Gastric supply manipulation to modulate ghrelin production and enhance vascularization to the cardia: proof of the concept in a porcine model.

Introduction. Selective embolization of the left-gastric artery (LGA) reduces levels of ghrelin and achieves significant short-term weight loss. However, embolization of the LGA would prevent the performance of bariatric procedures because the high-risk leakage area (gastroesophageal junction [GEJ]) would be devascularized. Aim. To assess an alternative vascular approach to the modulation of ghrelin levels and generate a blood flow manipulation, consequently increasing the vascular supply to the GEJ. Materials and methods. A total of 6 pigs underwent a laparoscopic clipping of the left gastroepiploic artery. Preoperative and postoperative CT angiographies were performed. Ghrelin levels were assessed perioperatively and then once per week for 3 weeks. Reactive oxygen species (ROS; expressed as ROS/mg of dry weight [DW]), mitochondria respiratory rate, and capillary lactates were assessed before and 1 hour after clipping (T0 and T1) and after 3 weeks of survival (T2), on seromuscular biopsies. A celiac trunk angiography was performed at 3 weeks. Results. Mean (±standard deviation) ghrelin levels were significantly reduced 1 hour after clipping (1902 ± 307.8 pg/mL vs 1084 ± 680.0; P = .04) and at 3 weeks (954.5 ± 473.2 pg/mL; P = .01). Mean ROS levels were statistically significantly decreased at the cardia at T2 when compared with T0 (0.018 ± 0.006 mg/DW vs 0.02957 ± 0.0096 mg/DW; P = .01) and T1 (0.0376 ± 0.008mg/DW; P = .007). Capillary lactates were significantly decreased after 3 weeks, and the mitochondria respiratory rate remained constant over time at the cardia and pylorus, showing significant regional differences. Conclusions. Manipulation of the gastric flow targeting the gastroepiploic arcade induces ghrelin reduction. An endovascular approach is currently under evaluation.

Characterization of a plant-derived peptide displaying water clarifying and antimicrobial activities

SUMMARY Drinking water is currently a scarce world resource, the preparation of which requires complex treatments that include clarification of suspended particles and disinfection. Seed extracts of Moringa oleifera Lam., a tropical tree, have been proposed as an environment- friendly alternative, due to their traditional use for the clarification of drinking water. However, the precise nature of the active components was unknown. Here, we show that recombinant or synthetic forms of a cationic seed polypeptide mediate efficient sedimentation of suspended mineral particles and bacteria. Unexpectedly, the polypeptide was also found to possesses a bactericidal activity capable of disinfecting heavily contaminated water. Furthermore, the polypeptide has been shown to efficiently kill several pathogenic bacteria, including antibiotic-resistant isolates of Pseudomona, Streptococcus and Legionella species. Structural modeling of the peptide coupled to the functional analysis of synthetic peptide derivatives delineated distinct structural determinants for the flocculation and antibacterial activities. Our results suggest that a glutamine-rich portion of the polypeptide is involved in the sedimentation process; alternatively, the antibacterial activity depends on a amphiphilic loop. Assembly of multiple copies of this loop into a branched peptide derivative strongly enhances antibacterial activity without displaying hemolytic effect. In conclusion, this polypeptide displays the unprecedented feature of combining efficient water purification and disinfectant properties indicating different molecular mechanisms involved in each case. This work not only identified the features responsible for these activities but also provides useful information that has implications for the further development of this cationic polypeptide as a potent antibacterial agent. RESUME L'eau potable est actuellement une ressource limitée dans le monde. La production d'eau propre à la consommation exige des traitements complexes, incluant la clarification des particules en suspension ainsi que sa désinfection par des additifs chimiques. Les extraits de la graine d'un arbre tropical, Moringa oleifera, sont utilisés traditionnellement en Afrique afin de clarifier l'eau. Quoique la nature exacte des composants actifs était inconnue, on a pu mettre en évidence un polypeptide cationique contenu dans ces graines, capable de sédimenter de manière efficace des particules minérales en suspension ainsi que des bactéries. Ce travail a aussi mis en évidence que ce polypeptide a une activité bactéricide, permettant une désinfection d'eau fortement contaminée. De plus, nous avons démontré que ce polypeptide est efficace contre de nombreuses souches bactériennes pathogènes, également celles résistantes aux antibiotiques comme Pseudomonas, Streptococcus et Legionella. L'analyse de la structure moléculaire de ce polypeptide, couplée à son analyse fonctionnelle a mis en évidence deux domaines structuraux distinct, un pour l'activité de floculation et l'autre pour l'activité antibactérienne. Nos résultats suggèrent que le domaine riche en glutamine est impliqué dans le processus de sédimentation et que l'activité antimicrobienne dépend d'un domaine formant une boucle amphiphilique. En ramifiant plusieurs copies de cette boucle on a pu augmenter de manière significative l'activité antibactérienne. En conclusion, nous avons pu démontrer que ce polypeptide à la capacité unique de combiner des propriétés de purification et de désinfection de l'eau, ce qui implique des mécanismes moléculaires distincts pour ces deux activités. Ce travail a permis d'identifier les domaines du polypeptide qui sont responsables de ses activités et offre une perspective pour le développement d'un nouvel agent antimicrobien.

