. RESULTS: The 185 respondents included parents (N=98), children aged 12-18 years (N=66) and children aged 8 to <12 years (N=21). Test-retest reliability was excellent for ChIMES Total Score and ChIMES Percentage Score with r>0.8 for all respondent types. Criteria for construct validation were met across all measures. ChIMES also demonstrated responsiveness with significant differences between baseline and Day 14. CONCLUSION: ChIMES is a paediatric-specific measure of mucositis with favourable psychometric properties. It exhibits reliability, construct validity and responsiveness. ChIMES should be incorporated into clinical trials of mucositis prevention and treatment in paediatric cancer and SCT.

Epigenetics in Prader-Willi Syndrome.

Prader-Willi Syndrome (PWS) is a rare neurodevelopmental disorder that affects approximately 1 in 20,000 individuals worldwide. Symptom progression in PWS is classically characterized by two nutritional stages. Stage 1 is hypotonia characterized by poor muscle tone that leads to poor feeding behavior causing failure to thrive in early neonatal life. Stage 2 is followed by the development of extreme hyperphagia, also known as insatiable eating and fixation on food that often leads to obesity in early childhood. Other major features of PWS include obsessive-compulsive and hoarding behaviors, intellectual disability, and sleep abnormalities. PWS is genetic disorder mapping to imprinted 15q11.2-q13.3 locus, specifically at the paternally expressed SNORD116 locus of small nucleolar RNAs and noncoding host gene transcripts. SNORD116 is processed into several noncoding components and is hypothesized to orchestrate diurnal changes in metabolism through epigenetics, according to functional studies. Here, we review the current status of epigenetic mechanisms in PWS, with an emphasis on an emerging role for SNORD116 in circadian and sleep phenotypes. We also summarize current ongoing therapeutic strategies, as well as potential implications for more common human metabolic and psychiatric disorders.

Dental disease in type 3 Von Willebrand disease: a neglected problem.

Type 3 Von Willebrand disease (VWD) is a rare, severe, autosomal recessive bleeding disorder. In our institution, we follow 17 children with type 3 VWD. We have observed a high prevalence of dental disease in these patients prompting us to undertake a retrospective review of our cohort of patients with type 3 VWD to catalogue the extent of their dental disease. Sixteen of these patients have been assessed by our dentistry department. Five children have undergone minor dental procedures (e.g. restorations, stainless steel crowns) and seven major procedures (e.g. dental extractions, pulpotomies and root canal treatments). These patients have collectively used 85,400 (ristocetin cofactor) IU of Humate-P on dental procedures alone. In addition to the considerable costs of factor are the cost of operating room time, dentists' costs, and the cost of other topical haemostatic agents (e.g. Tisseel) used during their dental procedures. As such there is considerable morbidity and cost from dental disease in these patients that is much higher than what is seen in patients with haemophilia or in the normal paediatric population. We speculate that the combination of these patients having a significant mucosal bleeding disorder together with various socioeconomic factors contribute to the significant degree of dental disease seen in this group of patients. We would suggest that better preventive dental care needs to be provided to these patients to avoid the considerable morbidity and very high burden of dental disease in type 3 VWD.

Knowledge, compliance and serum phenylalanine concentrations in adolescents and adults with phenylketonuria and the effect of a patient-focused educational resource.

BACKGROUND: There is a lack of dedicated resources for adolescent and adult patients with phenylketonuria (PKU) and few studies have examined dietary practices within this group. METHODS: One hundred and seventy-seven PKU patients were sent questionnaires to assess dietary compliance and the preferred format for an educational resource. Seventy-one patients responded; 32 following diet were recruited to assess the resource's impact on test variables. The results were compared for the intervention group (n = 22) and control group (n = 10) at baseline, and 1 and 6 months after resource intervention. RESULTS: Most patients were aware of dietary recommendations, although this did not always result in compliance. The preferred resource format was a filofax-style folder with inserts (P < 0.05). There was a significant difference in the extent of change in knowledge score between baseline and 1 month in favour of the intervention group (P < 0.05). The improvement in knowledge was not accompanied by a significant improvement in measures of compliance. CONCLUSIONS: These findings add to the knowledge base about this patient group and support the use of patient involvement in resource development. It is likely that the test parameters used were not sensitive enough to pick up subtle and longer-term effects on compliance.

