Splenocyte subsets in normal and protein malnourished mice after long-term exposure to cocaine or morphine

An experimental model which resembles human drug addiction was developed to study the effect of chronic drug (cocaine or morphine) administration on the immune system. As malnutrition has been associated with drug use, a low protein diet has been evaluated for its contribution to the impairment of the immune system during cocaine/morphine addiction. Female C57BL/6 mice that received a 20% or 4% casein diet were studied. Both drugs were administered intraperitoneally daily for 11 weeks and drugs were administered in increasing daily doses, beginning after 3 weeks of diet consumption. Doses of cocaine began with 5 mg/kg body weight and reached the maximum dose of 40 mg/kg/day at the fourth week. Doses of morphine gradually increased from 10 mg/kg to 75 mg/kg body weight with the maximum dose reached after 5 weeks of treatment. Cocaine administration reduced body weight, particularly in the low protein diet group, and spleen weight in protein malnourished mice. Cocaine as well as saline injected mice showed a decrease in the percentage of CD4+ CD8+ and Mac-1+ cells and an increase in B cells in the spleens of well nourished mice. Morphine-treated mice showed similar results to those observed in cocaine or saline treated mice. These results suggest that cocaine, morphine or saline injection can alter the percentage of cells that express a defined phenotype independently of the nutritional status of the subject. Moreover, the effect appears dependent on a stress mediated process.     

Simulation of field scale denitrification losses from soils under grass ley and barley

Denitrification losses from soils under barley and grass ley crops were simulated. The model, which includes the major processes determining inputs, transformations and outputs of nitrogen in arable soils, represents a scale compatible with information generally available in agricultural field research. The denitrification part of the model includes a field potential denitrification rate and functions for the effect of soil aeration status, soil temperature and soil nitrate content. Easily metabolizable organic matter is assumed not to limit denitrification. Simulated values were compared with denitrification measurements made during two growing seasons in the barley and grass ley treatments of a field experiment in central Sweden.Calibration revealed that the optimal parameter values describing the effect of soil aeration on denitrification rates were similar for both treatments. The response function derived agreed well with two data sets found in the literature. The potential denitrification rate constant, derived in the simulations, was higher for grass ley than for barley, which was consistent with the differences in overall rates of carbon and nitrogen turnover found between treatments.The simulated mean denitrification rates for the two seasons were within 20% of the mean of the measured values. However, simulated denitrification showed less temporal variability and a less skewed frequency distribution than measured denitrification. Some of the measured denitrification events not explained by the model could have been due to the stimulating effects of soil drying/wetting and freezing/thawing on microbial activity.     

The effect of estradiol valerate on follicular dynamics and superovulatory response in cows with Syncro-Mate-B implants

Two experiments were designed to evaluate the effect of estradiol valerate on follicular dynamics and superovulatory response in cows with Syncro-Mate-B (SMB)implants. In Experiment 1, 5 mg estradiol valerate (E(2)), injected at the same time as superstimulation treatments were initiated, resulted in fewer corpora lutea (CL), ova/embryos collected and fertilized ova (P<0.05) than if E(2) was administered with the SMB implant 7 days earlier. In Experiment 2, 31 beef cows and 26 Holstein cows were placed in one of four treatment groups. Group I (control) cows were superstimulated on Day 9 (estrus=Day 0). On Day 2, cows in Groups II, III, and IV received SMB and cows in Group III received E(2). On Day 9, cows in Group IV received E(2), and all cows were superstimulated with Folltropin. The number of CL did not differ (P>0.19) among groups. However, there were more follicles < 10 mm and fewer fertilized ova and transferable embryos (P<0.02) in Group IV cows. Ovarian ultrasonography revealed that the diameter of the largest follicle in Group III cows declined from Day 2 to Day 7 and subsequently increased until Day 13. In contrast, Groups I, II and IV were characterized by apparently linear growth between Days 2 and 13. Differences (P<0.05) were detected between Days 5 and 9. Mean diameter of the largest follicle was smaller for cows in Group III than for the remaining groups on Day 9. It was concluded that SMB did not adversely affect superovulatory response and that E(2) administration resulted in atresia of the antral follicles in the cows with SMB implants.     

Detection and activity of a bacteriocin produced by Leuconostoc mesenteroides.

