Biomedical subjects

R Lefebvre

Publications and source records attributed to R Lefebvre.

At least 19 recordsLinked to original sources

PubMed2002

The 13C-octanoic acid breath test: validation of a new noninvasive method of measuring gastric emptying in rats.

Currently available rat models for measuring gastric emptying are hampered by the necessity to kill the animals at the end of each experiment, which makes repetitive testing impossible. We have developed and validated a noninvasive test model, adapted from the 13C-octanoic breath test in humans, for repetitive measurements of gastric emptying in rats. Male Wistar rats were trained on a fixed protocol to eat a piece of pancake doped with 1 microg 13C-octanoic acid after 12 h fasting, and to stay thereafter in cylindrical glass cages. Breath tests were performed by a fully automated system of computer-guided switching valves, which collected consecutive breath samples. All breath samples were analysed by gas chromatography and isotope mass spectrometry. The area under the curve (AUC) from the cumulative 13CO2 excretion from 0 to 6 h was determined by the trapezium method to calculate the gastric half-emptying times (t(1/2)). Inter-day variability was determined. The effect of subcutaneous or intraperitoneal injection of saline was studied. The test was further validated for pharmacological interventions by oral administration of cisapride and parenteral administration of atropine, to induce, respectively. acceleration and delay of gastric emptying. Mean gastric emptying times +/- SD of 24 rats were 119.3 +/- 28.2 min, 138.7 +/- 26.0 min, and 124.5 +/- 30.9 min on three different test days. The mean coefficient of variation of three repeated measurements in the same 24 rats was 17.5%. No significant differences were observed after subcutaneous or intraperitoneal injection of saline. In a second test series of eight rats, cisapride significantly accelerated gastric emptying (mean t(1/2) 112.7 +/- 33.1 min, P < 0.05), while atropine caused a significant delay (mean t(1/2) 205.9 +/- 24.9 min, P < 0.05) when compared to control test results (mean t(1/2) 140.7 +/- 16.7 min) in the same rats. We validated the 13C-octanoic breath test to study gastric emptying in rats. This test method obviates the necessity to kill laboratory animals and allows repetitive measurements of gastric emptying to study its physiology or pathophysiology as well as the effect of pharmacological agents.

Animals↗

PubMed2001

Multiphase transfer processes in waste rock piles producing acid mine drainage 1: Conceptual model and system characterization.

Acid mine drainage (AMD) results from the oxidation of sulfides, mainly pyrite, present in mine wastes, either mill tailings or waste rock. This is the first of two papers describing the coupled physical processes taking place in waste rock piles undergoing AMD production. Since the oxidation of pyrite involves the consumption of oxygen and the production of heat, the oxidation process initiates coupled processes of gas transfer by diffusion and convection as well as heat transfer. These processes influence the supply of oxygen that is required to sustain the oxidation process. This first paper describes a general conceptual model of the interaction of these coupled transfer processes. This general conceptual model is illustrated by the physicochemical conditions observed at two large sites where extensive characterization programs revealed widely different properties. The South Dump of the Doyon mine in Canada is permeable and has a high pyrite oxidation rate leading to high temperatures (over 65 degrees C), thus making temperature-driven air convection the main oxygen supply mechanism. The Nordhalde of the Ronnenberg mining district in Germany contains lower permeability material which is less reactive, thus leading to a more balanced contribution of gaseous diffusion and convection as oxygen supply mechanisms. The field characterization and monitoring data at these sites were thoroughly analyzed to yield two coherent sets of representative physical properties. These properties are used in the second paper as a basis for applications of numerical simulation in AMD-producing waste rock piles.

Chemical Phenomena↗

PubMed2001

Multiphase transfer processes in waste rock piles producing acid mine drainage 2. Applications of numerical simulation.

Acid mine drainage (AMD) results from the oxidation of sulfides, mainly pyrite, present in mine wastes, either mill tailings or waste rock. This is the second of two papers describing the coupled physical processes taking place in waste rock piles undergoing AMD production. Since the oxidation of pyrite involves the consumption of oxygen and the production of heat, the oxidation process initiates coupled processes of gas transfer by diffusion and convection as well as heat transfer. These processes influence the supply of oxygen that is required to sustain the oxidation process. This second paper describes a numerical simulator used to represent the interaction of these coupled transfer processes. Numerical simulations are applied to two large sites with extensive characterization programs and widely different properties and behavior that were described in the first paper. The South Dump of the Doyon mine in Canada is permeable and has a high pyrite oxidation rate, thus making temperature-driven air convection the main oxygen supply mechanism. The Nordhalde of the Ronnenberg mining district in Germany contains lower permeability material which is less reactive, thus leading to a more balanced contribution of gaseous diffusion and convection as oxygen supply mechanisms. Overall, simulations allow a coherent representation of the conditions monitored within the waste rock piles and the confirmation of their physical properties. Conceptual simulations are also carried out to illustrate the potential effect of border membranes and layered co-mingling as mitigation methods used to control AMD production in either active or future waste rock piles.

