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Background It has been proposed that the use of gene expression microarrays in nonrecombinant parental or congenic strains can accelerate the process of isolating individual genes underlying quantitative trait loci (QTL). However, the effectiveness of this approach has not been assessed. Results Thirty-seven studies that have implemented the QTL/microarray approach in rodents were reviewed. About 30% of studies showed enrichment for QTL candidates, mostly in comparisons between congenic and background strains. Three studies led to the identification of an underlying QTL gene. To complement the literature results, a microarray experiment was performed using three mouse congenic strains isolating the effects of at least 25 biometric QTL. Results show that genes in the congenic donor regions were preferentially selected. However, within donor regions, the distribution of differentially expressed genes was homogeneous once gene density was accounted for. Genes within identical-by-descent (IBD) regions were less likely to be differentially expressed in chromosome 2, but not in chromosomes 11 and 17. Furthermore, expression of QTL regulated in cis (cis eQTL) showed higher expression in the background genotype, which was partially explained by the presence of single nucleotide polymorphisms (SNP). Conclusions The literature shows limited successes from the QTL/microarray approach to identify QTL genes. Our own results from microarray profiling of three congenic strains revealed a strong tendency to select cis-eQTL over trans-eQTL. IBD regions had little effect on rate of differential expression, and we provide several reasons why IBD should not be used to discard eQTL candidates. In addition, mismatch probes produced false cis-eQTL that could not be completely removed with the current strains genotypes and low probe density microarrays. The reviewed studies did not account for lack of coverage from the platforms used and therefore removed genes that were not tested. Together, our results explain the tendency to report QTL candidates as differentially expressed and indicate that the utility of the QTL/microarray as currently implemented is limited. Alternatives are proposed that make use of microarray data from multiple experiments to overcome the outlined limitations.

Mitochondrial hyperpolarization inhibits the electron transport chain and increases incomplete reduction of oxygen, enabling production of reactive oxygen species (ROS). The consequence is mitochondrial damage that eventually causes cell death. Uncoupling proteins (UCPs) are inner mitochondrial membrane proteins that dissipate the mitochondrial proton gradient by transporting H+across the inner membrane, thereby stabilizing the inner mitochondrial membrane potential and reducing the formation of ROS. The role of UCP2 in neuroprotection is still in debate. This study seeks to clarify the role of UCP2 in transient focal ischemia (tFI) and to further understand the mechanisms of ischemic brain damage. Both wild-type and UCP2-knockout mice were subjected to tFI. Knocking out UCP2 significantly increased the infarct volume to 61% per hemisphere as compared with 18% in wild-type animals. Knocking out UCP2 suppressed antioxidant, cell-cycle, and DNA repair genes, including Sod1 and Sod2, Gstm1, and cyclins. Furthermore, knocking out UCP2 significantly upregulated the protein levels of the inflammatory cytokines, including CTACK, CXCL16, Eotaxin-2, fractalkine, and BLC. It is concluded that knocking out the UCP2 gene exacerbates neuronal death after cerebral ischemia with reperfusion and this detrimental effect is mediated by alteration of antioxidant genes and upregulation of inflammatory mediators.

We have previously shown that 90% of outbred obese Zucker Leprfa/farats die prematurely of renal disease. Thus, renal disease in obese Zucker Leprfa/farats may be caused by the LEPR mutation on chromosome 5, by the obesity, or it may be influenced by Zucker susceptibility alleles of genes on other chromosomes. We have searched for susceptibility genes on other chromosomes using urinary albumin excretion (UAE) as an early indicator of altered renal function in a backcross of (Brown Norway × inbred Zucker) F1 × inbred Zucker, which we name the BZZ cross. We killed 237 BZZ backcross animals at 15 wk of age. All included animals were homozygous for the fatty mutation of LEPR and were obese. Urinary creatinine measurements were used to calculate the albumin-to-creatinine ratio (ACR). We identified direct effect quantitative trait loci (QTLs) for UAE and ACR on chromosome 1 (LOD scores = 3.6 and 2.86, respectively) in males, and chromosome 4 (LOD score = 2.9) in females. Significant QTLs were identified for left kidney weight for females on chromosomes 3 and 12. We also demonstrated that kidneys from 15 wk old obese inbred Zucker rats already show evidence of kidney pathology: tubular dilation, proteinaceous fluid accumulation, evidence for inflammation, and mild mesangial and tubular membrane basement membrane thickening. Both lean Zucker rats and the Brown Norway rats showed no evidence for these changes. Thus, by removing the influence of the Leprfa/famutation from analysis we have identified UAE QTLs unlinked to LEPR.

Inbred strains of mice exhibit large genetic variations in hepatic 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase activity. A tissue-specific genetic variation between the strains BALB/c and C57BL/6, resulting in about 5-fold higher levels in hepatic reductase activity in strain C57BL/6, was examined in detail. The activity difference between these two strains could be explained entirely by differences in hepatic reductase mRNA levels. In genetic crosses, the variation segregated as a single major Mendelian element. Surprisingly, the mode of inheritance was recessive since F1 mice exhibited the BALB/c levels of enzyme activity. Despite the fact that the rates of hepatic sterol synthesis also differed between the strains by a factor of about five, the altered hepatic reductase expression did not significantly influence plasma lipoprotein levels. The response to a high cholesterol, high fat diet between the strains was remarkably different. Thus, in BALB/c mice, both hepatic reductase activity and mRNA levels were affected only slightly, if at all, by cholesterol feeding, while in strain C57BL/6 mice both were reduced more than 10-fold by cholesterol feeding. Several lines of evidence, including analysis of cis-acting regulatory elements, the nonadditive mode of inheritance, and genetic studies of the HMG-CoA reductase gene locus on mouse chromosome 13, support the possibility that the variation in reductase expression is not due to a mutation of the structural gene but, rather, is determined by a trans-acting factor controlling reductase mRNA levels. The variation provides a striking example, at the molecular level, of the importance of dietary-genetic interactions in the control of cholesterol metabolism.

This article presents biochemical data on the BSB mouse model of multigenic obesity indicating increased percentage body fat, increased fasting plasma insulin, and increased insulin resistance in male and female obese mice compared with lean controls. Plasma glucose was significantly increased only in male obese mice. Morphological and morphometrical analyses of pancreatic islets showed increased islet size and number in all obese mice compared with lean controls. Immuno-staining results for insulin-positive islet cells showed greater levels of insulin in male and female obese versus lean mice, while the percent or proportion of insulin immuno-staining, as expected, was not significantly different between obese and lean. The percent or proportion of immuno-staining for islet glucagon and somatostatin showed reduced staining in islets from obese compared with lean mice. The significance of these findings shows, for the first time, the morphologic appearance of pancreatic islets and the quantitative distribution of the three major islet cell hormonal populations in BSB obese mice. The correlation between this descriptive information and physiological data might lend insights to the cause of obesity-related diabetes.

