Centrally Administered Ghrelin Acutely Influences Food Choice in Rodents

We sought to determine whether the orexigenic hormone, ghrelin, is involved in the intrinsic regulation of food choice in rats. Ghrelin would seem suited to serve such a role given that it signals hunger information from the stomach to brain areas important for feeding control, including the hypothalamus and reward system (e.g. ventral tegmental area, VTA). Thus, in rats offered a choice of palatable foods (sucrose pellets and lard) superimposed on regular chow for 2 weeks, we explored whether acute central delivery of ghrelin (intracerebroventricular (ICV) or intra-VTA) is able to redirect their dietary choice. The major unexpected finding is that, in rats with high baseline lard intake, acute ICV ghrelin injection increased their chow intake over 3-fold, relative to vehicle-injected controls, measured at both 3 hr and 6 hr after injection. Similar effects were observed when ghrelin was delivered to the VTA, thereby identifying the VTA as a likely contributing neurobiological substrate for these effects. We also explored food choice after an overnight fast, when endogenous ghrelin levels are elevated, and found similar effects of dietary choice to those described for ghrelin. These effects of fasting on food choice were suppressed in models of suppressed ghrelin signaling (i.e. peripheral injection of a ghrelin receptor antagonist to rats and ghrelin receptor (GHSR) knock-out mice), implicating a role for endogenous ghrelin in the changes in food choice that occur after an overnight fast. Thus, in line with its role as a gut-brain hunger hormone, ghrelin appears to be able to acutely alter food choice, with notable effects to promote “healthy” chow intake, and identify the VTA as a likely contributing neurobiological substrate for these effects.     

Cardiac Fibroblasts Contribute to Myocardial Dysfunction in Mice with Sepsis: The Role of NLRP3 Inflammasome Activation

Myocardial contractile dysfunction in sepsis is associated with the increased morbidity and mortality. Although the underlying mechanisms of the cardiac depression have not been fully elucidated, an exaggerated inflammatory response is believed to be responsible. Nucleotide-binding oligomerization domain-like receptor containing pyrin domain 3 (NLRP3) inflammasome is an intracellular platform that is involved in the maturation and release of interleukin (IL)-1β. The aim of the present study is to evaluate whether sepsis activates NLRP3 inflammasome/caspase-1/IL-1β pathway in cardiac fibroblasts (CFs) and whether this cytokine can subsequently impact the function of cardiomyocytes (cardiac fibroblast-myocyte cross-talk). We show that treatment of CFs with lipopolysaccharide (LPS) induces upregulation of NLRP3, activation of caspase-1, as well as the maturation (activation) and release of IL-1β. In addition, the genetic (small interfering ribonucleic acid [siRNA]) and pharmacological (glyburide) inhibition of the NLRP3 inflammasome in CFs can block this signaling pathway. Furthermore, the inhibition of the NLRP3 inflammasome in cardiac fibroblasts ameliorated the ability of LPS-chalenged CFs to impact cardiomyocyte function as assessed by intracellular cyclic adenosine monophosphate (cAMP) responses in cardiomyocytes. Salient features of this the NLP3 inflammasome/ caspase-1 pathway were confirmed in in vivo models of endotoxemia/sepsis. We found that inhibition of the NLRP3 inflammasome attenuated myocardial dysfunction in mice with LPS and increased the survival rate in mice with feces-induced peritonitis. Our results indicate that the activation of the NLRP3 inflammasome in cardiac fibroblasts is pivotal in the induction of myocardial dysfunction in sepsis.     

Methods of tracing hydrogen in polyketide biosynthesis: High field NMR spectroscopy of citrinin produced in a D2O based medium

Penicillium citrinum cultures have been germinated on an H₂O-based medium, resuspended on a D₂O-based medium and treated with [l,2-¹³C₂] acetate. The resulting citrinin (1) has been analysed by²H and¹³C nuclear magnetic resonance spectroscopy and information about the metabolism of hydrogen in citrinin biosynthesis has been deduced.     

