Exclusive Human Milk Diets and the Reduction of Necrotizing Enterocolitis.

BACKGROUND: Necrotizing enterocolitis (NEC) is common in preterm infants, especially infants less than 32 weeks gestation. Mortality from NEC is 7% and occurs in 1 out of 1000 preterm infants. Studies have shown the efficacy of an exclusive milk from mother diet in decreasing rates of NEC and associated mortality. PURPOSE: To evaluate the effectiveness of an existing exclusive human milk diet (EHMD) protocol on the incidence of NEC in extremely premature infants. EHMD, for the purposes of this project is defined as breast milk of mother, with or without human milk-based fortifier. METHODS: A single-center retrospective quasi-experimental study. The sample included 201 infants born less than 32 weeks gestation, weighing less than 1250 grams, small for gestational age (SGA) and with low Apgar scores. Outcomes measured included incidences of NEC, mortality, and co-morbidities in infants pre- and postinitiation of an EHMD protocol. RESULTS: Just 4.8% of the EHMD group had a NEC diagnosis compared to 10.5% of the bovine-based (BOV) group. There was a 1% mortality rate of the EHMD group as compared to 6% in the BOV group. The EHMD group had a statistically significant greater weight gain during hospitalization as compared to infants fed BOV ( P = < .05). IMPLICATIONS FOR PRACTICE AND RESEARCH: Neonatal intensive care units should consider EHMDs for use in this infant population. Future research is needed to support dissemination of the use of EHMD as standard of practice.

Identification and Characterization of a DmoB Flavin Oxidoreductase from a Putative Two-Component DMS C-Monooxygenase.

The compound dimethyl sulfide (DMS) links terrestrial and oceanic sulfur with the atmosphere because of its volatility. Atmospheric DMS is responsible for cloud formation and radiation backscattering and has been implicated in climate control mitigation. The enzyme DMS C-monooxygenase degrades DMS and has been classified as a two-component FMNH2-dependent monooxygenase. This enzyme requires a flavin reductase B subunit to supply electrons to the monooxygenase A subunit where DMS conversion occurs. One form of the enzyme from Hyphomicrobium sulfonivorans has been isolated and characterized. In this work, a putative DMS C-monooxygenase has been identified with bioinformatics in Arthrobacter globiformis. We report the expression, purification, and characterization of the DmoB flavin reductase subunit, termed DmoB, from A. globiformis. Data support DmoB preference and optimal activity for the cosubstrates flavin mononucleotide (FMN) and NADH. FMN binds at a 1:1 stoichiometry with high affinity (K d = 1.11 µM). The reductase is able to generate product with the A subunit from H. sulfonivorans expressed in Escherichia coli, albeit at a lower turnover than the natively expressed enzyme. No static protein-protein interactions were observed under the conditions tested between the two subunits. These results provide new details in the classification of enzymes involved in the sulfur cycling pathway and emerging forms of the enzyme DMS monooxygenase.

Biliopancreatic Diversion Induces Greater Metabolic Improvement Than Roux-en-Y Gastric Bypass.

Diabetes remission is greater after biliopancreatic diversion (BPD) than Roux-en-Y gastric bypass (RYGB) surgery. We used a mixed-meal test with ingested and infused glucose tracers and the hyperinsulinemic-euglycemic clamp procedure with glucose tracer infusion to assess the effect of 20% weight loss induced by either RYGB or BPD on glucoregulation in people with obesity (ClinicalTrials.gov number: NCT03111953). The rate of appearance of ingested glucose into the circulation was much slower, and the postprandial increases in plasma glucose and insulin concentrations were markedly blunted after BPD compared to after RYGB. Insulin sensitivity, assessed as glucose disposal rate during insulin infusion, was ∼45% greater after BPD than RYGB, whereas ß cell function was not different between groups. These results demonstrate that compared with matched-percentage weight loss induced by RYGB, BPD has unique beneficial effects on glycemic control, manifested by slower postprandial glucose absorption, blunted postprandial plasma glucose and insulin excursions, and greater improvement in insulin sensitivity.

How accessible are you? A hospital-wide audit of the accessibility and professionalism of Facebook profiles.