Chemical Phenomena↗

PubMed2001

Gas production and migration in landfills and geological materials.

Landfill gas, originating from the anaerobic biodegradation of the organic content of waste, consists mainly of methane and carbon dioxide, with traces of volatile organic compounds. Pressure, concentration and temperature gradients that develop within the landfill result in gas emissions to the atmosphere and in lateral migration through the surrounding soils. Environmental and safety issues associated with the landfill gas require control of off-site gas migration. The numerical model TOUGH2-LGM (Transport of Unsaturated Groundwater and Heat-Landfill Gas Migration) has been developed to simulate landfill gas production and migration processes within and beyond landfill boundaries. The model is derived from the general non-isothermal multiphase flow simulator TOUGH2, to which a new equation of state module is added. It simulates the migration of five components in partially saturated media: four fluid components (water, atmospheric air, methane and carbon dioxide) and one energy component (heat). The four fluid components are present in both the gas and liquid phases. The model incorporates gas-liquid partitioning of all fluid components by means of dissolution and volatilization. In addition to advection in the gas and liquid phase, multi-component diffusion is simulated in the gas phase. The landfill gas production rate is proportional to the organic substrate and is modeled as an exponentially decreasing function of time. The model is applied to the Montreal's CESM landfill site, which is located in a former limestone rock quarry. Existing data were used to characterize hydraulic properties of the waste and the limestone. Gas recovery data at the site were used to define the gas production model. Simulations in one and two dimensions are presented to investigate gas production and migration in the landfill, and in the surrounding limestone. The effects of a gas recovery well and landfill cover on gas migration are also discussed.

Biodegradation, Environmental↗

PubMed2001

Role of nitric oxide during swallow-induced esophageal shortening in cats.

Swallowing induces esophageal shortening due to contraction of the longitudinal muscle (LM) layer. Experiments in the opossum have shown an excitatory effect of nitric oxide (NO) on esophageal LM strips. We evaluated the role of NO in swallow-induced esophageal shortening and assessed the effect of NO in vitro on feline LM strips. Swallow-induced esophageal shortening was studied before and after NO synthase blockade with L-NAME. In five cats esophageal shortening was measured using two endoscopically affixed mucosal clips. In another five cats LM contraction was measured by a strain gauge sutured on the serosal side at 2 cm above the LES; muscle strips from that region were obtained for in vitro studies. Swallowing induced esophageal shortening of 48.3+/-8.3% and LM contraction of 4.4+/-0.8 g in the control period and 32.1+/-8% and 3.0+/-0.4 g after L-NAME (P < 0.05). Nitric oxide and SNP did not change the basal tone of esophageal LM strips but provoked inhibition of metacholine-induced tonic and phasic activity. Electrical field stimulation induced frequency-dependent contractions that were reduced by atropine without further reduction after L-NAME. In conclusion, the reduction of esophageal shortening after L-NAME during the in vivo experiments suggested an excitatory effect of NO on the feline esophagus. The in vitro experiments, however, showed no contractile effect of NO or SNP on LM strips, but an inhibitory effect on the precontracted tissue. The influence of NO synthase blockade on in vivo esophageal LM shortening might be secondary to its effect on circular muscle contractility.

Animals↗

PubMed2001

Fat absorption and metabolism in bile duct ligated rats.

BACKGROUND: Bile excretion is obstructed in children with extrahepatic bile duct atresia (EHBA) resulting in fat malabsorption and disturbed lipid metabolism. AIM: Investigate if the bile duct ligated rat exhibits similar deviations as patients with EHBA under different feeding conditions. METHODS: 6 bile duct ligated Wistar rats and 12 matched paired controls were randomised over 3 feeding groups. Rats were killed 16 or 30 days postsurgery. Faeces, blood and livers were collected. Fat absorption was evaluated, markers for cholestasis and the fatty acid composition of serum phospholipids (PL) and cholesterol esters (CE) were determined. Fatty acid desaturation activities in liver microsomes were measured. RESULTS: Cholestatic bile duct ligated rats have a lower fat absorption coefficient and a lower fraction of 18:2n-6 and 18:3n-3 in serum triglycerides than their controls. This demonstrates that bile duct ligated rats suffer from fat malabsorption. In contrast to the observations in serum triglycerides, 18:2n-6 and 18:3n-3 were not reduced in serum PL and CE of cholestatic rats. Overflow of 18:2n-6 rich biliary PL in the general circulation could contribute to this observation. In agreement with what was found in man, serum PL of cholestatic rats have a higher 16:0/18:0 ratio, increased monoenes and reduced unsaturated fatty acids. However, no differences were observed in microsomal desaturation activities. CONCLUSION: Cholestatic bile duct ligated rats exhibit similar deviations in serum fatty acid composition as found in patients with EHBA, therefore they can be used as a model for this human disease.

