Interaction of digoxin with antihypertensive drugs via MDR1

The multidrug transporter MDR1 (P-glycoprotein)-mediated interaction between digoxin and 29 antihypertensive drugs of various types was examined by using the MDR1 overexpressing LLC-GA5-COL150 cells, which were established by transfecting MDR1 cDNA into porcine kidney epithelial LLC-PK1 cells. These cells construct monolayers with tight junctions, and enable the evaluation of transcellular transport. The MDR1 was highly expressed on the apical membrane (urine side). The basal-to-apical and apical-to-basal transcellular transport of [3H]digoxin in LLC-GA5-COL150 cells was time- and temperature-dependent. The basal-to-apical transport of [3H]digoxin was markedly increased, whereas the apical-to-basal transport was decreased in LLC-GA5-COL150 cells, compared with the host LLC-PK1 cells, suggesting that [3H]digoxin was a substrate for MDR1. Most of the Ca2+ channel blockers used here markedly inhibited basal-to-apical transport and increased apical-to-basal transport. Exceptions were diltiazem, nifedipine and nitrendipine, which hardly showed inhibitory effects on transcellular transport of [3H]digoxin. Alpha-blocker doxazosin and beta-blocker carvedilol also inhibited transcellular transport of [3H]digoxin, but none of the angiotensin converting enzyme inhibitors and AT1 angiotensin II receptor antagonists used here were active. These observations will promote understanding of the digoxin-drug interactions resulting from their actions on MDR1, and which may aid in avoiding these unexpected effects of digoxin.     

Lethal mutagenesis in a structured environment

We analyze how lethal mutagenesis operates in a compartmentalized host. We assume that different compartments receive different amounts of mutagen and that virions can migrate among compartments. We address two main questions: (1) To what extent can refugia, i.e., compartments that receive little mutagen, prevent extinction? (2) Does migration among compartments limit the effectiveness of refugia? We find that if there is little migration, extinction has to be achieved separately in all compartments. In this case, the total dose of mutagen administered to the host needs to be so high that the mutagen is effective even in the refugia. By contrast, if migration is extensive, then lethal mutagenesis is effective as long as the average growth in all compartments is reduced to below replacement levels. The effectiveness of migration is governed by the ratio of virion replication and death rates, R₀. The smaller R₀, the less migration is necessary to neutralize refugia and the less mutagen is necessary to achieve extinction at high migration rates.     

Influence of ethoxiquin on the utilization of selenium in growing pigs

In a 2 × 3 faktorial arrangement with 30 male castrates (German Landrace; 13.7±1.7 kg b.w.), after a 2 week feeding period with a maize/torula yeast basal diet poor in selenium and without vitamin E supplementation, the effect of two ethoxiquin concentrations (0 and 150 mg/kg) und 3 selenium concentrations (0.075, 0.10 and 0.125 mg/kg) on growth, the activity of the Se dependent GSH‐Px in the erthrocytes and the Se concentrations in liver, kidney, heart and diaphragm was measured after 4 weeks on the experimental diets. Under the experimental conditions chosen ethoxiquin had no effect of any of the parameters studied. Selenium supplementation significantly increased the enzyme activity and the Se concentrations in the organes and tissues analyzed.     

Lethal gram-negative bacterial superinfection in Guinea pigs given bacitracin

Farrar, W. Edmund, Jr. (Walter Reed Army Institute of Research, Washington, D.C.), Thomas H. Kent, and Van B. Elliott. Lethal gram-negative bacterial superinfection in guinea pigs given bacitracin. J. Bacteriol. 92:496-501. 1966.-Oral administration of a single dose of bacitracin (either 2,000 or 10,000 units) was lethal to more than 80% of guinea pigs. Within the first 12 hr, there was a 2,000-fold fall in the number of gram-positive organisms in the cecum. An increase in the number of coliform bacteria in the cecum was demonstrable within 6 hr, and, by 48 hr, these organisms had increased from the normal level of less than 100 per gram to approximately 1 billion per gram. The changes in intestinal bacterial flora were associated with development of a severe cecitis, mild ileitis, and acute regional lymphadenitis. Bacteremia, primarily due to coliform bacteria, was demonstrated in approximately 40% of the animals killed between 72 and 96 hr after administration of bacitracin. Development of this disease syndrome was suppressed by the administration of neomycin and polymyxin B, nonabsorbable antibiotics effective against coliform bacteria. The lethal disease produced by bacitracin in the guinea pig is similar to that produced by penicillin.     