Introduction Social media is an integral part of modern society and is increasingly being used by patients and clinical staff alike. The General Dental Council and most employers have social media guidelines to ensure employees use social media responsibly and do not bring their organisations into disrepute.Aims To establish the accessibility of Facebook profiles of the staff and students at Eastman Dental Hospital (EDH) and to assess compliance with the latest social media guidance from the General Dental Council as well as the employing trust and university.Design and setting A hospital-wide prospective audit conducted from January to March 2018 at Eastman Dental Hospital.Gold standard One-hundred percent compliance with: a) General Dental Council (GDC); b) University College London (UCL); and c) University College London Hospitals (UCLH) social media policies.Materials and methods The names of all clinical staff/students at EDH were searched for on Facebook using a dummy profile account, to simulate a member of the public. The profiles were assessed for the accessibility of personal information, affiliations to UCL/UCLH, personal views, social networking and conduct.Results A total of 219 out of 440 profiles (50%) were identified and were accessible to varying degrees. Many accessible profiles had a profile picture and personal information visible such as gender and location. No violations of patient confidentiality were seen. Overall, 6% (n = 14) and 2% (n = 4) of profiles displayed unprofessional behaviour and substance abuse, respectively.Discussion This audit highlighted Eastman Dental Hospital's compliance to local and national social media policies.Conclusion It was not possible to identify all staff/students through the Facebook search function. No serious professional misconduct was identified, however a small minority displayed unprofessional behaviour.

Roux-en-Y Gastric Bypass Surgery Has Unique Effects on Postprandial FGF21 but Not FGF19 Secretion.

Context: Fibroblast growth factor (FGF)19 and FGF21 are secreted by the intestine and liver in response to macronutrient intake. Intestinal resection and reconstruction via bariatric surgery may alter their regulation. Objective: We tested the hypothesis that weight loss induced by Roux-en-Y gastric bypass (RYGB) surgery, but not matched weight loss induced by laparoscopic adjustable gastric banding (LAGB), increases postprandial plasma FGF19 and FGF21 concentrations. Design: Glucose kinetics and plasma FGF19 and FGF21 responses to mixed meal ingestion and to glucose-insulin infusion during a hyperinsulinemic-euglycemic clamp procedure, with stable isotope tracer methods, were evaluated in 28 adults with obesity before and after 20% weight loss induced by RYGB (n = 16) or LAGB (n = 12). Results: LAGB- and RYGB-induced weight loss increased postprandial plasma FGF19 concentrations (P < 0.05). However, weight loss after RYGB, but not LAGB, increased postprandial plasma FGF21 concentrations (1875 ± 330 to 2976 ± 682 vs 2150 ± 310 and 1572 ± 265 pg/mL × 6 hours, respectively). The increase in plasma FGF21 occurred ∼2 hours after the peak in delivery of ingested glucose into systemic circulation. Glucose-insulin infusion increased plasma FGF21, but not FGF19, concentrations. The increase in plasma FGF21 during glucose-insulin infusion was greater after than before weight loss in both surgery groups without a difference between groups, whereas plasma FGF19 was not affected by either procedure. Conclusions: RYGB-induced weight loss has unique effects on postprandial FGF21 metabolism, presumably due to rapid delivery of ingested macronutrients to the small intestine and delivery of glucose to the liver.

Alterations in 3-Hydroxyisobutyrate and FGF21 Metabolism Are Associated With Protein Ingestion-Induced Insulin Resistance.

Systemic hyperaminoacidemia, induced by either intravenous amino acid infusion or protein ingestion, reduces insulin-stimulated glucose disposal. Studies of mice suggest that the valine metabolite 3-hydroxyisobutyrate (3-HIB), fibroblast growth factor 21 (FGF21), adiponectin, and nonesterified fatty acids (NEFAs) may be involved in amino acid-mediated insulin resistance. We therefore measured in 30 women the rate of glucose disposal, and plasma 3-HIB, FGF21, adiponectin, and NEFA concentrations, under basal conditions and during a hyperinsulinemic-euglycemic clamp procedure (HECP), with and without concomitant ingestion of protein (n = 15) or an amount of leucine that matched the amount of protein (n = 15). We found that during the HECP without protein or leucine ingestion, the grand mean ± SEM plasma 3-HIB concentration decreased (from 35 ± 2 to 14 ± 1 µmol/L) and the grand median [quartiles] FGF21 concentration increased (from 178 [116, 217] to 509 [340, 648] pg/mL). Ingestion of protein, but not leucine, decreased insulin-stimulated glucose disposal (P < 0.05) and prevented both the HECP-mediated decrease in 3-HIB and increase in FGF21 concentration in plasma. Neither protein nor leucine ingestion altered plasma adiponectin or NEFA concentrations. These findings suggest that 3-HIB and FGF21 might be involved in protein-mediated insulin resistance in humans.

Evaluating public involvement in research design and grant development: Using a qualitative document analysis method to analyse an award scheme for researchers.