Serial controlled N-of-1 trials of topical vitamin E as prophylaxis for chemotherapy-induced oral mucositis in paediatric patients.

The objectives were (1) to determine whether in children undergoing doxorubicin-containing chemotherapy, topical vitamin E decreases an objective measurement of oral mucositis compared to placebo, and (2) to assess the feasibility of an innovative trial design in paediatric cancer, combining N-of-1 trials using Bayesian meta-analysis. We conducted a series of N-of-1, double-blinded, randomised controlled trials in children > or = 6 years of age receiving repeated cycles of identical doxorubicin-containing chemotherapy. Each study cycle was followed by topical vitamin E (800 mg) or placebo. We enroled 16 children and 45 post chemotherapy cycles were randomised to vitamin E (N=22) or placebo (N=23). There was no difference in objective mucositis scores with a mean score of 0.2 with vitamin E and 0.3 with placebo. Topical vitamin E does not reduce doxorubicin-induced oral mucositis in children. The use of N-of-1 studies and Bayesian meta-analysis may facilitate the study of some therapies in paediatric oncology.

The effect of rolled oats on blood lipids and fecal steroid excretion in man.

Rolled oats (125 g daily) were substituted for breakfast cereals and wheat flour in the metabolically controlled diets of 10 subjects for 3 wk. Fat and energy intakes in the 2-wk control periods before and after the oat period were adjusted by addition of an oil with a similar fatty acid composition to the lipid in the oats. Plasma total cholesterol concentrations were reduced in seven of 10 subjects, but over the whole group the mean reduction of 8% was not significant (0.05 less than p less than 0.01). High-density lipoprotein cholesterol concentrations and plasma triglyceride levels were unchanged. Fecal fat excretion was increased by 47% (p less 0.005) and fecal bile acid excretion by 35% (p less than 0.01) but neutral steroid excretion was unchanged on the oatmeal diet.

Wheat bread supplemented with depolymerized guar gum reduces the plasma cholesterol concentration in hypercholesterolemic human subjects.

Recent human studies have shown that the physiologic effects of guar gum are not diminished by partial depolymerization of its galactomannan fraction. We evaluated the effect of depolymerized guar galactomannan on fasting plasma cholesterol and triacylglycerol concentrations in healthy volunteers with moderately raised plasma cholesterol concentrations (range: 5.2-8.0 mmol/L). This study was designed as a randomized, double-blind crossover of two 3-wk feeding periods separated by a 4-wk washout period. Control and guar wheat breads were prepared by a commercial bread-making process. Subjects (n = 11) were asked to replace their normal bread with that provided, receiving control bread for one 3-wk period and guar bread for the other period, without altering their baseline diet. Subjects recorded their intake of foods for 6 consecutive days on three occasions during the study. Fasting venous blood samples (10 mL) were taken from subjects on two consecutive mornings at the start and end of each feeding period. No significant changes in body weight or dietary intake were recorded in the control and guar bread periods. There was a significant reduction (10%) in total plasma cholesterol concentration after the guar treatment (P < 0.001), mainly because of a reduction in the low-density-lipoprotein-cholesterol fraction. No changes in plasma high-density-lipoprotein-cholesterol or triacylglycerol concentrations were seen. The cholesterol-lowering effect of partially depolymerized guar gum appears to be of a magnitude similar to that of high-molecular-weight guar gum used in earlier studies.

Randomized trial of the effects of cholesterol-lowering dietary treatment on psychological function.

PURPOSE: Epidemiological studies have suggested that cholesterol lowering could affect psychological functioning. This study was designed to test whether cholesterol-lowering diets adversely affect mood and cognitive function.5.2 mM [198 mg/dL]) to either a low-fat diet, a Mediterranean diet, or a waiting-list control. Cholesterol levels, psychological well-being (depression, anxiety, hostility), and cognitive function were assessed at baseline, 6 weeks, and 12 weeks. RESULTS: Total serum cholesterol levels fell significantly more in the intervention groups (8.2% reduction) than in the control group (P <0.001). All three groups showed a modest improvement in psychological well-being during the 12-week treatment period, but there were no differences among the groups. There were no between-group differences on three measures of cognitive function, but for a fourth measure, which involved the task with the greatest processing load, the two intervention groups did significantly worse (P <0.001) than the control group. The change in performance was correlated with the change in total serum cholesterol level (r = 0. 21, P = 0.01). CONCLUSIONS: Two dietary interventions that successfully lowered serum cholesterol levels had no adverse effect on mood. There was some evidence for a relative impairment in cognitive function in the treated groups in one of four cognitive tests, but additional studies will be required to determine the relevance of this finding.