Gastrointestinal bleeding is a common occurrence in patients with cancer and is a frequent indicator of a gastrointestinal malignancy. Rapid evaluation and treatment is key for the hemodynamically unstable patient. Endoscopy remains the cornerstone of diagnosis and management for cancer patients with gastrointestinal bleeding. The emergency physician should also be aware of other diagnostic and treatment modalities that may be needed to take care of these patients.

The anorexigenic neuromodulator alpha-melanocyte-stimulating hormone (alpha-MSH; referred to here as alpha-MSH1-13) undergoes extensive posttranslational processing, and its in vivo activity is short lived due to rapid inactivation. The enzymatic control of alpha-MSH1-13 maturation and inactivation is incompletely understood. Here we have provided insight into alpha-MSH1-13 inactivation through the generation and analysis of a subcongenic mouse strain with reduced body fat compared with controls. Using positional cloning, we identified a maximum of 6 coding genes, including that encoding prolylcarboxypeptidase (PRCP), in the donor region. Real-time PCR revealed a marked genotype effect on Prcp mRNA expression in brain tissue. Biochemical studies using recombinant PRCP demonstrated that PRCP removes the C-terminal amino acid of alpha-MSH1-13, producing alpha-MSH1-12, which is not neuroactive. We found that Prcp was expressed in the hypothalamus in neuronal populations that send efferents to areas where alpha-MSH1-13 is released from axon terminals. The inhibition of PRCP activity by small molecule protease inhibitors administered peripherally or centrally decreased food intake in both wild-type and obese mice. Furthermore, Prcp-null mice had elevated levels of alpha-MSH1-13 in the hypothalamus and were leaner and shorter than the wild-type controls on a regular chow diet; they were also resistant to high-fat diet-induced obesity. Our results suggest that PRCP is an important component of melanocortin signaling and weight maintenance via control of active alpha-MSH1-13 levels.

The workshop reviewed the literature indicating that natural alleles influence a substantial percentage of responses to nutrition and exercise in both humans and animal models. Human genetic studies provide evidence that body weight response to over- and underfeeding and to exercise is associated with specific genes. Studies in animal models, primarily rodents, prove the genetic control of responsiveness to diet and exercise and provide the tools to examine specific mechanisms. Limitations of the animal literature include lack of studies of allelic contributions to weight loss in response to diet restriction and data on evidence-based diets. Discussion of the relative merits of sample size constraints vs. precision of phenotype measures in human genetic studies concluded that imprecise measures such as body weight and body mass index identify different genes than will specific measures of fat mass. Validation and limitations of whole genome association studies in humans was discussed, as was the role of animal models in discovery and mechanistic studies of gene/nutrition/exercise interactions. The workshop concluded that genetics has a substantial impact on responses to both diet and exercise. However, current knowledge does not allow individual diet and exercise recommendations. New resources and technologies, including cost-effective phenotyping for humans and whole genome sequencing in both humans and rodents, are needed.

Mitochondrial proton leak may modulate reactive oxygen species (ROS) production and play a role in aging. The purpose of this study was to determine proton leak across the life span in skeletal mitochondria from calorie-restricted and UCP2/3 overexpressing mice. Proton leak in isolated mitochondria and markers of oxidative stress in whole tissue were measured in female C57BL/6J mice fed ad-libitum (WT-Control) or a 30% calorie-restricted (WT-CR) diet, and in mice overexpressing UCP2 and UCP3 (Positive-TG), their non-overexpressing littermates (Negative-TG) and UCP3 knockout mice (UCP3KO). Proton leak in WT-CR mice was lower than that of control mice at 8 and 26 months of age. The Positive-TG mice had greater proton leak than the Negative-TG and UCP3KO mice at 8 months of age, but this difference disappeared by 19 and 26 months. Lipid peroxidation was generally lower in WT-CR vs. WT-Control mice and UCP3KO mice had greater concentrations of T-BARS (thiobarbituric acid reactive substances, a measure of lipid peroxidation) than did Positive-TG and Negative-TG. The results of this study indicate that sustained increases in muscle mitochondrial proton leak are not responsible for alterations in life span with calorie restriction or UCP3 overexpression in mice. However, UCP3 may contribute to the actions of CR through mechanisms distinct from increasing basal proton leak.

OBJECTIVE: The emergency physician should be familiar with the wide spectrum of pediatric mental health emergencies because they are commonly encountered in emergency medical practice. METHODS: A review of the literature was done in order to develop an approach for dealing with children presenting with mental health disorders in the emergency department (ED). RESULTS: Children' mental health emergencies have a wide spectrum from behavioral disturbances to major depression. An approach to the issues involved in caring for these patients is discussed which acknowledges the essential role of the emergency physician and the importance of integrating ED care with multidisciplinary services. CONCLUSIONS: The actions and directions taken in the ED are a crucial part of the child's long-term care and treatment. The ED evaluation and management of pediatric mental health emergencies may vary depending on the complaint and includes differentiation from organic etiologies, medical stabilization, and occasionally in depth psychosocial interview. Language: en

High density lipoproteins (HDL) are heterogeneous particles consisting of about equal amounts of lipid and protein that are thought to mediate the transport of cholesterol from peripheral tissues to liver. We show that a previously identified polymorphism affecting HDL electrophoretic mobility in mice is due to a monogenic variation controlling HDL size and apolipoprotein composition. Thus, the HDL particles of various inbred strains of mice exhibit a striking difference in the ratio fo the two major apolipoproteins of HDL, apoA-I and apoA-II. HDL particles in all strains examined contain an average of about five apoA-I molecules; however, whereas the strains with small HDL contain two to three apoA-II molecules per particle, the strains with large HDL contain about five apoA-II molecules per particle. This increase in the protein content of the large HDL is also accompanied by increased lipid content. The HDL size polymorphism and apoA-II levels cosegregate with the apoA-II structural gene on mouse chromosome 1, indicating that a mutation of the apoA-II gene locus is responsible. The rates of synthesis of apoA-II are increased in the strains with large HDL and high apoA-II levels as compared to the strains with small HDL and low apoA-II levels. On the other hand, the fractional catabolic rates of both apoA-I and apoA-II among the strains are very similar, confirming that apoA-II concentrations are controlled at the level of synthesis. Despite the difference in rates of apoA-II synthesis between strains, the apoA-II mRNA levels in the strains are not discernibly different, suggesting that a mutation of the apoA-II structural gene controls apoA-II translational efficiency. This was confirmed by translating apoA-II mRNA in vitro using a rabbit reticulocyte lysate system. Sequencing of apoA-II cDNA from the strains revealed a number of nucleotide substitutions, which may affect translational efficiency. We conclude that the assembly of apoA-II into HDL does not have a set stoichiometry but, rather, is controlled by the production of apoA-II. As apoA-II levels increase, the HDL particles become larger and acquire more lipid, but apoA-I content per particle remains unchanged. These studies with mice provide a model for the metabolic relationships between apoA-I, apoA-II, and HDL lipid in humans.

