Rational design of a combination medication for the treatment of obesity

Existing obesity therapies are limited by safety concerns and modest efficacy reflecting a weight loss plateau. Here, we explore combination therapy with bupropion (BUP), a putative stimulator of melanocortin pathways, and an opioid antagonist, naltrexone (NAL), to antagonize an inhibitory feedback loop that limits sustained weight reduction. In vitro electrophysiologic experiments were conducted to determine the extent to which BUP+NAL stimulated hypothalamic pro-opiomelanocortin (POMC) neurons in mouse brain. A subsequent study further characterized the effect of combination BUP+NAL treatment on food intake in lean and obese mice. Finally, a randomized, blinded, placebo-controlled trial in obese adult subjects was conducted. Randomization included: BUP (300 mg) + NAL (50 mg), BUP (300 mg) + placebo (P), NAL (50 mg) + P or P+P for up to 24 weeks. BUP+NAL stimulated murine POMC neurons in vitro and caused a greater reduction in acute food intake than either monotherapy, an effect consistent with synergism. Combined BUP+NAL provided sustained weight loss without evidence of an efficacy plateau through 24 weeks of treatment. BUP+NAL completers diverged from NAL+P (P < 0.01) and P+P (P < 0.001) at week 16 and from BUP+P by week 24 (P < 0.05). The combination was also well tolerated. Translational studies indicated that BUP+NAL therapy produced synergistic weight loss which exceeded either BUP or NAL alone. These results supported the hypothesis that NAL, through blockade of beta-endorphin mediated POMC autoinhibition, prevents the classic weight loss plateau observed with monotherapies such as BUP. This novel treatment approach (BUP+NAL) holds promise for the treatment of obesity.\     

Social Structure Predicts Genital Morphology in African Mole-Rats

Background

African mole-rats (Bathyergidae, Rodentia) exhibit a wide range of social structures, from solitary to eusocial. We previously found a lack of sex differences in the external genitalia and morphology of the perineal muscles associated with the phallus in the eusocial naked mole-rat. This was quite surprising, as the external genitalia and perineal muscles are sexually dimorphic in all other mammals examined. We hypothesized that the lack of sex differences in naked mole-rats might be related to their unusual social structure.

Methodology/Principal Findings

We compared the genitalia and perineal muscles in three African mole-rat species: the naked mole-rat, the solitary silvery mole-rat, and the Damaraland mole-rat, a species considered to be eusocial, but with less reproductive skew than naked mole-rats. Our findings support a relationship between social structure, mating system, and sexual differentiation. Naked mole-rats lack sex differences in genitalia and perineal morphology, silvery mole-rats exhibit sex differences, and Damaraland mole-rats are intermediate.

Conclusions/Significance

The lack of sex differences in naked mole-rats is not an attribute of all African mole-rats, but appears to have evolved in relation to their unusual social structure and reproductive biology.     

Genetic Variation in the Familial Mediterranean Fever Gene (MEFV) and Risk for Crohn's Disease and Ulcerative Colitis

Background and Aims

The familial Mediterranean fever (FMF) gene (MEFV) encodes pyrin, a major regulator of the inflammasome platform controlling caspase-1 activation and IL-1β processing. Pyrin has been shown to interact with the gene product of NLRP3, NALP3/cryopyrin, also an important active member of the inflammasome. The NLRP3 region was recently reported to be associated with Crohn''s disease (CD) susceptibility. We therefore sought to evaluate MEFV as an inflammatory bowel disease (IBD) susceptibility gene.