PLAIN ENGLISH SUMMARY: The National Institute for Health Research (NIHR) Research Design Service (RDS) for Yorkshire and Humber has been running a public involvement funding scheme since 2008. This scheme awards researchers a small amount of money to help them get involvement from patients and/or the public. Involvement activities take place at the time when researchers are planning studies, and when they are completing application forms to request funding for a proposed research project. After the public involvement activities researchers are asked to write a report for the RDS describing what they did with the public involvement funding. This study analysed those reports using an approach which included members of a public involvement panel in the data analysis process. The aim of the work was to see what the views and experiences of researchers who received funding were, and what might be learned for the future of the scheme. Twenty five reports were analysed. Four main themes were identified, these described: the added value of public involvement; aspects to consider when planning and designing public involvement; different roles of public contributors; and aspects of valuing public member contributions. The group approach to analysis was successful in enabling involvement of a variety of individuals in the process. The findings of the study provide evidence of the value of public involvement during the development of applications for research funding. The results also indicate that researchers recognise the variety in potential roles for the public in research, and acknowledge how involvement adds value to studies. ABSTRACT: Background A regional Research Design Service, funded by the National Institute for Health Research, introduced a small grant in 2008, to support public involvement (often known as patient and public involvement [PPI]) activities during the development of applications for research funding. Successful applicants are requested to submit a report detailing how the grant money was used, including a description of the aims and outcomes of the public involvement activities. The purpose of this study was to analyse the content of these reports. We aimed to find out what researcher views and experiences of public involvement activities were, and what lessons might be learned. Methods We used an innovative method of data analysis, drawing on group participatory approaches, qualitative content analysis, and Framework Analysis to sort and label the content of the reports. We developed a framework of categories and sub-categories (or themes and sub-themes) from this process. Results Twenty five documents were analysed. Four main themes were identified in the data: the added value of public involvement; planning and designing involvement; the role of public members; and valuing public member contributions. Within these themes, sub-themes related to the timing of involvement (prior to the research study/intended during the research study), and also specific benefits of public involvement such as: validating ideas; ensuring appropriate outcomes; ensuring the acceptability of data collection methods/tools and advice regarding research processes. Other sub-themes related to: finding and approaching public members; timing of events; training/support; the format of sessions; setting up public involvement panels: use of public contributors in analysis and interpretation of data; and using public members to assist with dissemination and translation into practice. Conclusions The analysis of reports submitted by researchers following involvement events provides evidence of the value of public involvement during the development of applications for research funding, and details a method for involving members of the public in data analysis which could be of value to other researchers The findings of the analysis indicate recognition amongst researchers of the variety in potential roles for public members in research, and also an acknowledgement of how involvement adds value to studies.

Perilipin 5-Driven Lipid Droplet Accumulation in Skeletal Muscle Stimulates the Expression of Fibroblast Growth Factor 21.

Perilipin 5 (PLIN5) is a lipid droplet protein and is highly expressed in oxidative tissue. Expression of the PLIN5 gene is regulated by peroxisome proliferator-activated receptor-α, fasting, and exercise. However, the effect of increased muscle PLIN5 expression on whole-body energy homeostasis remains unclear. To examine this, we developed a mouse line with skeletal muscle PLIN5 overexpression (MCK-Plin5). We show that MCK-Plin5 mice have increased energy metabolism and accumulate more intramyocellular triacylglycerol but have normal glucose and insulin tolerance. MCK-Plin5 mice fed high-fat chow manifest lower expression of inflammatory markers in their liver and increased expression of "browning" factors in adipose tissue. This muscle-driven phenotype is, at least in part, mediated by myokines; the MCK-Plin5 mice have 80-fold higher FGF21 gene expression in muscle and increased serum FGF21 concentration. The increase in FGF21 occurs mainly in muscles with a predominance of fast-twitch fibers, suggesting that fiber type-specific lipid storage may be part of the mechanism conferring metabolic protection in MCK-Plin5 mice. In conclusion, upregulating the PLIN5 level in skeletal muscle drives expression of the FGF21 gene in fast-twitch fibers and is metabolically protective. These findings provide insight into the physiology of PLIN5 and the potential contribution of its upregulation during exercise.

Oral verruciform xanthoma.

Verruciform xanthoma (VX) of the oral cavity is a benign mucosal growth that often presents as a pink, yellow or grey raised plaque or papule with granular, papillary or verrucous surface morphology. Intraorally this often presents on the masticatory mucosa and extraorally often involves the skin and anogenital mucosa. There are several proposed aetiological factors and the clinical features of VX can be misleading; clinically it can resemble malignancy. Histopathological diagnosis is a key for the correct management of this lesion. Excision of this lesion is curative.