Animal models are invaluable resources for biomedical research, including research on the effects of diet on metabolism and disease. Usually, great care is taken to ensure comparable genetic backgrounds and environmental conditions when performing studies using animal models, since this minimizes introduction of variability that can confound detection of treatment-related phenotypic differences. However, many papers using animal models draw conclusions about dietary effects from comparisons of natural-ingredient chow with defined diets, despite marked differences in micro- and macronutrient content. When comparing the effects of a chow diet with a defined high-fat diet, the effects of the dietary fat will be confounded with the effects of other components in the diets. This issue is highlighted by a limited literature survey that was conducted to identify common problems in the use and reporting of rodent diets.All original research papers identified by the keywords “mouse high fat” published in 2007 in five high-impact journals were included in this evaluation. Of the 35 papers examined, only 14% (5 papers) compared diets using identical nutrients differing only in relative amounts of fat and carbohydrate (Figure 1Figure 1). Specific details regarding the dietary comparisons made were often lacking, and frequently conclusions were drawn from comparisons of defined high-fat diets to chow.Figure 1Diet Comparisons in Recent Research PapersPie chart showing the percentage of 35 original research papers evaluated that used appropriate diet comparisons (14%), that compared chow and defined high-fat diets (43%), and that presented insufficient information to evaluate diet comparisons (34%). In the “chow and defined” category (9%), both diet types were used but no direct comparison was made between them. The journals examined were Cell Metabolism (7 papers), Diabetes (the first 11 of 36 papers), The Journal of Clinical Investigation (12 papers), Nature (2 papers), and Nature Medicine (3 papers).View Large Image | View Hi-Res Image | Download PowerPoint SlideRegular chow is composed of agricultural byproducts such as ground wheat, corn, or oats, alfalfa and soybean meal, a protein source such as fish, and vegetable oil and is supplemented with minerals and vitamins. Thus, chow is a high-fiber diet containing complex carbohydrates, with fats from a variety of vegetable sources. Chow is inexpensive to manufacture and is palatable to rodents. In contrast, defined high-fat diets consist of amino acid-supplemented casein, corn starch, maltodextrose or sucrose, and soybean oil or lard, also supplemented with minerals and vitamins. Fiber is often provided by cellulose. Chow and defined diets may exert significant separate and independent unintended effects on the measured phenotypes in any research protocol.Two important differences between chow and defined diets are the phytoestrogen content from soy, which is high but variable in chow diets but absent from defined diets (reviewed in Thigpen et al., 2004xThigpen, J.E., Setchell, K.D., Saunders, H.E., Haseman, J.K., Grant, M.G., and Forsythe, D.B. ILAR J. 2004; 45: 401–416Crossref | PubMedSee all ReferencesThigpen et al., 2004), and from sucrose, which is used as a carbohydrate source in defined diets but is absent from chow. Dietary phytoestrogens influence food and water intake, anxiety-related behaviors, locomotor activity, learning and memory, fat deposition, blood insulin, leptin and thyroid levels, and lipogenesis and lipolysis in isolated rat adipocytes (Torre-Villalvazo et al., 2008xTorre-Villalvazo, I., Tovar, A.R., Ramos-Barragan, V.E., Cerbon-Cervantes, M.A., and Torres, N. J. Nutr. 2008; 138: 462–468PubMedSee all References, Lephart et al., 2004axLephart, E.D., Porter, J.P., Lund, T.D., Bu, L., Setchell, K.D., Ramoz, G., and Crowley, W.R. Nutr. Metab. (Lond). 2004; 1: 16Crossref | PubMed | Scopus (34)See all References; reviewed in Lephart et al., 2004bxLephart, E.D., Setchell, K.D., Handa, R.J., and Lund, T.D. ILAR J. 2004; 45: 443–454Crossref | PubMedSee all ReferencesLephart et al., 2004b). Sucrose is 50% fructose, and there is abundant evidence that fructose can exacerbate weight gain and contribute to insulin resistance and dyslipidemia (reviewed in Stanhope and Havel, 2008xStanhope, K.L. and Havel, P.J. Curr. Opin. Lipidol. 2008; 19: 16–24Crossref | PubMed | Scopus (121)See all ReferencesStanhope and Havel, 2008). Other effects that differ between chow and defined diets and may be related to either phytoestrogen or fructose content include bone-related changes, plasma estradiol, urinary calcium and corticosterone (Tou et al., 2005xTou, J.C., Arnaud, S.B., Grindeland, R., and Wade, C. Exp. Biol. Med. (Maywood). 2005; 230: 31–39PubMedSee all ReferencesTou et al., 2005), and solution taste preferences (Tordoff et al., 2002xTordoff, M.G., Pilchak, D.M., Williams, J.A., McDaniel, A.H., and Bachmanov, A.A. J. Nutr. 2002; 132: 2288–2297PubMedSee all ReferencesTordoff et al., 2002).When comparing the effects of chow with a defined high-fat diet, the effects of the dietary fat will be confounded with the effects of other components that differ between the diets. Just as it is essential that mouse strains be specified, constituents of experimental diets must be specified (Thigpen et al., 2004xThigpen, J.E., Setchell, K.D., Saunders, H.E., Haseman, J.K., Grant, M.G., and Forsythe, D.B. ILAR J. 2004; 45: 401–416Crossref | PubMedSee all References, Tordoff et al., 1999xTordoff, M.G., Bachmanov, A.A., Friedman, M.I., and Beauchamp, G.K. Science. 1999; 285: 2069PubMedSee all References, Everitt and Foster, 2004xEveritt, J.I. and Foster, P.M. ILAR J. 2004; 45: 417–424Crossref | PubMedSee all References). Specifics to be considered are the objectives of the study and whether the endpoints are affected by several diet components, the maintenance diet, or the palatability of the diet (Thigpen et al., 2004xThigpen, J.E., Setchell, K.D., Saunders, H.E., Haseman, J.K., Grant, M.G., and Forsythe, D.B. ILAR J. 2004; 45: 401–416Crossref | PubMedSee all ReferencesThigpen et al., 2004). Although this correspondence focuses on high-fat diet comparisons, these issues are applicable to any specific nutrient comparisons.