The sigma factor AlgU (AlgT) controls exopolysaccharide production and tolerance towards desiccation and osmotic stress in the biocontrol agent Pseudomonas fluorescens CHA0.

A variety of stress situations may affect the activity and survival of plant-beneficial pseudomonads added to soil to control root diseases. This study focused on the roles of the sigma factor AlgU (synonyms, AlgT, RpoE, and sigma(22)) and the anti-sigma factor MucA in stress adaptation of the biocontrol agent Pseudomonas fluorescens CHA0. The algU-mucA-mucB gene cluster of strain CHA0 was similar to that of the pathogens Pseudomonas aeruginosa and Pseudomonas syringae. Strain CHA0 is naturally nonmucoid, whereas a mucA deletion mutant or algU-overexpressing strains were highly mucoid due to exopolysaccharide overproduction. Mucoidy strictly depended on the global regulator GacA. An algU deletion mutant was significantly more sensitive to osmotic stress than the wild-type CHA0 strain and the mucA mutant were. Expression of an algU'-'lacZ reporter fusion was induced severalfold in the wild type and in the mucA mutant upon exposure to osmotic stress, whereas a lower, noninducible level of expression was observed in the algU mutant. Overexpression of algU did not enhance tolerance towards osmotic stress. AlgU was found to be essential for tolerance of P. fluorescens towards desiccation stress in a sterile vermiculite-sand mixture and in a natural sandy loam soil. The size of the population of the algU mutant declined much more rapidly than the size of the wild-type population at soil water contents below 5%. In contrast to its role in pathogenic pseudomonads, AlgU did not contribute to tolerance of P. fluorescens towards oxidative and heat stress. In conclusion, AlgU is a crucial determinant in the adaptation of P. fluorescens to dry conditions and hyperosmolarity, two major stress factors that limit bacterial survival in the environment.

Estimating traveltimes and groundwater flow patterns using 3D time-lapse crosshole ERT imaging of electrical resistivity fluctuations induced by infiltrating river water

The infiltration of river water into aquifers is of high relevance to drinking-water production and is a key driver of biogeochemical processes in the hyporheic and riparian zone, but the distribution and quantification of the infiltrating water are difficult to determine using conventional hydrological methods (e.g., borehole logging and tracer tests). By time-lapse inverting crosshole ERT (electrical resistivity tomography) monitoring data, we imaged groundwater flow patterns driven by river water infiltrating a perialpine gravel aquifer in northeastern Switzerland. This was possible because the electrical resistivity of the infiltrating water changed during rainfall-runoff events. Our time-lapse resistivity models indicated rather complex flow patterns as a result of spatially heterogeneous bank filtration and aquifer heterogeneity. The upper part of the aquifer was most affected by the river infiltrate, and the highest groundwater velocities and possible preferential flow occurred at shallow to intermediate depths. Time series of the reconstructed resistivity models matched groundwater electrical resistivity data recorded on borehole loggers in the upper and middle parts of the aquifer, whereas the resistivity models displayed smaller variations and delayed responses with respect to the logging data. in the lower part. This study demonstrated that crosshole ERT monitoring of natural electrical resistivity variations of river infiltrate could be used to image and quantify 3D bank filtration and aquifer dynamics at a high spatial resolution.

Water, energy and early postnatal growth in preterm infants.