Specific minor groove solvation is a crucial determinant of DNA binding site recognition.

The DNA sequence preferences of nearly all sequence specific DNA binding proteins are influenced by the identities of bases that are not directly contacted by protein. Discrimination between non-contacted base sequences is commonly based on the differential abilities of DNA sequences to allow narrowing of the DNA minor groove. However, the factors that govern the propensity of minor groove narrowing are not completely understood. Here we show that the differential abilities of various DNA sequences to support formation of a highly ordered and stable minor groove solvation network are a key determinant of non-contacted base recognition by a sequence-specific binding protein. In addition, disrupting the solvent network in the non-contacted region of the binding site alters the protein's ability to recognize contacted base sequences at positions 5-6 bases away. This observation suggests that DNA solvent interactions link contacted and non-contacted base recognition by the protein.

Imaging of neutral lipids and neutral lipid associated proteins.

Intracellular fat droplets are large and have a distinct morphology, which makes their imaging at the light level simple and informative. We detail how to image the fat droplet core by metabolic labeling with fluorescent fatty acids or lipophilic fluorochromes. Further, we describe the use of indirect immunostaining to image fat droplet proteins and fat cores in the same field. We also address the use of appropriate controls for determining signal specificity and other practical considerations for optimizing image quality.

Perilipin 1 moves between the fat droplet and the endoplasmic reticulum.

Perilipin 1, unlike the other perilipins, is thought to be restricted to the fat droplet. We reassessed its cellular distribution using the fat droplet marker CGI-58 in OP9 and 3T3-L1 adipocyte lines and in brown adipose tissue (BAT). As expected, we found perilipin 1 in the fat droplet-enriched floating fraction from centrifuged adipocyte or BAT homogenates. However, about half of perilipin 1 was suspended in the cytosol/infranate or pelleted with cellular membranes. In these fractionations, most of the fat droplet-associated protein CGI-58 was in the floating fraction. In BAT and OP9 adipocytes about a third of perilipin 1 pellets, compared with a much smaller fraction of CGI-58. Co-imaging perilipin 1 and smooth endoplasmic reticulum (ER) markers reveals both ER and fat droplet associated perilipin 1 in OP9 adipocytes. Consistent with these observations, perilipin 1 overexpressed in COS7 cells mostly fractionates with cellular membranes and imaging shows it on the ER. In 3T3-L1 adipocytes almost half of perilipin 1 floats, half is suspended as infranate and small amounts pellet. Finally, driving rapid fat droplet synthesis in OP9 adipocytes increases the intensity of perilipin 1 on fat droplets, while decreasing non-fat droplet immunolabeling. Confirming the morphological findings, fractionation shows perilipin 1 moving from the pelleted to the floated fractions. In conclusion, this study documents an expanded intracellular distribution for perilipin 1 and its movement from ER to fat droplet during lipid synthesis.

Indirect readout of DNA sequence by p22 repressor: roles of DNA and protein functional groups in modulating DNA conformation.

The repressor of bacteriophage P22 (P22R) discriminates between its various DNA binding sites by sensing the identity of non-contacted base pairs at the center of its binding site. The "indirect readout" of these non-contacted bases is apparently based on DNA's sequence-dependent conformational preferences. The structures of P22R-DNA complexes indicate that the non-contacted base pairs at the center of the binding site are in the B' state. This finding suggests that indirect readout and therefore binding site discrimination depend on P22R's ability to either sense and/or impose the B' state on the non-contacted bases of its binding sites. We show here that the affinity of binding sites for P22R depends on the tendency of the central bases to assume the B'-DNA state. Furthermore, we identify functional groups in the minor groove of the non-contacted bases as the essential modulators of indirect readout by P22R. In P22R-DNA complexes, the negatively charged E44 and E48 residues are provocatively positioned near the negatively charged DNA phosphates of the non-contacted nucleotides. The close proximity of the negatively charged groups on protein and DNA suggests that electrostatics may play a key role in the indirect readout process. Changing either of two negatively charged residues to uncharged residues eliminates the ability of P22R to impose structural changes on DNA and to recognize non-contacted base sequence. These findings suggest that these negatively charged amino acids function to force the P22R-bound DNA into the B' state and therefore play a key role in indirect readout by P22R.

A single centrifugation method for isolating fat droplets from cells and tissues.