To assess: (1) the relative importance of prehospital physiological measures in identifying high-risk children; (2) whether different age-based criteria should be used for each prehospital physiological measure; and (3) outcome-based appropriate ranges of physiological measures in injured children.This was a retrospective cohort analysis of injured children 0 to 14 years transported by emergency medical services to 48 statewide hospitals from January 1, 1998, through December 31, 2003. We analyzed prehospital physiological measures, including Glasgow Coma Scale score (GCS), systolic blood pressure (SBP), respiratory rate (RR), heart rate, shock index (heart rate/SBP), and airway intervention. "High-risk" children were defined as those with in-hospital mortality, major nonorthopedic surgery, intensive care unit stay greater than or equal to 2 days, or Injury Severity Score greater than or equal to 16. Specific age groups included 0 to 2 years, 3 to 5 years, 6 to 10 years, and 11 to 14 years.A total of 3877 injured children were included in the analysis, of which 1111 (29%) were high risk. Prehospital GCS was the variable of greatest importance in identifying high-risk children, followed by (in order) airway intervention, RR, heart rate, SBP, and shock index. Age modified the effect of prehospital RR (P = 0.0046), heart rate (P = 0.01), and SBP (P = 0.02). There was a linear relationship between GCS and outcome that was consistent across all ages. Specific age-based ranges of other physiological measures were identified for high-risk children.Prehospital GCS and respiratory compromise were the most important physiological measures in identifying high-risk injured children. Age-specific criteria should be considered for RR, heart rate, and SBP.

We previously constructed a congenic mouse, B6.S- D2Mit194-D2Mit311 (B6.S-2) with 27 Mb of SPRET/Ei donor DNA on distal chromosome 2 in a C57BL/6J background that captured an obesity quantitative trait locus (QTL). Mice homozygous for SPRET/Ei alleles at the donor region had decreased body weight and obesity-related phenotypes (Diament AL, Farahani P, Chiu S, Fisler J, Warden CH. Mamm Genome 15: 452–459, 2004). In this study, we constructed five overlapping subcongenics with smaller SPRET/Ei donor regions to fine map the underlying gene(s). One of the five subcongenic lines derived from the B6.S-2 founding congenic, B6.S-2A, captured the body weight and adiposity phenotypes in a donor region with a maximum size of 7.4 Mb. Homozygous SPRET/Ei donor alleles in both the founding congenic and the derived B6.S-2A subcongenic exhibited significant decreases in body weight, multiple fat pad weights, and adiposity index (total fat pad weight divided by body weight). Interval-specific microarray analysis in four tissues for donor region genes from the founding B6.S-2 congenic identified several differentially expressed genes mapping to the B6.S-2A subcongenic donor region, including prohormone convertase 2 (PC2; gene name: Pcsk2). Quantitative real-time PCR confirmed a modest decrease of PC2 expression in brains of mice homozygous for SPRET/Ei donor alleles. Analysis of the relative levels of mRNA for B6 and SPRET/Ei in heterozygous congenic mice showed differentially higher expression of the C57BL/6J allele over the SPRET/Ei allele, indicating a cis regulation of differential expression. Using subcongenic mapping, we successfully narrowed a body weight and obesity QTL interval and identified PC2 as a positional candidate gene.

We have shown that heat-stressed birds exhibit increased superoxide production in skeletal muscle mitochondria. To determine the precise mechanism for this effect, here we studied not only progressive, but also sequential changes in superoxide production, anion carriers and substrate oxidation in mitochondria of heat-stressed chickens. Exposure to acute heat stress (34 degrees C for 6, 12 and 18h) stimulated pectoralis muscle mitochondrial superoxide production. Heat stress-induced downregulations of avUCP gene transcripts and mitochondrial avUCP protein content were time-dependent: avUCP gene transcript was decreased after 6h, while avUCP protein content was only downregulated after 12h of heat stress. Avian adenine nucleotide translocator (avANT) gene transcripts were not changed on exposure to heat stress, suggesting that avANT may not be involved in the regulation of superoxide production in the muscle mitochondria of heat-stressed chickens. During the initial stage of acute heat stress beta-oxidation enzymes gene transcripts and activity were upregulated, with elevated plasma non-esterified fatty acid levels and increased expression of mitochondrial fatty acid transport genes. This sudden surge in mitochondrial substrate oxidation resulted in higher superoxide production: the avUCP expression at 6h after heat stress might have not been large enough to alleviate the overproduction of reactive oxygen species (ROS) even though a small amount of endogenous FFA, a potential uncoupler, might have been present in the mitochondria. Thereafter, avUCP content was downregulated while substrate oxidation returned to control levels. This downregulation of avUCP may have caused increased mitochondrial superoxide production, keeping the superoxide production high in the later stages of heat stress. These results suggest that overproduction of mitochondrial ROS in chicken skeletal muscle under the heat stress might result from enhanced substrate oxidation and downregulation of avUCP in a time-dependent manner.

Publisher Summary This chapter demonstrates that there are many expressed non-protein coding regions in the mammalian genome. The core power of positional genetics approaches is that one can identify the causative factor whether it is an expressed gene, microRNA, antisense, or other unknown mechanisms. New phenotyping resources may also facilitate gene discovery programs. The Mouse Phenome Database includes extensive phenotyping information on dozens of strains. One possibility is that this phenotyping can be combined with high-density mouse haplotyping to find genes for many complex traits. Using genetic approaches leads to unpredictable results; only a small fraction may identify genes that can be targeted by weight loss drugs. Although each obesity gene may have its own interesting story, not all genes have human orthologs that cause obesity; and not all human obesity genes provide safe and effective drug targets. It is not always clear that weight gain or loss in mice can be predicted from measurements of food intake and energy expenditure, possibly because the assays used are not accurate enough to capture the small imbalances of intake and expenditure that cause fat accumulation or because the assays are used for cross-sectional studies at one time point, but the obesity-causing energy imbalance occurs at another time. Settling these questions will allow for a more precise understanding of the mechanisms by which alleles cause fat accumulation.

To evaluate cardiac arrest survival using geographical information systems (GIS) methodology.Patient data were obtained from a fire district Utstein-style adult cardiac arrest registry that also included address data. All incident locations were geocoded and fire station first-due areas were mapped by using the new computer-aided dispatch geographic data. Retrospective assignment of first-due versus second-due fire response unit was done by using a GIS "point-in-polygon" algorithm Survival to hospital admission was the primary outcome measure for incidents responded to by first-due versus second-due apparatus controlling for other potential predictors of survival using logistic regression. Cluster analysis was also performed to evaluate potential areas of high or low rates of survival.There were 461 eligible patients with an average age of 67+/-17 years, 63% were male, 53% had a witnessed arrest, bystander cardiopulmonary resuscitation was performed in 38%, bystander automatic external defibrillator (AED) Page: 1 was used in 0.01%, ventricular fibrillation or ventricular tachycardia were the presenting rhythms in 44%, the average response time was 5.5+/-2.1 minutes, and survival to hospital admission was 17%. There was no significant difference in response time between survivors (4.97 minutes) and non-survivors (5.52 minutes), (difference 0.55 minutes, 95%CI -0.08 to 1.18 min). The number of cardiac arrest calls varied from 1 to 49 for each station and the rate of second-due response varied from 0 to 19%. There was a nonsignificant association of survival to hospital admission for the first-due area cohort: odds ratio 0.70, 95% CI 0.38-1.29.GIS is a new methodology for analyzing EMS incident data. It adds a spatial component of analysis to traditional statistical techniques. No spatial difference was found on patient survival in this analysis.

