A novel peripheral cannabinoid receptor 1 antagonist, BPR0912, reduces weight independently of food intake and modulates thermogenesis.

AIM: To investigate the in vivo metabolic effects of treatment with BPR0912, a novel and potent peripheral cannabinoid receptor 1 (CB1R) antagonist, on both normal mice and diet-induced obese (DIO) mice. METHODS: The acute peripheral effects of BPR0912 administration on gastrointestinal transit and energy metabolism in normal mice were investigated. The effects of chronic BPR0912 treatment were compared with those of rimonabant using DIO mice. Alterations to body weight and biochemical and metabolic variables were determined. RESULTS: Acute treatment with BPR0912 did not alter food intake or energy metabolism, but efficiently reversed CB1R-mediated gastrointestinal delay. Chronic treatment of DIO mice with BPR0912 showed that BPR0912 exerts a food intake-independent mechanism, which contributes to weight loss. Genes involved in ß-oxidation and thermogenesis were upregulated in white adipose tissue (WAT) in addition to increased lipolytic activity, whereas Ucp1 expression was induced in brown adipose tissue (BAT) and body temperature was elevated. Expression of the ß2-adrenoceptor was specifically elevated in both WAT and BAT in a manner dependent on the BPR0912 dose. Lastly, chronic BPR0912 treatment was more efficacious than rimonabant in reducing hepatic triglycerides in DIO mice. CONCLUSION: BPR0912 exhibits significant in vivo efficacy in inducing food intake-independent weight loss in DIO mice, while tending to reduce their hepatic steatosis. The thermogenic effects of BPR0912, as well as its modulation of protein and gene expression patterns in WAT and BAT, may enhance its efficacy as an anti-obesity agent. The results of the present study support the benefits of the use of peripheral CB1R antagonists to combat metabolic disorders.

Induction of fatty acid oxidation resists weight gain, ameliorates hepatic steatosis and reduces cardiometabolic risk factors.

OBJECTIVE: Fatty acid oxidation has been implicated in amelioration of obesity by burning off excessive accumulated lipid. BPR697, a peripheral cannabinoid receptor 1 (CB1) antagonist, elevated fat oxidation without added energy expenditure. Its impact on food intake, body weight changes and metabolic alterations were examined in rats fed standard chow and in diet-induced obesity (DIO) mice. MATERIALS AND METHODS: CB1 agonist-induced hypothermia and analgesia responses were measured to examine the brain activity of BPR697. The acute effects of BPR697 on food intake, body weight change and post-absorptive metabolic profiles were investigated in rats. Energy utilization with BPR697 was examined by indirect calorimetry. Chronic treatment of DIO mice was used to evaluate the long-term effects of BPR697. RESULTS: Distribution of BPR697 was significantly biased in favor of the periphery instead of the brain, as shown by its low brain/plasma concentration ratio and confirmed by the negative response of BPR697 in CB1 agonist-induced hypothermia and analgesia. When administered to rats at 20 mg kg(-1), BPR697 showed a unique spectrum of effects with significant weight loss without altered food intake. Furthermore, BPR697 increased serum levels of free fatty acids and ketone bodies and reduced hepatic lipid accumulation with preservation of liver glycogen in postprandial rats. Indirect calorimetric profiling of BPR697 revealed a similar trend, shifting whole-body energy catabolism toward fat oxidation, but without elevated energy expenditure. In DIO mice with chronic treatment, animals treated with BPR697 at 20 mg kg(-1) resisted weight gain and showed a reduction of high-fat-induced cardiometabolic abnormalities such as hyperglycemia, abdominal fat and liver steatosis. CONCLUSION: The induction of fatty acid oxidation without concomitant elevation of energy expenditure by the peripheral CB1 antagonist BPR697 is sufficient to cause substantial weight loss in chow-fed rats. In the presence of high-dietary fat intake, BPR697 resists weight gain and alleviates obesity-related cardiometabolic risk factors.

Film cooling performance on curved walls with compound angle hole configuration.