Assessment of fecal output in patients receiving enteral tube feeding: validation of a novel chart.

OBJECTIVE: To develop a practical, valid and reliable chart to assist in the accurate visual characterization of fecal output in patients receiving enteral tube feeding (ETF). DESIGN: A chart incorporating verbal and pictorial descriptors of fecal output was developed. Validity and reliability were assessed by a questionnaire survey of health professionals and a clinical study of patients commencing ETF. Content validity was assessed from the results of the questionnaire, construct validity by contrasting groups analysis, concurrent validity by comparison of the chart with actual fecal weight and inter-rater reliability by independent characterization of the same fecal sample by two nurses. SETTING: St George's Hospital, London, UK. SUBJECTS: In all, 35 health professionals completed a questionnaire with respect to the chart. The chart was then used to monitor fecal output in 36 patients commencing ETF for a total of 171 patient-days, during which time nurses used the chart to characterize 269 fecal samples, of which 59 were subsequently weighed. RESULTS: The results of the questionnaire suggested good content validity. The chart demonstrated statistically significant differences in fecal frequency, fecal consistency, fecal score and incidence of diarrhea for contrasting patient groups expected to have different fecal output (P < 0.05). The inter-rater reliability was almost perfect for fecal consistency (95% agreement, kappa = 0.91) and substantial for fecal weight (83%, kappa = 0.75). In all, 83% of fecal samples were assigned to the correct weight category (kappa = 0.75). CONCLUSION: The chart has good content, construct and concurrent validity and inter-rater reliability, and is suitable for both research and clinical use. SPONSORSHIP: This research was jointly supported by King's College London and Nestlé, UK.

Frequency of consumption and nutrient composition of composite dishes commonly consumed in the UK by South Asian Muslims originating from Bangladesh, Pakistan and East Africa (Ismailis).

INTRODUCTION: This paper presents information on the nutrient composition of commonly consumed traditional dishes eaten by the three major South Asian Muslim groups residing in Britain, namely Bangladeshi, Pakistani and East African Ismaili Muslims. METHODS: Information regarding the most common dishes consumed by South Asian Muslims originating from Bangladesh, Pakistan and East Africa (Ismailis) and living in London was obtained from 7-day menu records over two seasons. For each common dish, weighed recipes were collected in triplicate and the composition (energy and selected nutrients) was calculated from the ingredients and cooked weight of the dish. RESULTS: The three Muslim groups showed considerable variation in traditional foods commonly consumed as well as variation in fat and energy contents of similar recipes both within and between groups. Nutrient composition of commonly consumed dishes is presented calculated from the recipe nearest the average in terms of fat and energy for a particular dish. DISCUSSION AND CONCLUSIONS: The potential uses of the data, one of which would be to improve dietary compliance (especially to lower fat intakes) amongst the three diverse South Asian Muslim groups, are discussed.

Composite resin short-post technique for primary anterior teeth.

A technique using a "mushroom shaped" composite resin short post constructed inside the pulp-treated root canal provides sufficient retention to build a composite resin crown on the reinforced superstructure of the remaining crown dentin. This technique was tested for 1 year in 92 teeth; they showed no failures of retention of the short post. Recurrent caries and severe bruxism--factors beyond operator control--posed some problems that were readily resolved.

The effectiveness of residential care facilities for adolescent boys.

This article reports the findings of a 1984 study evaluating the effectiveness of a well-established residential child-care facility for adolescent boys. Program outcomes for 101 males between the ages of eleven and seventeen were examined in relation to a wide range of demographic, social history, and program participation variables. Characteristics found to be significantly associated with program outcome included: race; probation; drug abuse; program intervention; home visits; and runaway behavior. In a separate analysis, three factors were found to be significantly related to runaway behavior, the chief cause of program failure: race; placement under court supervision; and program intervention.