The objective of this study was to compare the efficacy and adverse events associated with the use of diazepam and midazolam for the treatment of pediatric seizures in the prehospital setting.This was a retrospective cohort study of all patients younger than 18 years treated for a seizure with a benzodiazepine by emergency medical services in Multnomah County, Oregon, from 1998 to 2001. The emergency medical services system consists of a single private advanced life support transporting ambulance service with fire department first responders that are all advanced life support capable. The benzodiazepine used changed from diazepam to midazolam at the midpoint of this period. The primary outcomes were termination of the seizure by arrival to the emergency department (ED), recurrence of seizure while in the ED, or the requirement for active airway interventions including intubation. The two cohorts were also compared for demographics, past history of seizures, long-term use of seizure medications, response times, route of administration, use of second doses of benzodiazepines, and final disposition.Forty-five patients were treated with diazepam, and 48 were treated with midazolam. The two cohorts were comparable except the diazepam cohort had a significantly increased proportion of patients with previous afebrile seizures (53% vs. 25%; p = 0.005). The midazolam cohort had an increased use of nonintravenous route for initial dosing (65% vs. 42%; p = 0.02). The two cohorts were equivalent in rates of termination of seizures before to ED arrival, recurrence of seizures in the ED, requiring airway support or a second dose of benzodiazepine, and admission to the hospital.Diazepam and midazolam appear to be equivalent in treating seizures and causing adverse events. Paramedics appear to be administering midazolam intramuscularly more often than they use diazepam rectally.

There has been intense interest in defining the functions of UCP2 and UCP3 during the nine years since the cloning of these UCP1 homologues. Current data suggest that both UCP2 and UCP3 proteins share some features with UCP1, such as the ability to reduce mitochondrial membrane potential, but they also have distinctly different physiological roles. Human genetic studies consistently demonstrate the effect of UCP2 alleles on type-2 diabetes. Less clear is whether UCP2 alleles influence body weight or body mass index (BMI) with many studies showing a positive effect while others do not. There is strong evidence that both UCP2 and UCP3 protect against mitochondrial oxidative damage by reducing the production of reactive oxygen species. The evidence that UCP2 protein is a negative regulator of insulin secretion by pancreatic β-cells is also strong: increased UCP2 decreases glucose stimulated insulin secretion ultimately leading to β-cell dysfunction. UCP2 is also neuroprotective, reducing oxidative stress in neurons. UCP3 may also transport fatty acids out of mitochondria thereby protecting the mitochondria from fatty acid anions or peroxides. Current data suggest that UCP2 plays a role in the metabolic syndrome through down-regulation of insulin secretion and development of type-2 diabetes. However, UCP2 may protect against atherosclerosis through reduction of oxidative stress and both UCP2 and UCP3 may protect against obesity. Thus, these UCP1 homologues may both contribute to and protect from the markers of the metabolic syndrome.

The recent cloning of a gene that codes for a novel uncoupling protein, UCP2, which is expressed in a wide range of adult human tissues, has raised the possibility that it may be involved in regulation of energy balance. To explore this concept we have investigated potential linkage relationships between three microsatellite markers which encompass the UCP2 gene location on 11q13 with resting metabolic rate (RMR), body mass index, percentage body fat (%FAT) and fat mass (FM) in 640 individuals from 155 pedigrees from the Québec Family Study. Using a linkage analysis strategy based on sibling, avuncular, grandparental and cousin pairs, strong evidence of linkage was found between the marker D11S911 (P = 0.000002) and RMR, with more moderate evidence for D11S916 (P = 0.006) and D11S1321 (P = 0.02). Suggestive evidence of linkage was also observed between D11S1321 and %FAT (P = 0.04) and FM (P = 0.02). It is concluded that the three markers encompassing the UCP2 locus and spanning a 5 cM region on 11q13 are linked to resting energy expenditure in adult humans. The evidence is strong enough to warrant a search for DNA sequence variation in the gene itself.

There is growing awareness that complex interactions among multiple genes and environmental factors play an important role in controlling obesity traits. The BSB mouse, which is produced by the backcross of (lean C57BL/6J × lean Mus spretus) × C57BL/6J, provides an excellent model of epistatic obesity. To evaluate potential epistatic interactions among six chromosomal regions previously determined to influence obesity phenotypes, we performed novel Bayesian analyses on the basis of both epistatic and nonepistatic models for four obesity traits: percentage of body fat, adiposity index, total fat mass, and body weight, and also for plasma total cholesterol. The epistatic analysis detected at least one more QTL than the nonepistatic analysis did for all obesity traits. These obesity traits were variously influenced by QTL on chromosomes 2, 7, 12, 15, and 16. Interaction between genes on chromosomes 2 and 12 was present for all obesity traits, accounting for 3–4.8% of the phenotypic variation. Chromosome 12 was found to have weak main effects on all obesity traits. Several different epistatic interactions were also detected for percentage of body fat, adiposity index, and total fat mass. Chromosomes 6 and 12 have not only main effects but also strong epistatic effects on plasma total cholesterol. Our results emphasize the importance of modeling epistasis for discovery of obesity genes.

Objective: We searched for genes whose alleles cause obesity and novel pathways correlated with obesity. Research Methods and Procedures: BSB mice are a model of complex obesity due to interactions among genes from C57BL/6J (B) and Mus spretus (SPRET) in (B × SPRET) × B backcross mice. Stringent criteria identified 50 genes differentially expressed in epididymal adipose tissue from 7 pairs of lean vs. obese BSB mice. Quantitative reverse transcription-polymerase chain reaction of adipose tissue RNA from 48 BSB mice with a range of obesity was assayed. Leptin was evaluated in inbred (SPRET/Ei) and outbred (SPRET/Pt) BSB mice. Results: Leptin (Lep) and adipsin expressions had the greatest fold differences between obese and lean mice. Four genes involved in iron homeostasis were included in the 50 differentially expressed genes [hemochromatosis (Hfe), diaphorase 1, transferrin receptor (Trfr) 2, and protoporphyrinogen oxidase] and two additional iron-related genes did not quite meet the stringent criteria for differential expression (Trfr and lactotransferrin). Hfe and Trfr mRNA levels and liver iron were negatively correlated with fat mass. Variation in obesity phenotypes explained 49%, 40%, and 37%, respectively, of the variance in Hfe, Lep, and Trfr mRNA levels. Leptin differed by haplotype at the Lep locus in outbred BSB. The quantitative trait locus identified in the outbred cross did not occur in inbred BSB. Discussion: Our results suggest that iron homeostasis in BSB mice is coordinately regulated in vivo in adipose depots in response to obesity. Lep alleles derived from outbred, but not inbred, SPRET are a positional candidate for the chromosome 6 quantitative trait locus in BSB mice.