Leuconostoc mesenteroides UL5 was found to produce a bacteriocin, referred as mesenterocin 5, active against Listeria monocytogenes strains but with no effect on several useful lactic acid bacteria. The antimicrobial substance is a protein, since its activity was completely destroyed following protease (pronase) treatment. However, it was relatively heat stable (100 degrees C for 30 min) and partially denaturated by chloroform. The inhibitory effect of the bacteriocin on sensitive bacterial strains was determined by a critical-dilution micromethod. Mutants of L. mesenteroides UL5 which had lost the capacity to produce the bacteriocin were obtained. The mutant strain was stable and phenotypically identical to parental cells and remained resistant to the bacteriocin. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis was used to detect bacteriocin activity corresponding to an apparent molecular mass of about 4.5 kDa.     

Changes in pulmonary Cu−Zn contents in superior mesenteric artery occlusion shock of rabbit

Contents of Cu and Zn of into-pulmonary blood (IPB), out-pulmonary blood (OPB), Lung tissue, and supernatant and macrophages of Lung Lavage were determined in superior mesenteric artery occlusion (SMAO) shock of rabbits. Zn of pulmonary tissue was 11.42 +/- 0.60 and 14.52 +/- 1.78 (micrograms/g wet wt) in SMAO shock and control groups, respectively. Content of Zn was found to be lower, Cu was not changed, and Cu/Zn ratio increased in lung tissue in SMAO shock. Contents of Cu and Zn in other samples were not changed. The results suggest that lower Zn in lung tissue related to acute lung injury.     

Affinity thermoprecipitation of trypsin using soybean trypsin inhibitor conjugated with a thermo-reactive polymer,poly(N-vinyl caprolactam)

The soybean trypsin inhibitor conjugate with a thermo-reactive water soluble polymer, poly(N-vinyl caprolactam), was successfully used for thermally induced affinity precipitation of trypsin. The validity of the developed procedure was proven by a model separation of trypsin from dilute solution containing a large excess of bovine serum albumin.     

Crystal structure of the Y52F/Y73F double mutant of phospholipase A2: increased hydrophobic interactions of the phenyl groups compensate for the disrupted hydrogen bonds of the tyrosines.

The enzyme phospholipase A2 (PLA2) catalyzes the hydrolysis of the sn-2 ester bond of membrane phospholipids. The highly conserved Tyr residues 52 and 73 in the enzyme form hydrogen bonds to the carboxylate group of the catalytic Asp-99. These hydrogen bonds were initially regarded as essential for the interfacial recognition and the stability of the overall catalytic network. The elimination of the hydrogen bonds involving the phenolic hydroxyl groups of the Tyr-52 and -73 by changing them to Phe lowered the stability but did not significantly affect the catalytic activity of the enzyme. The X-ray crystal structure of the double mutant Y52F/Y73F has been determined at 1.93 A resolution to study the effect of the mutation on the structure. The crystals are trigonal, space group P3(1)21, with cell parameters a = b = 46.3 A and c = 102.95 A. Intensity data were collected on a Siemens area detector, 8,024 reflections were unique with an R(sym) of 4.5% out of a total of 27,203. The structure was refined using all the unique reflections by XPLOR to a final R-factor of 18.6% for 955 protein atoms, 91 water molecules, and 1 calcium ion. The root mean square deviation for the alpha-carbon atoms between the double mutant and wild type was 0.56 A. The crystal structure revealed that four hydrogen bonds were lost in the catalytic network; three involving the tyrosines and one involving Pro-68. However, the hydrogen bonds of the catalytic triad, His-48, Asp-99, and the catalytic water, are retained. There is no additional solvent molecule at the active site to replace the missing hydroxyl groups; instead, the replacement of the phenolic OH groups by H atoms draws the Phe residues closer to the neighboring residues compared to wild type; Phe-52 moves toward His-48 and Asp-99 of the catalytic diad, and Phe-73 moves toward Met-8, both by about 0.5 A. The closing of the voids left by the OH groups increases the hydrophobic interactions compensating for the lost hydrogen bonds. The conservation of the triad hydrogen bonds and the stabilization of the active site by the increased hydrophobic interactions could explain why the double mutant has activity similar to wild type. The results indicate that the aspartyl carboxylate group of the catalytic triad can function alone without additional support from the hydrogen bonds of the two Tyr residues.     

A general approach toward the design of inhibitors of serine proteinases: inhibition of human leukocyte elastase by substituted dihydrouracils

A general approach toward the rational design of potential inhibitors of serine proteinases is described. The approach is exemplified and validated through the use of appropriate heterocyclic systems in inhibiting human leukocytes elastase (HLE).