Effect of Voluntary Exercise on Genetically Obese Cpefat/fat Mice: Quantitative Proteomics of Serum

Objective: To compare the effect of voluntary exercise on body weight, food consumption, and levels of serum proteins between wild‐type and carboxypeptidase E‐deficient (Cpe^fat/fat) mice. Research Methods and Procedures: Study 1 consisted of three groups of female mice: Cpe^fat/fat mice with continuous access to exercise wheels for 3 weeks (n = 4); wild‐type C57BKS mice with access to exercise wheels for 3 weeks (n = 4); and sedentary Cpe^fat/fat mice (n = 3). Activity, body weight, and food consumption were monitored for this period and a subsequent 9‐week period without exercise wheels. Study 2 consisted of four groups of male mice (n = 6 to 7 each): Cpe^fat/fat mice with exercise wheels, wild‐type mice with exercise wheels, and Cpe^fat/fat and wild‐type mice without exercise wheels. Body weight and food consumption were measured over 4 weeks. Sera were collected, and the protein profile was determined by 2‐dimensional gel electrophoresis and mass spectrometry. Results: Cpe^fat/fat mice were moderately hyperphagic but lost weight during the initial exercise period because of greater energy expenditure. The effect of exercise was temporary, and the mice gained weight after the second week. Several serum proteins were found to be altered by exercise: haptoglobin was decreased by exercise in Cpe^fat/fat mice, and several kallikreins were increased by exercise in wild‐type mice. Discussion: The access to exercise wheels provided an initial weight loss in Cpe^fat/fat mice, but this effect was offset by elevated food consumption. The serum proteomics results indicated that Cpe^fat/fat and wild‐type mice differed in their response to exercise.     

Identification of coliform genera recovered from water using different technologies

Aims:  Methods for the detection of coliforms in water have changed significantly in recent years with procedures incorporating substrates for the detection of β- d -galactosidase becoming more widely used. This study was undertaken to determine the range of coliform genera detected with methods that rely on lactose fermentation and compare them to those recovered using methods based upon β- d -galactosidase.
Methods and Results:  Coliform isolates were recovered from sewage-polluted water using m-endo, membrane lauryl sulfate broth, tergitol TTC agar, Colilert-18^®, ChromoCult^® and ColiScan^® for primary isolation. Organisms were grouped according to whether they had been isolated based upon lactose fermentation or β- d -galactosidase production.
Conclusions:  A wide range of coliform genera were detected using both types of methods. There was considerable overlap between the two groups, and whilst differences were seen between the genera isolated with the two method types, no clear pattern emerged. Substantial numbers of 'new' coliforms (e.g. Raoutella spp.) were recovered using both types of methods.
Significance and Impact of the Study:  The results presented here confirm that both methods based on lactose fermentation or detection of β- d -galactosidase activity recover a range of coliform organisms. Any suggestion that only methods which are based upon fermentation of lactose recover organisms of public health or regulatory significance cannot be substantiated. Furthermore, the higher recovery of coliform organisms from sewage-polluted water using methods utilizing β- d -galactosidase-based methods does not appear to be because of the recovery of substantially more 'new' coliforms.     

Study of haemolytic activity of some Campylobacter spp. on blood agar plates

A total of 152 strains of Campylobacter jejuni, C. coli, C. laridis and C. fetus subsp. fetus were tested for haemolysis on blood agar plates. Distinct haemolysis was detected in 92.% (96/104) of strains of C. jejuni and 21.7% (5/23) of strains of C. coli on sheep blood heart infusion agar after incubation for 4 d microacrobically at 42°C. Haemolysis was also detected on horse blood heart infusion agar. Haemolysis was not detected at 37°C except with one of 50 strains of C. jejuni tested at this temperature, which was weakly positive. Campylobacter laridis was not haemolytic; C. fetus subsp. fetus , which does not grow at 42°C, showed no haemolysis at 37°C. Blood agar (Oxoid, BA Base No. 2) was not suitable for testing for haemolysis by these organisms. A microaerobic gas mixture containing hydrogen is better than that containing nitrogen because the medium has a brighter colour, making haemolysis casier to detect. There was no synergistic haemolysis with Staphylococcus aureus or Streptococcus agalactiae . The plate haemolysis test as described here may aid differentiation within the thermophilic campylobacters.     