Methodology and Results

MEFV colonic mucosal gene expression was significantly increased in experimental colitis mice models (TNBS p<0.0003; DSS p<0.006), in biopsies from CD (p<0.02) and severe ulcerative colitis (UC) patients (p<0.008). Comprehensive genetic screening of the MEFV region in the Belgian exploratory sample set (440 CD trios, 137 UC trios, 239 CD cases, 96 UC cases, and 107 healthy controls) identified SNPs located in the MEFV 5′ haplotype block that were significantly associated with UC (rs224217; p = 0.003; A allele frequency: 56% cases, 45% controls), while no CD associations were observed. Sequencing and subsequent genotyping of variants located in this associated haplotype block identified three synonymous variants (D102D/rs224225, G138G/rs224224, A165A/rs224223) and one non-synonymous variant (R202Q/rs224222) located in MEFV exon 2 that were significantly associated with UC (rs224222: p = 0.0005; A allele frequency: 32% in cases, 23% in controls). No consistent associations were observed in additional Canadian (256 CD trios, 91 UC trios) and Scottish (495 UC, 370 controls) sample sets. We note that rs224222 showed marginal association (p = 0.012; G allele frequency: 82% in cases, 70% in controls) in the Canadian sample, but with a different risk allele. None of the NLRP3 common variants were associated with UC in the Belgian-Canadian UC samples and no significant interactions were observed between NLRP3 and MEFV that could explain the observed flip-flop of the rs224222 risk allele.

Conclusion

The differences in association levels observed between the sample sets may be a consequence of distinct founder effects or of the relative small sample size of the cohorts evaluated in this study. However, the results suggest that common variants in the MEFV region do not contribute to CD and UC susceptibility.     

The CYPome (Cytochrome P450 complement) of Aspergillus nidulans

The cytochromes P450 (CYPs) are found in all biological kingdoms and genome sequencing projects continue to reveal an ever increasing number. The principle aim of this paper is to identify the complete CYPome of Aspergillus nidulans from the genome sequence version AN.3 deposited at the Broad institute, assign the appropriate CYP nomenclature and define function where possible. The completed analysis revealed a total of 111 CYP genes, 3 of which were previously unknown and 8 pseudogenes, representing 89CYP families, 21 of which are unique. We have identified 28 potential gene clusters associated with one or more CYP genes and discussed those with putative PKS and NRPS associated function. The chromosomal location of the genes, predicted cellular location of the proteins and possible function(s) are discussed.     

A Novel Role for MAPKAPK2 in Morphogenesis during Zebrafish Development

One of the earliest morphogenetic processes in the development of many animals is epiboly. In the zebrafish, epiboly ensues when the animally localized blastoderm cells spread, thin over, and enclose the vegetally localized yolk. Only a few factors are known to function in this fundamental process. We identified a maternal-effect mutant, betty boop (bbp), which displays a novel defect in epiboly, wherein the blastoderm margin constricts dramatically, precisely when half of the yolk cell is covered by the blastoderm, causing the yolk cell to burst. Whole-blastoderm transplants and mRNA microinjection rescue demonstrate that Bbp functions in the yolk cell to regulate epiboly. We positionally cloned the maternal-effect bbp mutant gene and identified it as the zebrafish homolog of the serine-threonine kinase Mitogen Activated Protein Kinase Activated Protein Kinase 2, or MAPKAPK2, which was not previously known to function in embryonic development. We show that the regulation of MAPKAPK2 is conserved and p38 MAP kinase functions upstream of MAPKAPK2 in regulating epiboly in the zebrafish embryo. Dramatic alterations in calcium dynamics, together with the massive marginal constrictive force observed in bbp mutants, indicate precocious constriction of an F-actin network within the yolk cell, which first forms at 50% epiboly and regulates epiboly progression. We show that MAPKAPK2 activity and its regulator p38 MAPK function in the yolk cell to regulate the process of epiboly, identifying a new pathway regulating this cell movement process. We postulate that a p38 MAPKAPK2 kinase cascade modulates the activity of F-actin at the yolk cell margin circumference allowing the gradual closure of the blastopore as epiboly progresses.     