Non-invasive methods, including stable isotope techniques, indirect calorimetry, nutritional balance and skinfold thickness, have given a new insight into early postnatal growth in neonates. Neonates and premature infants in particular, create an unusual opportunity to study the fluid and metabolic adaptation to extrauterine life because their physical environment can be controlled, fluid and energy balance can be measured and the link between metabolism and the energetics of their postnatal growth can be assessed accurately. Thus the postnatal time course of total body water, heat production, energy cost of growth and composition of weight gain have been quantified in a series of "healthy" low-birth-weight premature infants. These results show that total body water is remarkably stable between postnatal days 3-21. Energy expenditure and heat production rates increase postnatally from mean values of 40 kcal/kg/day during the first week to 60 kcal/kg/day in the third week. An apparent energy balance deficit of 180 kcal/kg can be ascribed to premature delivery. The cost of protein metabolism is the highest energy demanding process related to growth. The fact that nitrogen balance becomes positive within 72 h after birth places the newborn in a transitional situation of dissociated balance between energy and protein metabolism during early postnatal growth: skinfold thickness, dry body mass and fat decrease, while there is a gain in protein and increase in supine length. This particular situation ends during the second postnatal week and soon thereafter the rate of weight gain matches statural growth. The goals of the following review are to summarize data on total body water and energy metabolism in premature infants and to discuss how they correlate with physiological aspects of early postnatal growth.

Effects of infused amino acids on glucose production and utilization in healthy human subjects.

Amino acids have been reported to increase endogenous glucose production in normal human subjects during hyperinsulinemia: however, controversy exists as to whether insulin-mediated glucose disposal is inhibited under these conditions. The effect of an amino acid infusion on glucose oxidation rate has so far not been determined. Substrate oxidation rates, endogenous glucose production, and [13C]glucose synthesis from [13C]bicarbonate were measured in six normal human subjects during sequential infusions of exogenous glucose and exogenous glucose with (n = 5) or without (n = 5) exogenous amino acids. Amino acids increased endogenous glucose production by 84% and [13C]glucose synthesis by 235%. Glucose oxidation estimated from indirect calorimetry decreased slightly after amino acids, but glucose oxidation estimated from [13C]glucose-13CO2 data was increased by 14%. It is concluded that gluconeogenesis is the major pathway of amino acid degradation. During amino acid administration, indirect calorimetry underestimates the true rate of glucose oxidation, whereas glucose oxidation calculated from the 13C enrichment of expired CO2 during [U-13C]glucose infusion does not. A slight stimulation of glucose oxidation during amino acid infusion, concomitant with an increased plasma insulin concentration, indicates that amino acids do not inhibit glucose oxidation.

Amplification of the housekeeping sigma factor in Pseudomonas fluorescens CHA0 enhances antibiotic production and improves biocontrol abilities.

Pseudomonas fluorescens CHA0 produces a variety of secondary metabolites, in particular the antibiotics pyoluteorin and 2,4-diacetylphloroglucinol, and protects various plants from diseases caused by soilborne pathogenic fungi. The rpoD gene encoding the housekeeping sigma factor sigma 70 of P. fluorescens was sequenced. The deduced RpoD protein showed 83% identity with RpoD of Pseudomonas aeruginosa and 67% identity with RpoD of Escherichia coli. Attempts to inactivate the single chromosomal rpoD gene of strain CHA0 were unsuccessful, indicating an essential role of this gene. When rpoD was carried by an IncP vector in strain CHA0, the production of both antibiotics was increased severalfold and, in parallel, protection of cucumber against disease caused by Pythium ultimum was improved, in comparison with strain CHA0.

Expression of a plant-derived peptide harboring water-cleaning and antimicrobial activities.

Drinking water is currently a scarce world resource, the preparation of which requires complex treatments that include clarification of suspended particles and disinfection. Seed extracts of Moringa oleifera Lam., a tropical tree, have been proposed as an environment-friendly alternative, due to their traditional use for the clarification of drinking water. However, the precise nature of the active components of the extract and whether they may be produced in recombinant form are unknown. Here we show that recombinant or synthetic forms of a cationic seed polypeptide mediate efficient sedimentation of suspended mineral particles and bacteria. Unexpectedly, the polypeptide was also found to possesses a bactericidal activity capable of disinfecting heavily contaminated water. Furthermore, the polypeptide has been shown to efficiently kill several pathogenic bacteria, including antibiotic-resistant isolates of Staphylococcus, Streptococcus, and Legionella species. Thus, this polypeptide displays the unprecedented feature of combining water purification and disinfectant properties. Identification of an active principle derived from the seed extracts points to a range of potential for drinking water treatment or skin and mucosal disinfection in clinical settings.

Dominant negative MyD88 proteins inhibit interleukin-1beta /interferon-gamma -mediated induction of nuclear factor kappa B-dependent nitrite production and apoptosis in beta cells.