Inhibition of matrix metalloproteinase-9 secretion by dimethyl sulfoxide and cyclic adenosine monophosphate in human monocytes

BACKGROUND Matrix metalloproteinases(MMPs),including MMP-9,are an integral part of the immune response and are upregulated in response to a variety of stimuli.New details continue to emerge concerning the mechanistic and regulatory pathways that mediate MMP-9 secretion.There is significant evidence for regulation of inflammation by dimethyl sulfoxide(DMSO)and 3',5'-cyclic adenosine monophosphate(cAMP),thus investigation of how these two molecules may regulate both MMP-9 and tumor necrosis factorα(TNFα)secretion by human monocytes was of high interest.The hypothesis tested in this study was that DMSO and cAMP regulate MMP-9 and TNFαsecretion by distinct mechanisms.AIM To investigate the regulation of lipopolysaccharide(LPS)-stimulated MMP-9 and tumor necrosis factorαsecretion in THP-1 human monocytes by dimethyl sulfoxide and cAMP.METHODS The paper describes a basic research study using THP-1 human monocyte cells.All experiments were conducted at the University of Missouri-St.Louis in the Department of Chemistry and Biochemistry.Human monocyte cells were grown,cultured,and prepared for experiments in the University of Missouri-St.Louis Cell Culture Facility as per accepted guidelines.Cells were treated with LPS for selected exposure times and the conditioned medium was collected for analysis of MMP-9 and TNFαproduction.Inhibitors including DMSO,cAMP regulators,and anti-TNFαantibody were added to the cells prior to LPS treatment.MMP-9 secretion was analyzed by gel electrophoresis/western blot and quantitated by ImageJ software.TNFαsecretion was analyzed by enzyme-linked immuno sorbent assay.All data is presented as the average and standard error for at least 3 trials.Statistical analysis was done using a two-tailed paired Student t-test.P values less than 0.05 were considered significant and designated as such in the Figures.LPS and cAMP regulators were from Sigma-Aldrich,MMP-9 standard and antibody and TNFαantibodies were from R&D Systems,and amyloid-βpeptide was from rPeptide.RESULTS In our investigation of MMP-9 secretion from THP-1 human monocytes,we made the following findings.Inclusion of DMSO in the cell treatment inhibited LPSinduced MMP-9,but not TNFα,secretion.Inclusion of DMSO in the cell treatment at different concentrations inhibited LPS-induced MMP-9 secretion in a dosedependent fashion.A cell-permeable cAMP analog,dibutyryl cAMP,inhibited both LPS-induced MMP-9 and TNFαsecretion.Pretreatment of the cells with the adenylyl cyclase activator forskolin inhibited LPS-induced MMP-9 and TNFαsecretion.Pretreatment of the cells with the general cAMP phosphodiesterase inhibitor IBMX reduced LPS-induced MMP-9 and TNFαin a dose-dependent fashion.Pre-treatment of monocytes with an anti-TNFαantibody blocked LPSinduced MMP-9 and TNFαsecretion.Amyloid-βpeptide induced MMP-9 secretion,which occurred much later than TNFαsecretion.The latter two findings strongly suggested an upstream role for TNFαin mediating LPS-stimulate MMP-9 secretion.CONCLUSION The cumulative data indicated that MMP-9 secretion was a distinct process from TNFαsecretion and occurred downstream.First,DMSO inhibited MMP-9,but not TNFα,suggesting that the MMP-9 secretion process was selectively altered.Second,cAMP inhibited both MMP-9 and TNFαwith a similar potency,but at different monocyte cell exposure time points.The pattern of cAMP inhibition for these two molecules suggested that MMP-9 secretion lies downstream of TNFαand that TNFαmay a key component of the pathway leading to MMP-9 secretion.This temporal relationship fit a model whereby early TNFαsecretion directly led to later MMP-9 secretion.Lastly,antibody-blocking of TNFαdiminished MMP-9 secretion,suggesting a direct link between TNFαsecretion and MMP-9 secretion.     

Saprotrophic cord systems: dispersal mechanisms in space and time

In natural terrestrial environments, nutrients are often patchily and sparsely distributed, and the microclimate is constantly changing both temporally and spatially. To survive, fungi must be able to transfer to a new resource before the nutrient supplies in their current food base are exhausted. While the majority of fungi propagate as spores, some basidiomycetes can grow out of a resource as mycelium in search of new resources. The mycelium of these fungi typically aggregates to form linear organs, termed cords or rhizomorphs, that ramify at the soil-litter interface in forests, interconnecting disparate litter components to form extensive (many square meters or even hectares), long-lived (many years) systems. These mycelial systems form effective dispersal mechanisms in space and time. This article reviews the two main, but not mutually exclusive, mycelial dispersal (resource capture) strategies: (1) a “sit and wait” strategy, whereby a large mycelial network waits for resources to land on it and then actively colonises those resources; and (2) growing and searching actively for new resources. The way in which mycelia balance exploration and nutrient transport, and robustness to damage, against “cost” of production and speed with which an area can be colonised, is explored using techniques borrowed from graph theory and statistical mechanics.