Objectives: We previously demonstrated coincident quantitative trait loci (QTLs) for percentage body fat, plasma hepatic lipase (HL) activity, and plasma cholesterol on mouse chromosome 7. In the present study, we investigated whether hepatic lipase (Lipc) is an obesity gene, whether Lipc interacts with an unknown gene on chromosome 7, and how HL activity is linked to the chromosome 7 locus. Research Methods and Procedures: BSB mice are a model of complex obesity due to interactions among genes from C57BL/6J and Mus spretus (SPRET) in (C57BL/6J × SPRET) × C57BL/6J backcross mice. Five crosses tested the impact on obesity of combinations of inactive (knockout) and wild-type Lipc alleles from C57BL/6J or SPRET in a reciprocal hemizygosity analysis. Results: The combined data from this allelic series suggest that Lipc alleles, and not alleles from a gene linked to Lipc, influence obesity. No interaction between Lipc and chromosome 7 was demonstrated. We confirmed the chromosome 7 QTLs for obesity, HL activity, and cholesterol. Because obesity and HL activity are not consistently associated in the BSB model, linkage of HL activity to chromosome 7 is not secondary to obesity per se. We also report, for the first time to our knowledge, a QTL in mammals for food intake. Discussion: This use of reciprocal hemizygosity analysis in mammals, which, to our knowledge, is the first reported, reveals its power to detect previously unknown effects of Lipc on obesity.

Our primary objective was to discover simplified mouse models corresponding to human obesity linkages. We used the B10.UW- H3(b) we Pax1(un) a(t)/Sn (B10.UW) congenic strain, a subcongenic strain with a reduced UW strain donor region, and their C57BL/10SnJ background strain. The congenic and subcongenic UW strain donor regions are on mouse Chr 2. We measured body length [anal-nasal (AN) length], summed fat depot weights normalized for body weight (Adiposity Index, AI), and percentage of body weight that is lipid. The B10.UW congenic and subcongenic strains have significantly smaller AN lengths ( p < 0.0001) and have a significantly lower AI and percentage of body weight as fat than the background strain ( p < 0.0001). In an F(2) intercross of the congenic and background strains, AN and AI were both linked to the distal half of the donor region with LOD scores greater than 19 and 5, respectively. F(2) haplotypes identified a minimal region for AN linkage of 0.8 megabases (Mb) that is estimated to express four genes in the current Celera mouse genome assembly. We narrowed the most likely location of the obesity gene to 15 Mb whose homologous genes are all located on human Chr 20 in the region surrounding the centromere. Since a previous study identified human obesity linkage peaking near the centromere, then the B10.UW mice may exhibit obesity due to the homologous gene.

BACKGROUND: Congenic mouse strains contain donor mouse strain DNA in genomes otherwise identical to a background strain. They can be used to identify defined chromosomal regions containing obesity genes with small effects. OBJECTIVE: The objective of this study was to discover congenic strains containing genes that influence body fat in mice and to examine interactions between these genes. DESIGN: A survey of congenic strains showed that the B6.C-Tyrc H1b Hbbd/By (B6.C-H1) congenic strain, with a 24 centiMorgan (cM) donor region from strain BALB/cBy on chromosome 7, had 50% less fat than background C57BL/6By (B6By) mice. The congenic donor region was then divided into 11 smaller overlapping subcongenic regions. Genotype effects on obesity traits in the subcongenics were determined by breeding heterozygotes for each line and comparing phenotypes of littermates with different donor genotypes. RESULTS: At least three subcongenic strains, two with overlapping donor regions and one with a nonoverlapping donor strain region, were found to exhibit significant influences of donor region genotype on obesity. A cross of the two overlapping subcongenics demonstrated that a single gene in the overlap region could not account for the observed obesity effects. We also observed significant obesity differences between genetically identical progeny that were contingent on the genotype of their subcongenic mothers. CONCLUSIONS: These results demonstrate the existence of at least three genes influencing obesity in three subcongenic strains with donor strain chromosomal regions whose size ranges from 0.5 to 5 cM. A maternal effect gene influencing obesity may be present in some subcongenic strains.

We have previously reported that glucose metabolism mediates the effects of insulin to increase leptin gene expression and leptin secretion by isolated adipocytes. The aim of the present study was to investigate the role of transcription and translation in the regulation of basal and insulin-stimulated leptin production. The short-term (4 h) and long-term (24-48 h) effects of actinomycin D, a transcriptional inhibitor, and cycloheximide, an inhibitor of protein synthesis, on leptin gene expression and leptin secretion by isolated adipocytes were determined. Actinomycin D (5 microg/ml) increased both basal and insulin-stimulated (1.6 nM) leptin secretion at 4 and 24h (193+/-14.9% and 153.8+/-10.4% of respective controls at 24h, both p

OBJECTIVE: To test the hypothesis that either uncoupling protein-2 UCP2 or UCP3 or both together influence obesity and inflammation in transgenic mice. DESIGN: We generated 12 lines of transgenic mice for both human UCP2 and 3 using native promoters from a human bacterial artificial chromosome (BAC) clone. The BAC expresses no genes other than UCP2 and 3. Mice used for experiments are N4 or higher of backcross to C57BL/6J (B6). Each experiment used transgenic mice and their nontransgenic littermates. RESULTS: Northern blots confirmed expression on human UCP2 in adipose and spleen, while human UCP3 expression was detectable in gastrocnemius muscle. Western blots demonstrated a four-fold increase of UCP2 protein in spleens of Line 32 transgenic animals. Heterozygous mice of four lines showing expression of human UCP2 in spleen were examined for obesity phenotypes. There were no significant differences between Lines 1 and 32, but female transgenics of both lines had significantly smaller femoral fat depots than the control (littermate) mice (P=0.015 and 0.005, respectively). In addition, total fat of transgenic females was significantly less in Line 1 (P=0.05) and almost significantly different in Line 32 (P=0.06). Male Line 1 mice were leaner (P=0.04) while male Line 32 mice were almost significantly leaner (P=0.06). Heterozygous mice of Lines 35 and 44 showed no significant differences from the nontransgenic littermate controls. Effects of the UCP2/UCP3 transgene on obesity in Line 32 mice were confirmed by crossing transgenic mice with the B6.Cg-Ay agouti obese mice. B6.Cg-Ay carrying the UCP2/UCP3 transgene from Line 32 were significantly leaner than nontransgenic B6.Cg-Ay mice. Line 32 UCP2/UCP3 transgenics showed increased hypothalamic Neuropeptide (NPY) levels and food intake, with reduced spontaneous physical activity. Transgenic baseline interleukin4 (IL-4) and interleukin6 (IL-6) levels were low with lower or later increases after endotoxin injection compared to wild-type littermates. Endotoxin-induced fever was also diminished in transgenic male animals. Low-density lipoprotein (LDL) cholesterol levels were significantly higher in both Line 1 and 32 transgenics (P=0.05 and 0.001, respectively) after they had been placed on a moderate fat-defined diet containing 32% of calories from fat for 5 weeks. CONCLUSION: Moderate overexpression of UCP2 and 3 reduced fat mass and increased LDL cholesterol in two independent lines of transgenic mice. Thus, the reduced fat mass cannot be due to insertional mutagenesis since virtually identical fat pad weights and masses were observed with the two independent lines. Line 32 mice also have altered inflammation and mitochondrial function. We conclude that UCP2 and/or 3 have small but significant effects on obesity in mice, and that their mechanism of action may include alterations of metabolic rate.