Insulin-dependent diabetes mellitus is an autoimmune disease in which pancreatic islet beta cells are destroyed by a combination of immunological and inflammatory mechanisms. In particular, cytokine-induced production of nitric oxide has been shown to correlate with beta cell apoptosis and/or inhibition of insulin secretion. In the present study, we investigated whether the interleukin (IL)-1beta intracellular signal transduction pathway could be blocked by overexpression of dominant negative forms of the IL-1 receptor interacting protein MyD88. We show that overexpression of the Toll domain or the lpr mutant of MyD88 in betaTc-Tet cells decreased nuclear factor kappaB (NF-kappaB) activation upon IL-1beta and IL-1beta/interferon (IFN)-gamma stimulation. Inducible nitric oxide synthase mRNA accumulation and nitrite production, which required the simultaneous presence of IL-1beta and IFN-gamma, were also suppressed by approximately 70%, and these cells were more resistant to cytokine-induced apoptosis as compared with parental cells. The decrease in glucose-stimulated insulin secretion induced by IL-1beta and IFN-gamma was however not prevented. This was because these dysfunctions were induced by IFN-gamma alone, which decreased cellular insulin content and stimulated insulin exocytosis. These results demonstrate that IL-1beta is involved in inducible nitric oxide synthase gene expression and induction of apoptosis in mouse beta cells but does not contribute to impaired glucose-stimulated insulin secretion. Furthermore, our data show that IL-1beta cellular actions can be blocked by expression of MyD88 dominant negative proteins and, finally, that cytokine-induced beta cell secretory dysfunctions are due to the action of IFN-gamma.

Experimental calcium oxalate crystal production and dissolution by selected wood rot fungi.

Twenty-six species of white-rotting Agaricomycotina fungi (Basidiomycota) were screened for their ability to produce calcium-oxalate (CaOx) crystals in vitro. Most were able to produce CaOx crystals in malt agar medium in the absence of additional calcium. In the same medium enriched with Ca2+, all the species produced CaOx crystals (weddellite or whewellite). Hyphae of four species (Ganoderma lucidum, Polyporus ciliatus, Pycnoporus cinnabarinus, and Trametes versicolor) were found coated with crystals (weddellite/whewellite). The production of CaOx crystals during the growth phase was confirmed by an investigation of the production kinetics for six of the species considered in the initial screening (Pleurotus citrinopileatus, Pleurotus eryngii, Pleurotus ostreatus, P. cinnabarinus, Trametes suaveolens, and T. versicolor). However, the crystals produced during the growth phase disappeared from the medium over time in four of the six species (P. citrinopileatus, P. eryngii, P. cinnabarinus, and T. suaveolens). For P. cinnabarinus, the disappearance of the crystals was correlated with a decrease in the total oxalate concentration measured in the medium from 0.65 μg mm−2 (at the maximum accumulation rate) to 0.30 μg mm−2. The decrease in the CaOx concentration was correlated with a change in mycelia morphology. The oxalate dissolution capability of all the species was also tested in a medium containing calcium oxalate as the sole source of carbon (modified Schlegel medium). Three species (Agaricus blazei, Pleurotus tuberregium, and P. ciliatus) presented a dissolution halo around the growth zone. This study shows that CaOx crystal production is a widespread phenomenon in white-rot fungi, and that an excess of Ca2+ can enhance CaOx crystal production. In addition, it shows that some white-rot fungal species are capable of dissolving CaOx crystals after growth has ceased. These results highlight a diversity of responses around the production or dissolution of calcium oxalate in white-rot fungi and reveal an unexpected potential importance of fungi on the oxalate cycle in the environment.

Role of glutathione in macrophage activation: effect of cellular glutathione depletion on nitrite production and leishmanicidal activity.

We have examined the effects of two agents depleting the intracellular pool of glutathione (GSH) on macrophage activation induced by IFN-gamma + LPS, as measured by nitrite production and leishmanicidal activity. Diethylmaleate (DEM), which depletes intracellular GSH by conjugation via a reaction catalyzed by the GSH-S-transferase, strongly inhibited nitrite secretion and leishmanicidal activity when added before or at the time of addition of IFN-gamma + LPS; this inhibition was progressively lost when addition of DEM was delayed up to 10 hr. A close correlation was observed between levels of intracellular soluble GSH during activation and nitrite secretion. Inhibition was partially reversed by the addition of glutathione ethyl ester (GSH-Et). Buthionine sulfoximine (BSO), a specific inhibitor of gamma-glutamylcysteine synthetase, also inhibited macrophage activation, although to a lesser extent than DEM despite a more pronounced soluble GSH depletion. This inhibition was completely reversed by the addition of GSH-Et. DEM and BSO did not alter cell viability or PMA-triggered O2- production by activated macrophages, suggesting that the inhibitory effects observed on nitrite secretion and leishmanicidal activity were not related to a general impairment of macrophage function. DEM and BSO treatment reduced iNOS specific activity and iNOS protein in cytosolic extracts. DEM also decreased iNOS mRNA expression while BSO had no effect. Although commonly used as a GSH-depleting agent, DEM may have additional effects because it can also act as a sulhydryl reagent; BSO, on the other hand, which depletes GSH by enzymatic inhibition, has no effect on protein-bound GSH. Our results suggest that both soluble and protein-bound GSH may be important for the induction of NO synthase in IFN-gamma + LPS-activated macrophages.