In order to explore the effect of compound angle holes on film cooling over a convex wall and a concave wall, the present study adopts the transient liquid crystal thermography for conducting the film cooling measurement on simple hole and expanded-hole configurations. Two compound angles of 0 and 45 deg are tested at an elevated mainstream turbulence condition (Tu) of 3.8%. The test pieces have the different radius of curvature (2r/D) of 92.5 on convex and 86.5 on concave, and the same pitch to diameter ratio (P/D) of 3 on both convex and concave walls. All measurements were conducted under the mainstream Reynolds number (Re(d)) of 1700 on convex and 2300 on concave with the density ratio between coolant and mainstream (rho c/rho m) of 0.98. In current study, the effect of blowing ratio (M) on film cooling performance is investigated by varying the range of blowing ratio from 0.5 and 2.0. The present measured results show that the forward-expanded hole injection provides better surface protection than the simple hole injection. As far as the injection angle is concerned, compound angle injection provides higher film effectiveness than simple angle injection. However, the forward-expanded hole in injection (beta = 0 degree) has the best performance on both convex and concave surfaces.

Transcriptional repression by Drosophila methyl-CpG-binding proteins.

C methylation at genomic CpG dinucleotides has been implicated in the regulation of a number of genetic activities during vertebrate cell differentiation and embryo development. The methylated CpG could induce chromatin condensation through the recruitment of histone deacetylase (HDAC)-containing complexes by methyl-CpG-binding proteins. These proteins consist of the methylated-DNA binding domain (MBD). Unexpectedly, however, several studies have identified MBD-containing proteins encoded by genes of Drosophila melanogaster, an invertebrate species supposed to be void of detectable m(5)CpG. We now report the genomic structure of a Drosophila gene, dMBD2/3, that codes for two MBD-containing, alternatively spliced, and developmentally regulated isoforms of proteins, dMBD2/3 and dMBD2/3Delta. Interestingly, in vitro binding experiments showed that as was the case for vertebrate MBD proteins, dMBD2/3Delta could preferentially recognize m(5)CpG-containing DNA through its MBD. Furthermore, dMBD2/3Delta as well as one of its orthologs in mouse, MBD2b, could function in human cells as a transcriptional corepressor or repressor. The activities of HDACs appeared to be dispensable for transcriptional repression by dMBD2/3Delta. Finally, dMBD2/3Delta also could repress transcription effectively in transfected Drosophila cells. The surprisingly similar structures and characteristics of the MBD proteins as well as DNA cytosine (C-5) methyltransferase-related proteins in Drosophila and vertebrates suggest interesting scenarios for their roles in eukaryotic cellular functions.

Estimating probabilities of diabetes mellitus using neural networks.

Classification problems are often encountered in medical diagnosis. This paper presents an introduction to classification theory and shows how artificial neural networks can be used for classification. We also map out a bootstrapped procedure for interval estimation of posterior probabilities. The entire procedure is illustrated using the diabetes mellitus data in Pima Indians.

Drosophila proteins related to vertebrate DNA (5-cytosine) methyltransferases.

DNA methylation at CpG residues is closely associated with a number of biological processes during vertebrate development. Unlike the vertebrates, however, several invertebrate species, including the Drosophila, do not have apparent DNA methylation in their genomes. Nor have there been reports on a DNA (5-cytosine) methyltransferase (CpG MTase) found in these invertebrates. We now present evidence for two CpG MTase-like proteins expressed in Drosophila cells. One of these, DmMTR1, is a protein containing peptide epitopes immunologically related to the conserved motifs I and IV in the catalytic domain of the mammalian dnmt1. DmMTR1 has an apparent molecular mass of 220 kDa and, similar to mammalian dnmt1, it also interacts in vivo with the proliferating cell nuclear antigen. During interphase of the syncytial Drosophila embryos, the DmMTR1 molecules are located outside the nuclei, as is dnmt1 in the mouse blastocyst. However, DmMTR1 appears to be rapidly transported into, and then out of the nuclei again, as the embryos undergo mitotic waves. Immunofluorescent data indicate that DmMTR1 molecules "paint" the whole set of condensed Drosophila chromosomes throughout the mitotic phase, suggesting they may play an essential function in the cell-cycle regulated condensation of the Drosophila chromosomes. Through search in the genomic database, we also have identified a Drosophila polypeptide, DmMT2, that exhibits high sequence homology to the mammalian dnmt2 and the yeast CpG MTase homolog pmt1. The expression of DmMT2 appears to be developmentally regulated. We discuss the evolutionary and functional implications of the discovery of these two Drosophila proteins related to mammalian CpG MTases.