Fat droplets (FDs) have important roles in cellular energy regulation. Isolating FDs from either cells or tissue continues to be important for studying these organelles. Here, we describe a procedure wherein whole homogenates of cultured cells or tissue are fractionated with a single centrifugation step in a standard microcentrifuge. This procedure reproducibly yields three fractions highly enriched in either FDs, soluble cellular components, or sedimentable organelles/membranes.

Effect of peroxides on [3H]D-aspartate release from bovine isolated retinae.

In the present study, we investigated the effect of naturally occurring and synthetic peroxides on K+-depolarization-evoked release of [3H]D-aspartate from bovine isolated retinae. Furthermore, effect of peroxides on endogenous glutamate concentrations were measured by HPLC in bovine neural retinae and vitreous humor of eyes treated with hydrogen peroxide (H2O2) ex vivo. Both naturally occurring H2O2 (1-100 microM) and synthetic (cumene hydroperoxide, cuOOH; 1-100 microM) peroxides caused a concentration-dependent inhibition of K+-evoked [3H]D-aspartate release without affecting basal tritium efflux. The antioxidant, trolox (2 mM) prevented the inhibition of evoked [3H]D-aspartate overflow elicited by both H2O2 (30 microM) and cuOOH (10 microM). Inhibition of catalase by 3-amino-triazole (3- AT 100 mM) enhanced an inhibitory effect of a low concentration of H2O2 (1 microM) but antagonized the effect of H2O2 (30 microM) on K+-induced [3H]D-aspartate release. In ex vivo experiments, exogenously applied H2O2 (1-100 microM) also caused a concentration-related decrease in glutamate levels in the bovine retina. We conclude that peroxides can inhibit K+-evoked release of [3H]D-aspartate and also decrease endogenous glutamate concentrations in the bovine retina.

Glucose-deprivation-induced [3H]D-aspartate release from isolated bovine and human retinae.

The glucose deprivation-induced release of [3H]D-aspartate was studied in bovine and human retinas in a superfusion apparatus. [3H]D-aspartate release was significantly increased upon omitting glucose in the superfusion buffer. This effect was dependent on external Ca2+ because L- and N-type Ca2+-channel blockers, such as diltiazem (1 microM), nitrendipine (1 microM), and omega-conotoxin (100 nM), significantly reduced the effect of glucose-deprivation induced release of [3H]D-aspartate. Furthermore, while glutamate receptor agonists (L-glutamate, N-methyl-D-aspartate, but not kainate) potentiated the effects of glucose deprivation, antagonists (MK-801, MCPG, ifenprodil, and L-AP3) at these receptors blocked the glucose deprivation-induced release process. Taken together, these studies have demonstrated that under conditions of glucose deprivation, as may happen during ischemic events in vivo, the retinal glutamatergic nerve endings and/or glial cells promote the efflux of [3H]D-aspartate into the extracellular environment. This process appears to be receptor-mediated and dependent on extracellular Ca2+ and is similar to previous reports pertaining to brain tissues.

Pharmacology of serotonin receptors modulating electrically-induced [3h]-norepinephrine release from isolated mammalian iris-ciliary bodies.

The pharmacology of prejunctional serotonin (5-HT) heteroreceptors that regulate the release of norepinephrine (NE) was studied in isolated bovine and human iris-ciliary bodies. The effect of exogenous 5-HT and various 5-HT receptor agonists was examined on the release of [3H]-norepinephrine ([3H]NE). Both 5-HT and m-chlorophenyl-biguanide (m-CPBG) caused enhancement in the field-stimulated release of [3H]NE from bovine tissues whereas 5-carboxamidotryptamine (5-CT) had no such effect. On the other hand, 8hydroxy-dipropylaminotetralin (8-OH-DPAT), caused a significant dose-related inhibition of evoked [3H]NE release. In human iris-ciliary bodies, 5-HT caused an inhibitory response on electrically-evoked [3H]NE release at low concentrations but produced an excitatory action at concentrations greater than 3 microM. To further confirm the nature of the prejunctional 5-HT heteroreceptors regulating [3H]NE release, effects of 5-HT3, 5-HT6 and 5-HT7 receptor antagonists were examined on a standard response to 5-HT. All antagonists examined caused a concentration-dependent inhibition of the response elicited by the standard 5-HT-induced response with the following rank order of potency (as measured by IC30 values): MDL-72222 >> SB-258719 > RO-04-690. We conclude that the excitatory prejunctional 5-HT heteroreceptors present in bovine iris-ciliary bodies belong to the 5-HT3 receptor subtype.