Variations of susceptibility to alloxan induced diabetes in the rabbit

The variations of susceptibility to alloxan induced Diabetes in a total of seventeen rabbits was described. Our study was designed to explore dosage schedules which might improve rabbit responsiveness to and survival after alloxan treatment. A wide range of response to intravenously administered alloxan was observed. Permanent diabetes (blood glucose 350 mg/dl) was found in three rabbits after a single injection (60 mg/kg in one, 100 mg/kg in two). This effect has persisted for eight months. By contrast, two other rabbits injected with a single dose of alloxan (60 mg/kg) developed only transient hyperglycemia. Similarly, four other rabbits either did not respond or had an incomplete response after receiving a total dose of 120 mg/kg. These data suggest that there is extreme variability in individual rabbits susceptibility to the diabetogenic affects of alloxan.     

HIV-specific antibody among voluntary blood donors in Lower Saxony (FRG)

Anti-HIV test results of the Red Cross Blood Transfusion Service of Lower Saxony from 1 June 1985 to 31 July 1986 inclusive were analysed retrospectively. Nine out of 70,936 donors who had not donated blood before 1 June 1985 (first-time donors) and 9 out of 261,231 donors who had donated blood before this date (repeating donors) were found anti-HIV confirmed positive at the time of the first blood donation during the study period. The prevalence of HIV antibody in first-time donors was significantly higher than in repeating donors (p less than 0.01). It was concluded that some members of risk groups used blood donation to obtain an anti-HIV test result. One out of 30,300 blood donations was confirmed anti-HIV positive. The results of this study justify the transfusion of blood donations that are reactive only in the initial ELISA test.     

Role of Ethylene Metabolism in Amaranthus retroflexus

¹⁴C-Ethylene was metabolized by etiolated pigweed seedlings (Amaranthus retroflexus L.) in the manner similar to that observed in other plants. The hormone was oxidized to ¹⁴CO₂ and incorporated into ¹⁴C-tissue components. Selected cyclic olefins with differing abilities to block ethylene action were used to determine if ethylene metabolism in pigweed is necessary for ethylene action. 2,5-Norbornadiene and 1,3-cyclohexadiene were effective inhibitors of ethylene action at 800 and 6400 microliters per liter, respectively, in the gas phase, while 1,4-cyclohexadiene and cyclohexene were not. However, all four cyclic olefins inhibited the incorporation and conversion of ¹⁴C-ethylene to ¹⁴CO₂ by 95% with I₅₀ values below 100 microliters per liter. The results indicate that total ethylene metabolism does not directly correlate with changes in ethylene action. Additionally, the fact that inhibition of ethylene metabolism by the cyclic olefins did not result in a corresponding increase in ethylene evolution indicates that ethylene metabolism does not serve to significantly reduce endogenous ethylene levels.     

Continuous microbial desulfurization of coal--application of a multistage slurry reactor and analysis of the interactions of microbial and chemical kinetics