vab-8 is a key regulator of posteriorly directed migrations in C. elegans and encodes a novel protein with kinesin motor similarity.

Nervous system assembly requires the directed migrations of cells and axon growth cones along the dorsoventral and anteroposterior axes. Although guidance mechanisms for dorsoventral migrations are conserved from nematodes to mammals, mechanisms for anteroposterior migrations are unknown. In C. elegans, the gene vab-8, which specifically functions in posteriorly directed migrations, encodes two isoforms of a novel intracellular protein that act cell-autonomously in different migrations. VAB-8L, which contains a domain similar to kinesin-like motors, functions in all vab-8-dependent axon growth cone migrations. VAB-8S, which lacks this N-terminal domain, functions in a subset of vab-8-dependent cell migrations. Continuous expression of VAB-8L in the ALM mechanosensory neuron, which normally requires vab-8 early in its development for posteriorly directed cell migration, redirects its anteriorly projecting axon posteriorly. We propose that regulation of vab-8 activity is a mechanism for controlling the direction of cell and axon growth cone migrations.

Suppressors of the unc-73 gene of Caenorhabditis elegans.

The unc-73 gene of Caenorhabditis elegans is necessary for proper axon guidance. Animals mutant in this gene are severely uncoordinated and also exhibit defects in cell migration and cell lineages. We have isolated coordinated revertants of unc-73 (e936). These fall into three classes: intragenic revertants, extragenic dominant suppressors (sup-39), and a single apparently intragenic mutation that is a dominant suppressor with a linked recessive lethal phenotype. sup-39 mutations cause early embryonic lethality, but escapers have a wild-type movement phenotype as larvae and adults. Gonads of sup-39 mutant animals show a novel defect: normal gonads have a single row of oocytes, but sup-39 gonads often have two rows of oocytes. This result suggests that the mutant gonad is defective in choosing on its surface only a single site form which nuclei will emerge to form oocytes. These results are interpreted in terms of an effect of unc-73 on determination of cell polarity.

Blunt gastrointestinal injury in trauma patients.

The evaluation, management, and final outcome of 34 patients with blunt gastrointestinal injury (BGI) were reviewed. Initial absence of symptoms and signs led to two delayed diagnoses. Sonography provided 80% (12/15) positive-predictive value, and three false-negative patients were subsequently detected by diagnostic peritoneal lavage (DPL). Besides repeated clinical surveillance, screening by sonography complemented with DPL provided early detection of blunt bowel injury in trauma patients. In the outcome analysis, BGI patients with high injury-severity scores, intraoperative hypotension, or accompanying major medical diseases were associated significantly with increased risk of infectious complications (P < .05).

Cell polarity and the mechanism of asymmetric cell division.

During development, one mechanism for generating different cell types is asymmetric cell division, by which a cell divides and contributes different factors to each of its daughter cells. Asymmetric cell division occurs throughout the eukaryotic kingdom, from yeast to humans. Many asymmetric cell divisions occur in a defined orientation. This implies a cellular mechanism for sensing direction, which must ultimately lead to differences in gene expression between two daughter cells. In this review, we describe two classes of molecules: regulatory factors that are differentially expressed upon asymmetric cell division, and components of a signal transduction pathway that may define cell polarity. The lin-11 and mec-3 genes of C. elegans, the Isl-1 gene of mammals and the HO gene of yeast, encode regulatory factors that determine cell type of one daughter after asymmetric cell division. The CDC24 and CDC42 genes of yeast affect both bud positioning and orientation of mating projections, and thus may define a general cellular polarity. We speculate that molecules such as Cdc24 and Cdc42 may regulate expression of genes such as lin-11, mec-3, Isl-1 and HO upon asymmetric cell division.