Note: Please be advised that the American Academy of Pediatrics and the American College of Emergency Physicians are in the process of independently developing technical reports on this issue that will provide more in-depth educational and clinical information for their respective members on the death of a child in the emergency department. When completed, these reports will be published separately by each organization to supplement this joint policy statement.

Abuse of ethanol is a major risk factor in medicine, in part because of its widespread effect on the activity of the central nervous system, including behavior, pain, and temperature sensation. Uncoupling protein 2 (UCP2) is a mitochondrial protonophore that regulates cellular energy homeostasis. Its expression in mitochondria of axons and axon terminals of basal forebrain areas suggests that UCP2 may be involved in the regulation of complex neuronal responses to ethanol. We employed a paradigm in which acute exposure to ethanol induces tolerance and altered pain and temperature sensation. In UCP2 overexpressing mice, sensitivity to ethanol was decreased compared to that of wild-type animals, while UCP2 knockouts had increased ethanol sensitivity. In addition, UCP2 expression was inversely correlated with the impairment of pain and temperature sensation induced by ethanol. Taken together, these results indicate that UCP2, a mitochondrial uncoupling protein previously associated with peripheral energy expenditure, is involved in the mediation of acute ethanol exposure on the central nervous system. Enhancement of UCP2 activation after acute alcohol consumption might decrease the time of recovery from intoxication, whereas UCP2 inhibition might decrease the tolerance to ethanol.

Injury is the number one cause of death and life-years lost for children. In children, injury mortality is greater than childhood mortality from all other causes combined. Modern injury prevention and control seeks to prevent and limit or control injuries through the 4 Es of injury prevention: engineering, enforcement, education, and economics. Emergency physicians are often placed in a critical role in the lives of individuals, are respected authorities on the health and safety of children and adults, and have daily exposure to high-risk populations. This gives emergency physicians a unique perspective and an opportunity to take an active role in injury control and prevention. Specific methods or strategies for promulgating injury prevention and control in our emergency medicine practices are suggested, ranging from education (for our patients and health professionals); screening and intervention for domestic violence, child maltreatment, drug-alcohol dependency and abuse; data collection; reporting unsafe products; research; legislation; serving in regulatory and governmental agencies; emergency medical services-community involvement; and violence prevention. Emergency physicians can play a significant role in decreasing pediatric injury and its concomitant morbidity and mortality.

Uncoupling proteins (UCPs) are composed of three repeated domains of approximately 100 amino acids each. We have used chimeras of UCP1 and UCP2, and electron paramagnetic resonance (EPR), to investigate domain specific properties of these UCPs. Questions include: are the effects of nucleotide binding on proton transport solely mediated by amino acids in the third C-terminal domain, and are the amino acids in the first two domains involved in retinoic or fatty acid activation? We first confirmed that our reconstitution system produced UCP1 that exhibited known properties, such as activation by fatty acids and inhibition of proton transport by purine nucleotides. Our results confirm the observations reported for recombinant yeast that retinoic acid, but not fatty acids known to activate UCP1, activates proton transport by UCP2 and that this activation is insensitive to nucleotide inhibition. We constructed chimeras in which the last domains of UCP1 or UCP2 were switched and tested for activation by fatty acids or retinoic acid and inhibition by nucleotides. U1U2 is composed of mUCP1 (amino acids 1-198) and hUCP2 (amino acids 211-309). Fatty acids activated proton transport of U1U2 and GTP mediated inhibition. In the other chimeric construct U2U1, hUCP2 (amino acids 1-210) and mUCP1 (amino acids 199-307), retinoic acid still acted as an activator, but no inhibition was observed with GTP. Using EPR, a method well suited to the analysis of the structure of membrane proteins such as UCPs, we confirmed that UCP2 binds nucleotides. The EPR data show large structural changes in UCP1 and UCP2 on exposure to ATP, implying that a putative nucleotide-binding site is present on UCP2. EPR analysis also demonstrated changes in conformation of UCP1/UCP2 chimeras following exposure to purine nucleotides. These data demonstrate that a nucleotide-binding site is present in the C-terminal domain of UCP2. This domain was able to inhibit proton transport only when fused to the N-terminal part of UCP1 (chimera U1U2). Thus, residues involved in nucleotide inhibition of proton transport are located in the two first carrier motifs of UCP1. While these results are consistent with previously reported effects of the C-terminal domain on nucleotide binding, they also demonstrate that interactions with the N-terminal domains are necessary to inhibit proton transport. Finally, the results suggest that proteins such as UCP2 may transport protons even though they are not responsible for basal or cold-induced thermogenesis.

Altered ambient force environments affect energy expenditure via changes in thermoregulation, metabolism, and body composition. Uncoupling proteins (UCPs) have been implicated as potential enhancers of energy expenditure and may participate in some of the adaptations to a hyperdynamic environment. To test this hypothesis, this study examined the homeostatic and circadian profiles of body temperature (Tb) and activity and adiposity in wild-type and UCP2/3 transgenic mice exposed to 1 and 2 G. There were no significant differences between the groups in the means, amplitudes, or phases of Tb and activity rhythms at either the 1- or 2-G level. Percent body fat was significantly lower in transgenic (5.2 ± 0.2%) relative to the wild-type mice (6.2 ± 0.1%) after 2-G exposure; mass-adjusted mesenteric and epididymal fat pads in transgenic mice were also significantly lower ( P < 0.05). The data suggest that 1) the actions of two UCPs (UCP2 and UCP3) do not contribute to an altered energy balance at 2 G, although 2) UCP2 and UCP3 do contribute to the utilization of lipids as a fuel substrate at 2 G.