Microbial desulfurization of coal by pyrite oxidizing bacterial enrichment cultures has been studied in air-agitated slurry reactors of 4- and 20-L volumes. Batch experiments showed that inoculation with an active bacterial culture is essential to minimize the lag phase, although a considerable number of pyrite oxidizing bacteria was found on the coal prior to desulfurization. For detailed investigations of kinetics, energy requirements, and technical applicability, a bioreactor equipment consisting of a cascade of eight stages was developed and operated continuously. Microbial desulfurization of coal-monitored by measuring the axial profile of dissolved iron concentration, real and maximum oxygen consumption rates, and cell concentration-at pulp densities to 30% was performed over a period of 200 days without any disturbances concerning the aeration system, fluidization, transport of solids and microbial growth. At a pulp density of 20%, a pyrite conversion of 68% was achieved after the third reactor stage at a total residence time of five days in the first three stages. The kinetics of pyrite degradation were found to be well described by a rate equation of first order in pyrite surface area concentration if the pyrite is directly accessible for microbial attack. Rate constants were determined to 0.48 mg pyrite/(cm(2) day) in the first and to 0.24 mg pyrite/(cm(2) day) in the following reactor stages. Kinetic models taking into account adsorption/desorption as well as growth kinetics failed to describe the observed reaction rates. However, a model treating pyrite degradation and microbial growth kinetics formalistically seems to be applicable when backmixing between the reactor stages can be avoided. The advantage of a multistage reactor in comparison to single-stage equipment was shown by calculation. To obtain a pyrite conversion of 68%, a three-stage reactor would require only 58% of the volume of single-stage equipment.Measurement of oxygen consumption rates proved to provide quickly and easily measurable parameters to observe microbial coal desulfurization in technical scale: the real oxygen consumption rate is correlated to the pyrite oxidation rate and the maximum oxygen consumption rate is correlated to the concentration of viable cells. The Y(o/s) coefficient for the amount of oxygen consumed per mass unit of pyrite oxygen was determined to approximately 0.33 in comparison to 1.0 which can be calculated from stoichiornetry. This could yet not be explained. Chemical leaching experiments as well as sulfur analyses of desulfurized coal samples showed that the microorganisms play the main role in degradation of pyrite from coal and that pyrite oxidation by ferric iron can be neglected.     

Inactivation of human arginine-143, lysine-143, and isoleucine-143 Cu,Zn superoxide dismutases by hydrogen peroxide: multiple mechanisms for inactivation

Site-specific mutants of human Cu,Zn superoxide dismutase (Cu,ZnSOD) have been prepared in which the active-site arginine at position 143 (i.e., SODR143) has been replaced by either lysine (SODK143) or isoleucine (SODI143). As reported previously (W.F. Beyer, Jr., et al. (1987) J. Biol. Chem. 262, 11182-11187), SODK143 and SODI143 have 43 and 11%, respectively, of the catalytic activity of SODR143. H2O2, at low concentrations, acts as an affinity reagent for the inactivation of SODR143. At pH 9.0 and 25 degrees C, the process is characterized by a half-saturation constant for H2O2, K50, of 5.1 mM and a maximum pseudo-first-order rate constant for inactivation, Kmax, of 0.53 min-1. At pH 11.5, the corresponding values are 0.63 mM and 1.23 min-1. The active species in the inactivation is likely HO2-, as previously found with yeast and bovine Cu,ZnSODs (see C.L. Borders, Jr., and I. Fridovich (1985) Arch. Biochem. Biophys. 241, 472-476). SODK143 is also inactivated by HO2- by an affinity mechanism, i.e., one where reversible binding of H2O2 (HO2-) is a prerequisite for inactivation. At pH values of 9.0 and 11.5, the kmax values are 0.92 and 1.08 min-1, respectively; however, the corresponding K50 values increase to 42.5 and 15.8 mM, respectively. SODI143 is also inactivated by H2O2, but no evidence for an affinity mechanism was found; instead, a second-order kinetic mechanism was observed. Inactivation of each of the three enzymes is accompanied by the loss of one histidine per subunit. At elevated concentrations of H2O2, a second nonaffinity mechanism of inactivation of both SODR143 and SODK143 was found, in which a second equivalent of H2O2 reacts with the Cu,ZnSOD.HO2- complex to give a competing second-order inactivation. It appears that the positive charge of arginine-143 plays a role in the binding of HO2- at the active site of human Cu,ZnSOD, and that replacement of the arginine by lysine gives an enzyme with a similar affinity mechanism of inactivation, but with a greatly reduced affinity for HO2-. However, replacement with isoleucine causes an entirely different mechanism of inactivation; this raises the possibility that the mechanism of enzyme catalysis of superoxide dismutation by SODI143 is also different.