Disease-Causing 7.4 kb Cis-Regulatory Deletion Disrupting Conserved Non-Coding Sequences and Their Interaction with the FOXL2 Promotor: Implications for Mutation Screening

To date, the contribution of disrupted potentially cis-regulatory conserved non-coding sequences (CNCs) to human disease is most likely underestimated, as no systematic screens for putative deleterious variations in CNCs have been conducted. As a model for monogenic disease we studied the involvement of genetic changes of CNCs in the cis-regulatory domain of FOXL2 in blepharophimosis syndrome (BPES). Fifty-seven molecularly unsolved BPES patients underwent high-resolution copy number screening and targeted sequencing of CNCs. Apart from three larger distant deletions, a de novo deletion as small as 7.4 kb was found at 283 kb 5′ to FOXL2. The deletion appeared to be triggered by an H-DNA-induced double-stranded break (DSB). In addition, it disrupts a novel long non-coding RNA (ncRNA) PISRT1 and 8 CNCs. The regulatory potential of the deleted CNCs was substantiated by in vitro luciferase assays. Interestingly, Chromosome Conformation Capture (3C) of a 625 kb region surrounding FOXL2 in expressing cellular systems revealed physical interactions of three upstream fragments and the FOXL2 core promoter. Importantly, one of these contains the 7.4 kb deleted fragment. Overall, this study revealed the smallest distant deletion causing monogenic disease and impacts upon the concept of mutation screening in human disease and developmental disorders in particular.     

A Neutralizing Human Monoclonal Antibody Protects against Lethal Disease in a New Ferret Model of Acute Nipah Virus Infection

Nipah virus is a broadly tropic and highly pathogenic zoonotic paramyxovirus in the genus Henipavirus whose natural reservoirs are several species of Pteropus fruit bats. Nipah virus has repeatedly caused outbreaks over the past decade associated with a severe and often fatal disease in humans and animals. Here, a new ferret model of Nipah virus pathogenesis is described where both respiratory and neurological disease are present in infected animals. Severe disease occurs with viral doses as low as 500 TCID₅₀ within 6 to 10 days following infection. The underlying pathology seen in the ferret closely resembles that seen in Nipah virus infected humans, characterized as a widespread multisystemic vasculitis, with virus replicating in highly vascular tissues including lung, spleen and brain, with recoverable virus from a variety of tissues. Using this ferret model a cross-reactive neutralizing human monoclonal antibody, m102.4, targeting the henipavirus G glycoprotein was evaluated in vivo as a potential therapeutic agent. All ferrets that received m102.4 ten hours following a high dose oral-nasal Nipah virus challenge were protected from disease while all controls died. This study is the first successful post-exposure passive antibody therapy for Nipah virus using a human monoclonal antibody.     

Understanding reporting bias in the dietary recall data of 11-year-old girls

Objective: This study describes patterns of bias in self‐reported dietary recall data of girls by examining differences among girls classified as under‐reporters, plausible reporters, and over‐reporters on weight, dietary patterns, and psychosocial characteristics. Research Methods and Procedures: Participants included 176 girls at age 11 and their parents. Girls’ weight and height were measured. Three 24‐hour dietary recalls and responses to psychosocial measures were collected. Plausibility cut‐offs for reported energy intake as a percentage of predicted energy requirements were used to divide the sample into under‐reporters, plausible reporters, and over‐reporters. Differences among these three groups on dietary and psychosocial variables were assessed to examine possible sources of bias in reporting. Results: Using a ±1 standard deviation cut‐off for energy intake plausibility, 50% of the sample was categorized as plausible reporters, 34% as under‐reporters, and 16% as over‐reporters. Weight status of under‐reporters was significantly higher than that of plausible reporters and over‐reporters. With respect to reported dietary intake, under‐reporters were no different from plausible reporters on intakes of foods with higher nutrient densities and lower energy densities and were significantly lower than plausible reporters on intakes of foods with lower nutrient densities and higher energy densities. Over‐reporters reported significantly higher intakes of all food groups and the majority of subgroups, relative to plausible reporters. Under‐reporters had significantly higher levels of weight concern and dietary restraint than both plausible reporters and over‐reporters. Discussion: Techniques to categorize plausible and implausible reporters can and should be used to provide an improved understanding of the nature of error in children's dietary intake data and account for this error in analysis and interpretation.