Distinct brain peptidergic circuits govern peripheral energy homeostasis and related behavior. Here we report that mitochondrial uncoupling protein 2 (UCP2) is expressed discretely in neurons involved in homeostatic regulation. UCP2 protein was associated with the mitochondria of neurons, predominantly in axons and axon terminals. UCP2-producing neurons were found to be the targets of peripheral hormones, including leptin and gonadal steroids, and the presence of UCP2 protein in axonal processes predicted increased local brain mitochondrial uncoupling activity and heat production. In the hypothalamus, perikarya producing corticotropin-releasing factor, vasopressin, oxytocin, and neuropeptide Y also expressed UCP2. Furthermore, axon terminals containing UCP2 innervated diverse hypothalamic neuronal populations. These cells included those producing orexin, melanin-concentrating hormone, and luteinizing hormone-releasing hormone. When c-fos-expressing cells were analyzed in the basal brain after either fasting or cold exposure, it was found that all activated neurons received a robust UCP2 input on their perikarya and proximal dendrites. Thus, our data suggest the novel concept that heat produced by axonal UCP2 modulates neurotransmission in homeostatic centers, thereby coordinating the activity of those brain circuits that regulate daily energy balance and related autonomic and endocrine processes.

The newly discovered uncoupling protein-2 (UCP2) has been proposed to play a critical role in thermoregulatory and substrate oxidation processes. Skeletal muscle mRNA expression and, more recently, the protein content of UCP2 were investigated in humans. These studies have shown that the content of this protein varies quite substantially and that several factors could be responsible for its variation in human skeletal muscle. The aim of this review is to determine whether obesity and low-intensity increased contractile activity contribute to variation in muscle UCP2 content. A recent study from our laboratories revealed that, in obesity, UCP2 content in skeletal muscle is over-expressed by about 1.5-fold compared to lean. Body weight loss in obese subjects did not cause any change in skeletal muscle UCP2 content. On the other hand, when increased muscular contractile activity of knee extensor muscles is induced by several weeks of low-frequency electrical stimulation, UCP2 content increased by about 15%. Obesity and increased contractile activity do not appear sufficient, however, to explain the magnitude of the human skeletal muscle variation in UCP2 content. Since intensive efforts are being devoted to this area of research, it is expected that our understanding of the causes contributing to its variation in humans will soon be substantially improved.

The molecular basis for variations in resting metabolic rate (RMR) within a species is unknown. One possibility is that variations in RMR occur because of variations in uncoupling protein 2 (UCP-2) and uncoupling protein 3 (UCP-3) expression, resulting in mitochondrial proton leak differences. We tested the hypothesis that UCP-2 and -3 mRNAs positively correlate with RMR and proton leak. We treated thyroidectomized and sham-operated mice with triiodothyronine (T3) or vehicle and measured RMR, liver, and skeletal muscle mitochondrial nonphosphorylating respiration and UCP-2 and -3 mRNAs. T3stimulated RMR and liver UCP-2 and gastrocnemius UCP-2 and -3 expression. Mitochondrial respiration was not affected by T3and did not correlate with UCP-2 and -3 mRNAs. Gastrocnemius UCP-2 and -3 expression did correlate with RMR. We conclude 1) T3did not influence intrinsic mitochondrial properties such as membrane structure and composition, and 2) variations in UCP-2 and -3 expression may partly explain variations in RMR. One possible explanation for these data is that T3stimulates the leak in vivo but not in vitro because a posttranslational regulator of UCP-2 and -3 is not retained in the mitochondrial fraction.

We describe an unusual case of a toddler diagnosed with an idiopathic seizure disorder that later was proved to be caused by deliberate administration of amitriptyline by his custodian. In spite of seizures associated with widened electrocardiographic wave (QRS) and right axis deviation on the electrocardiogram (EKG), the correct diagnosis eluded clinicians through a series of hospital admissions. Unfortunately, clinicians are quite accustomed to the fact that patients previously diagnosed with epilepsy have seizures and may not investigate other causes of seizure. This allowed classic signs of cyclic antidepressant poisoning to go unrecognized. Language: en

Aims/hypothesis. Linkage between markers close to the uncoupling protein 2 and 3 genes (11q13) and resting metabolic rate and a pre-diabetic phenotype have been found. The syntenic region in mouse has been found to be linked to quantitative traits associated with obesity and diabetes. UCP2 and UCP3 could therefore have an important role in body weight regulation and susceptibility to diabetes. We investigated a recently identified variant of the UCP2 gene in exon 8 as a marker for glucose and weight homeostasis. Methods. Length variation of the UCP2 exon 8 variant was studied by the polymerase chain reaction and agarose gel electrophoresis. Sequence variants of the UCP3 gene were sought by semi-automated DNA sequencing. Results. In 453 South Indian subjects, we found an association in women between the UCP2 exon variant and body mass index (p = 0.018). These findings were replicated in a separate group of South Indian subjects (n = 143, p < 0.001) irrespective of sex. Although no association was found between the UCP2 exon 8 variant and overt obesity in British subjects, the UCP2 genotype of obese women (n = 83) correlated with fasting serum leptin concentration (p = 0.006) in the presence of extreme obesity. These observations could not be explained by tight linkage disequilibrium with a coding region variant in the region of the UCP3 gene of biological significance. Lastly, no association was found between UCP2 and Type II (non-insulin-dependent) diabetes using either a family based design (85 families) or case control study (normal glucose tolerance n = 335, impaired glucose tolerance n = 42, Type II diabetes n = 76). Conclusion/interpretation. We have described a UCP2 gene exon 8 variant that may affect susceptibility to weight gain by influencing regulation of leptin. [Diabetologia (1999) 42: 688–693]

We report here the cloning and functional analysis of a novel homologue of the mitochondrial carriers predominantly expressed in the central nervous system and referred to as BMCP1 (brain mitochondrialcarrier protein-1). The predicted amino acid sequence of this novel mitochondrial carrier indicates a level of identity of 39, 31, or 30%, toward the mitochondrial oxoglutarate carrier, phosphate carrier, or adenine nucleotide translocator, respectively, and a level of identity of 34, 38, or 39% with the mitochondrial uncoupling proteins UCP1, UCP2, or UCP3, respectively. Northern analysis of mouse, rat, or human tissues demonstrated that mRNA of this novel gene is mainly expressed in brain, although it is 10–30-fold less expressed in other tissues.In situ hybridization analysis of brain showed it is particularly abundant in cortex, hippocampus, thalamus, amygdala, and hypothalamus. Chromosomal mapping indicates that BMCP1 is located on chromosome X of mice and at Xq24 in man. Expression of the protein in yeast strongly impaired growth rate. Analysis of respiration of total recombinant yeast or yeast spheroplasts and in particular of the relationship between respiratory rate and membrane potential of yeast spheroplasts revealed a marked uncoupling activity of respiration, suggesting that although BMCP1 sequence is more distant from the uncoupling proteins (UCPs), this protein could be a fourth member of the UCP family.