Feeding disorders research and teams: a new challenge for dentistry.

Feeding disorders, which include masticatory and cerebral disabilities, present a major problem for disabled persons, but only in the last decade have health professionals shown an increased interest in the diagnosis and management of patients with such problems. Dentists are experts in the diagnosis and treatment of disorders of the oral-facial region, yet they have either worked in intradisciplinary isolation or have not demonstrated their interest in patients with feeding disorders. Consequently, those involved in the assessment and treatment of persons with feeding disorders are not aware of the contributions dentists have made to the basic and clinical data base of feeding assessment and therapy. Dentistry is simply not mentioned in rehabilitation. The opportunity now exists for dentistry to inform others of our expertise in the field from both a research and a clinical perspective. The basic and clinical base of dental therapeutics and its role in the diagnosis and treatment of disorders of the masticatory apparatus and of swallowing are reviewed here. It is hoped that this review will stimulate interest among dentists to take part in the rehabilitation of patients with feeding disorders through the use of the extensive data base within the profession.

Characteristics and perspectives of night-eating behaviour in a severely obese population.

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT: Night-eating syndrome (NES) can be a feature of severe obesity. NES is a dysfunction of circadian rhythm and is associated with impaired sleep. WHAT THIS STUDY ADDS: Night eaters with severe obesity are more likely to be low in mood and unemployed compared with non-night eaters. Night eaters with severe obesity describe compulsive and uncontrolled eating. Research interest in night-eating syndrome (NES) has grown in recent years in line with increased rates of obesity. This study used a mixed-methods approach to investigate its characteristics in severe obesity. Eighty-one individuals (mean [standard deviation] age 44.6 [11.6] years, [body mass index] 50.0 [10.7] kg m(-2) ; 43% men) from a hospital-based UK obesity clinic were interviewed for NES based on 2003 criteria. Full and partial NES were combined into one night-eating behaviour (NEB) group (n = 31). Demographic and clinical characteristics were compared with those of non-NEB individuals (n = 50). NEB characteristics were also identified through exploratory thematic analysis of interview data. NEB individuals had lower mood (P = 0.01) and were less likely to be employed (P = 0.03). Differences in mean age and reported sleep duration were not significant. Thematic analysis of patient perceptions of NEB highlighted the potential heterogeneity of NEB development: NEB developed in childhood, adolescence and adulthood. Individuals reported long-standing and current sleep difficulties, negative affect and conflictful relationships. Night eating was solitary, compulsive and uncontrolled, and daytime eating patterns were chaotic. Accounts of awareness of night eating were conflicting. Severely obese night eaters are characterized by low mood and lack of employment. Further studies are required to explore behavioural and cognitive influences on night eating in severe obesity.

Night eating syndrome: implications for severe obesity.

Night eating syndrome (NES) was first identified in 1955 by Stunkard, a psychiatrist specialising in eating disorders (ED). Over the last 20 years considerable progress has been made in defining NES as a significant clinical entity in its own right and it has now been accepted for inclusion in the fifth edition of the Diagnostic and Statistical Manual of Mental Disorders (DSM-5) due for publication in 2013. NES is considered a dysfunction of circadian rhythm with a disassociation between eating and sleeping. Core criteria include a daily pattern of eating with a significantly increased intake in the evening and/or night time, as manifested by one or both of the following: at least 25% of food intake is consumed after the evening meal or at least two episodes of nocturnal eating per week. An important recent addition to core criteria includes the presence of significant distress and/or impairment in functioning. Stunkard's team recommend further investigation on the pathogenesis of NES, in particular its relationship with traumatic life events, psychiatric comorbidity, the age of onset of NES and course of NES over time. The relationship between NES and other ED also requires further clarification as night-eaters exhibit some features of other ED; previous guidance to separate NES from other ED may have hindered earlier characterisation of NES. Evidence from European and American studies suggests NES features strongly in populations with severe obesity. The complex interplay between depression, impaired sleep and obesity-related comorbidity in severely obese individuals makes understanding NES in this context even more difficult. This review examines evidence to date on the characterisation of NES and concludes by examining the applicability of current NES criteria to individuals with severe obesity.