Cyclical strain increases endothelin-1 secretion and gene expression in human endothelial cells.

We examined the effects of cyclical strain on endothelin-1 (Et-1) secretion and Et-1 mRNA levels in human umbilical vein endothelial cells. Cultured endothelial cells grown on a flexible membrane base were deformed by vacuum to 20% of maximum strain, at 60 cycles/min, for various time periods. The rate of Et-1 release from strain- treated cells and their Et-1 mRNA levels were measured. Cells subjected to strain increased their Et-1 secretion into the culture medium from 0.17 ng/hr/10(6) cells to 0.44 ng/hr/10(6) cells. Concomitantly, the Et-1 mRNA levels in strained cells increased 1.4-, 2.1- or 2.6- fold (vs control unstrained cells) after 0.25, 0.5 or 6 hours of strain, respectively. Cells exposed to longer periods of strain (> 6 hrs) did not further increase their mRNA expression. Pretreatment with calphostin C, a specific inhibitor of protein kinase C, before straining completely abolished Et-1 mRNA expression. These results indicate that mechanical strain can modulate the secretion of Et-1 from endothelial cells by increasing Et-1 mRNA levels via the protein kinase C pathway. Elevation of Et-1 secretion and gene expression in endothelial cells under physiological strain might influence normal or pathological states of the vasculature and cardiac growth.

Effects of PGE1 on platelet deformability.

Prostaglandin E1 (PGE1) is frequently used as a stabilizing agent while isolating platelets. The platelet-stabilizing effect of PGE1 is mediate through the PGE1 receptor and the consequent increase of intracellular cAMP. The discoid shape of platelet is thus maintained. Recently, we have studied the effect of PGE1 on platelet deformability with micropipette aspiration technics, using pipettes with radii (Rp) of 0.26-0.36 micron. Platelets, obtained from platelet-rich plasma, and suspended in Hank's buffer containing BSA, glucose, adenosine and theophylline, were subjected for micropipette aspiration test within 2 hr after sampling. In general, PGE1 treated platelets (greater than 75%) remained as non-activated form as revealed under video microscopy and maintained their discoid shape for as long as 6 hr. The time course of the extension length (Dp, in micron) of the platelets in response to aspiration with a negative pressure of 5 cm H2O (tension = 0.15 dynes/cm) was analyzed by using a digital image analyzer system (IRIS 3120 with IP-512, with a resolution of 0.076 microns). A computer program based on the algorithm of real-time reading of image intensity was developed to facilitate real-time data acquisition. The data of Dp/Rp vs time (sec) were fitted by using a second-order equation. The deformability of PGE1 (0.1 microM) treated platelet was decreased significantly as compared with control. These results indicate that the platelet stabilization by PGE1 is accompanied by a decrease in platelet deformability.

Purification and characterization of bilirubin UDP-glucuronyltransferase from the liver of eel, Anguilla japonica.

1. The enzyme bilirubin uridine diphosphate glucuronyltransferase (UDPGT) was purified and characterized from the liver of eel, Anguilla japonica. 2. The molecular weight of the enzyme was 88,000. 3. The optimal working pH of the enzyme was 7.5-8.0. The optimal working temperature of the enzyme was around 46 degrees C. 4. The Km of bilirubin and uridine diphosphate glucuronic acid for this enzyme was 1.6 mM and 1.8 mM respectively. 5. The concentration of bilirubin did not show any significant effect of inhibition on this enzyme up to 3.3 mM. 6. Since this is the first time UDPGT has been purified and characterized from poikilothermic aquatic animals, it provided interesting information on evolution and adaptation of this enzyme when compared to that of mammals.