The C76R transmembrane activator and calcium modulator cyclophilin ligand interactor mutation disrupts antibody production and B-cell homeostasis in heterozygous and homozygous mice.

BackgroundMutations in TNFRSF13B, the gene encoding transmembrane activator and calcium modulator cyclophilin ligand interactor (TACI), are found in 10% of patients with common variable immunodeficiency. However, the most commonly detected mutation is the heterozygous change C104R, which is also found in 0.5% to 1% of healthy subjects. The contribution of the C104R mutation to the B-cell defects observed in patients with common variable immunodeficiency therefore remains unclear.ObjectiveWe sought to define the functional consequences of the C104R mutation on B-cell function.MethodsWe performed in vitro studies of TACI C104R expression and signaling. A knock-in mouse with the equivalent mutation murine TACI (mTACI) C76R was generated as a physiologically relevant model of human disease. We examined homozygous and heterozygous C76R mutant mice alongside wild-type littermates and studied specific B-cell lineages and antibody responses to T cell-independent and T cell-dependent challenge.ResultsC104R expression and ligand binding are significantly diminished when the mutant protein is expressed in 293T cells or in patients' cell lines. This leads to defective nuclear factor κB activation, which is proportionally restored by reintroduction of wild-type TACI. Mice heterozygous and homozygous for mTACI C76R exhibit significant B-cell dysfunction with splenomegaly, marginal zone B-cell expansion, diminished immunoglobulin production and serological responses to T cell-independent antigen, and abnormal immunoglobulin synthesis.ConclusionsThese data show that the C104R mutation and its murine equivalent, C76R, can significantly disrupt TACI function, probably through haploinsufficiency. Furthermore, the heterozygous C76R mutation alone is sufficient to disturb B-cell function with lymphoproliferation and immunoglobulin production defects.

Influences of body weight, body composition, and substrate oxidation rate on resting postabsorptive glucose production and gluconeogenesis.

OBJECTIVE: To determine the influence of body weight, fat mass, and fat distribution on resting endogenous glucose production in healthy lean and overweight individuals. DESIGN: measurements were performed in the resting postabsorptive state in individuals receiving an unrestricted diet. SETTING: Institute of Physiology of Lausanne University. MEASUREMENTS: resting post absorptive glucose production, glycogenolysis and gluconeogenesis; resting energy expenditure and net substrate oxidation. RESULTS: Endogenous glucose production was positively correlated with body weight, lean body mass, energy expenditure and carbohydrate oxidation. Gluconeogenesis was positively correlated with net lipid oxidation and energy expenditure, and negatively correlated with net carbohydrate oxidation. No correlation with body fat or fat distribution was observed. CONCLUSIONS: Gluconeogenesis shows a large interindividual variability. Net lipid oxidation and not body fat appears to be a major determinant of gluconeogenesis.

Connexin40 regulates renin production and blood pressure.

Renin secretion is regulated by coordinated signaling between the various cells of the juxtaglomerular apparatus. The renin-secreting cells (RSC), which play a major role in the control of blood pressure, are coupled to each other and to endothelial cells by Connexin40 (Cx40)-containing channels. In this study, we show that Cx40 knockout (Cx40-/-) mice, but not their heterozygous littermates, are hypertensive due to the increase in the number of RSC, renin biosynthesis, and plasma renin. Treatment with the angiotensin II receptor AT1 antagonist candesartan or the angiotensin II-converting enzyme inhibitor ramipril reduced the blood pressure of the Cx40-/- mice to the same levels seen in wild-type (WT) mice. The elevated blood pressure of the knockout mice was not affected by clipping one renal artery (2K1C, renin-dependent model of hypertension) or after a high salt diet. Under these conditions, however, Cx40-/- mice showed an altered production and release of renin. The renin mRNA ratio between the clipped and the non-clipped kidney was lower in the knockout than in the WT 2K1C mice. This indicates that the response to a change in blood pressure was altered. The RSC of the Cx40-/- mice did not have a compensatory increase in the levels of either Cx43 or Cx37. Our data show that renin secretion is dependent on Cx40 and suggest the Cx40-/- mice may be a genetic model of renin-dependent hypertension.