Animals↗

PubMed1999

Postnatal development of nitrergic neurons in the myenteric plexus of rat stomach.

The effect of age on the proportion of nicotinamide adenine dinucleotide phosphate diaphorase (NADPHd)-positive neurons was investigated in the myenteric plexus of five different gastric areas of 1-day-, 1-week-, 2-week-, 1-month- and 2-month-old rats. Protein gene product 9.5 immunocytochemistry was used as a marker for the total enteric neuron population in order to establish the percentage of gastric nitrergic neurons in relation to age. The percentage of NADPHd-positive neurons in the proximal parts of the rat stomach (34-38%) is significantly higher than in the antral part (29%). This difference persists in all the age groups investigated. No significant relative increase with age of NADPHd-positive neurons could be observed in any of the areas studied. These findings imply that the increased nitrergic response in the rat proximal stomach as seen in pharmacological studies cannot be explained by an increased relative number of nitrergic neurons.

Animals↗

PubMed1997

Distribution of mucus and sperm in bovine oviducts after artificial insemination: the physical environment of the oviductal sperm reservoir.

There is a sperm reservoir in the caudal oviduct of cattle and other mammals. We had observed trapping of sperm by mucus produced by explants of bovine oviductal epithelium in vitro; therefore, we used techniques designed for preserving mucus and luminal dimensions to determine whether mucus is associated with the reservoir in vivo. Heifers were synchronized by prostaglandin injections and inseminated during estrus. Oviducts on the side of ovulation were surgically removed either 8-10 h after insemination (preovulatory) or 50-55 h after insemination (postovulatory). Segments (1 cm) taken from the uterotubal junction (UTJ), caudal 3 cm of isthmus, and mid ampulla were snap frozen. Frozen sections were coated with collodion and postfixed in phosphate-buffered formaldehyde containing cetyl-pyridinium chloride. Sections were alternately stained with periodic acid-Schiff stain (PAS) or alcian blue/PAS. Most of the oviductal lumen was highly branched with passages that measured only a few microns across and were filled with mucus. In limited areas, the lumen opened to 100 microm across and was only lightly stained for mucus. Overall, the lumen was much narrower than in sections prepared by standard fixation and paraffin embedding. Sperm were found scattered throughout the lumen of the UTJ and isthmus, in both the narrow, deeply stained luminal areas and the wider, lightly stained areas. The numbers tapered off cranially, especially prior to ovulation. In conclusion, the combination of narrow passages and mucus would appear to impede sperm progress, contributing to the creation of a reservoir.

Alcian Blue↗

PubMed1997

Bovine sperm binding to oviductal epithelium involves fucose recognition.

Sperm binding to oviductal epithelium probably serves to form the isthmic sperm reservoir. This interaction of sperm and oviductal epithelium may involve species-specific carbohydrate recognition. We tested a series of carbohydrates and glycoproteins for inhibition of bovine sperm binding to oviductal epithelium in vitro. Explants of isthmic and ampullar epithelium were obtained from oviducts that had been surgically removed from preovulatory heifers. The explants were incubated (39 degrees C, 5% CO2) with fetuin, asialofetuin, ovalbumin, fucoidan, fucose, N-acetyl glucosamine, or N-acetyl glucosamine sulfate dissolved in a modified Tyrode's balanced salt solution, termed sperm-TALP (pH 7.4, 295 mOsm) for 10 min before frozen-thawed motile sperm obtained by swim-up were added. After 15 min, the explants were rinsed, and sperm binding density was evaluated. Oviductal explants treated with fucoidan (3 mg/ml; p < 0.001, n = 5) or fucose (31 mM, p < 0.01, n = 6) had reduced densities of bound sperm compared to the controls. Incubation of explants in increasing concentrations of fucose (4-62 mM) resulted in increased inhibition of sperm binding. Pretreating explants with fucosidase also reduced sperm binding (p < 0.001, n = 3) compared to that in controls containing the fucosidase inhibitor deoxyfuconojirimycin. The presence of fucosylated molecules on the surface of the oviductal epithelium was confirmed by labeling with fucose lectins from Ulex europeus and Lotus tetragonolobus. We conclude that fucose is involved in a specific interaction between bovine sperm and oviductal epithelium.

Animals↗

PubMed1996

Effect of capacitation on bull sperm binding to homologous oviductal epithelium.

Sperm binding to oviductal epithelium is thought to be an important mechanism regulating sperm reservoir formation in the oviduct. On the basis of evidence in the hamster, we hypothesized that capacitation affects release of bovine sperm, allowing them to fertilize. Oviducts were obtained from the ovulatory side of estrous Holstein heifers. The isthmic and ampullar epithelia were milked out and reduced to fragments, which formed everted vesicles (explants). Explants were placed in tissue culture wells in TALP medium and incubated at 39 degrees C in 5% CO2. Frozen-thawed sperm were prepared by swim-up in TALP and capacitated by incubation for 4 h in TALP with 20 micrograms/ml heparin (without glucose). Uncapacitated sperm were used immediately after dilution into capacitation medium. Within 2 h of surgery, sperm were added to the explants and incubated with them for 15 min. Sperm and explants were videotaped, and the tapes were analyzed to determine the numbers of sperm bound per surface area. ANOVA did not show a difference between the number of sperm bound/0.1 mm2 in the isthmus and ampulla (p > 0.05); however, an effect of capacitation was detected (p = 0.0015). Also, the percentage of capacitated sperm, determined by chlortetracycline labeling, was greater in sperm that remained free-swimming in the presence of explants than in the absence of explants (p = 0.001). In conclusion, capacitation appears to be involved in the release of bovine sperm from oviductal epithelium and therefore could enable sperm to leave the reservoir and fertilize oocytes.

Animals↗

PubMed1995

Carbohydrates mediate the adherence of hamster sperm to oviductal epithelium.

The lower isthmus of the mammalian oviduct appears to serve as a reservoir for sperm that are retained by adherence to the epithelium. By inhibiting sperm binding within excised hamster oviducts and making use of carbohydrate probes, we have characterized the adherence of sperm in the reservoir and established a potential biochemical mechanism for the adherence and release of sperm. Fetuin and its terminal sugar, N-acetylneuraminic acid, interfered with the adherence of sperm to the oviductal epithelium. Labeled fetuin bound to the acrosomal region of fresh epididymal sperm, but not hyperactivated sperm, which have previously been reported to release from epithelial binding. Western blots labeled with fetuin and sialic acid-recognizing lectins identified proteins at several molecular masses that are candidates for the sperm surface component involved in adherence. Labeling of some of these candidates was reduced in samples from hyperactivated sperm. These results indicate that a sialylated oligosaccharide similar to that found on fetuin may be recognized by a sperm surface component and mediate binding to the oviductal epithelium. Release may be accomplished by loss or modification of the component during capacitation or hyperactivation.

Acrosome↗

PubMed1995

Characterization of the oviductal sperm reservoir in cattle.

A reservoir for sperm has been found in the oviductal isthmus in several species. Sperm are apparently trapped in the reservoir by binding to the oviductal epithelium, although other factors may be involved. We hypothesized that binding sites for bovine sperm are limited to the isthmus and are regulated by the hormonal state of the cow. Ipsilateral oviducts were obtained from heifers that were preovulatory (in estrus), had ovulated recently (within 12 h), or were in diestrus (Day 10). The isthmic and the ampullar epithelium were milked out and incubated separately in serum-free (SFRE-199-2) medium, at 39 degrees C in 5% CO2. Frozen-thawed sperm from bulls were added to the epithelium and coincubated for 15 min. The number of spermatozoa that bound to explants was not affected by stage of cycle or by anatomic origin of the explants (p > 0.05). In an additional experiment, oviducts were infused with sperm in vivo and then prepared for scanning electron microscopy, which revealed that sperm were associated with ciliated epithelium in both the isthmus and ampulla. Thus, bovine sperm may form a reservoir in the isthmic end of the oviduct because it is the first oviductal region that they encounter.

Animals↗

PubMed1994

Nitric oxide induces acetylcholine-mediated contractions in the guinea-pig small intestine.

In longitudinal muscle/myenteric plexus preparations of the guinea-pig ileum, exogenous nitric oxide (NO) induced concentration-dependent relaxations. In tissues at basal tone, NO (3 x 10(-6) M) induced a moderate relaxation followed by a pronounced contraction, consisting of a quick and sustained component. Tetrodotoxin (5 x 10(-7) M) abolished both phases of the contraction. Atropine (5 x 10(-7) M) abolished the quick component and reduced the sustained component of the contraction; the latter was further suppressed by the selective NK1 receptor antagonist CP 96,345. Hexamethonium (5 x 10(-5) M) failed to affect the contractile response to NO. It is concluded that administration of exogenous NO in the guinea-pig ileum can lead to activation cholinergic and to a lesser degree tachykininergic neurones.

Acetylcholine↗

PubMed1994