Lethal pneumonia in guinea pigs associated with a virus

No bacteria were observed in an epizootic of lethal pneumonia in guinea pigs. Necrotic bronchitis and bronchiolitis with basophilic intranuclear inclusion bodies in bronchial epithelial cells were characteristic. Although adenovirus infection of guinea pigs has not previously been reported, histological findings paralleled those found in adenovirus infections of other animals including man. Virus particles found by electron-microscopical examination of the lung tissue closely resembled adenoviruses. The disease seemed to have a low contagiousness, a low morbidity (about 0.7%), but an acute course and a high mortality (100%).     

A two-step drug repositioning method based on a protein-protein interaction network of genes shared by two diseases and the similarity of drugs

The present study proposed a two-step drug repositioning method based on a protein-protein interaction (PPI) network of twodiseases and the similarity of the drugs prescribed for one of the two. In the proposed method, first, lists of disease related geneswere obtained from a meta-database called Genotator. Then genes shared by a pair of diseases were sought. At the first step of themethod, if a drug having its target(s) in the PPI network, the drug was deemed a repositioning candidate. Because targets of manydrugs are still unknown, the similarities between the prescribed drugs for a specific disease were used to infer repositioningcandidates at the second step. As a first attempt, we applied the proposed method to four different types of diseases: hypertension,diabetes mellitus, Crohn disease, and autism. Some repositioning candidates were found both at the first and second steps.     

Aspects of cardiomyopathy in Copper-deficient pigs

Pigs were made copper(Cu)-deficient to evaluate cardiac function and pathology, and electrocardiography. Fifteen-day-old pigs were fed a Cu-restricted diet over an 8 wk period and compared to Cu-adequate diet-fed pigs. Cardiac effects were examined concerning gross morphometry and ultrastructure, echocardiography, and electrocardiography, as well as serum cholesterol levels. The Cu-restricted diet-fed pigs exhibited a marked deceleration of growth and lower hematocrit, hemoglobin, and liver and serum Cu concentrations compared to the Cu-adequate diet-fed pigs. The Cu-restricted diet-fed pigs developed a significantly greater heart weight:body weight ratio, along with greater diastolic measures of ventricular wall and internal dimension relative to body weight. Electrocardiography in the Cu-restricted diet-fed pigs revealed one instance of electrical alternans and an intraventricular conduction disturbance and several instances of T-wave inversion. The Cu-restricted pigs also displayed a prolonged QT interval at the closure of study. Increased mitochondrial volume density and mitochondria: myofibril volume density ratio were observed in the Cu-restricted pig electron micrographs along with excessive lipid and glycogen inclusion and focal degradation of Z-lines, intercalated disk, and sarcomeres. Copper-restriction in young pigs results in cardiac pathology and electrical disturbances. These alterations are similar to those reported for young Cu-restricted rodents. Given then that many cardiac manifestations of developed Cu-deficiency appear conserved across specie lines, the potential for human disturbances in response to severe Cu-deficiency may be plausible.     

Quercetin significantly decreased cyclosporin oral bioavailability in pigs and rats

Cyclosporin, an immunosuppressant with a narrow therapeutic window, is a substrate for both CYP3A4 and P-glycoprotein (Pgp). Quercetin is an inhibitor of CYP3A4 and a modulator of Pgp. This study aimed to measure the effect of quercetin on the absorption and disposition of cyclosporin in pigs and rats. Cyclosporin (Sandimmune, 10 mg/kg) was orally administered with and without a concomitant dose of quercetin (50 mg/kg) to pigs and rats. Cyclosporin concentrations in blood samples were determined by a specific monoclonal fluorescence polarization immunoassay. The pharmacokinetic parameters were calculated by noncompartmental analysis using WINNONLIN. A paired Student's t-test was conducted for statistical comparison. A study using the everted intestinal sac was carried out to evaluate the effect of quercetin on the function of intestinal Pgp. The coadministration of quercetin significantly decreased cyclosporin AUC(0-3) (area under the concentration-time curve from time zero to 3 h) by 56% and AUC(0-t) (area under the concentration-time curve from time zero to the last point) by 43% in pigs and rats, respectively, indicating that the coadministration of quercetin significantly decreased cyclosporin oral bioavailability. However, the inverted sac study showed that quercetin significantly inhibited the function of intestinal Pgp. It is suggested that concurrent use of quercetin or quercetin-containing dietary supplement or herbs with cyclosporin or other medications whose absorption and metabolism are mediated by Pgp and/or CYP3A4 should require close monitoring.     

Quercetin supplementation to the thawing and incubation media of boar sperm improves post-thaw sperm characteristics and the in vitro production of pig embryos

Elevated levels of reactive oxygen species can cause oxidative stress, which could lead to membrane damage, decreased fertility, and spermatozoan morphological deformities. Antioxidants can be supplemented to reduce the impacts of oxidative stress. The objective of this study was to determine the effects of supplementing quercetin (0.25, 0.50, 0.75 mM) during the thawing and incubation of frozen-thawed boar semen on spermatozoan characteristics, IVF kinetics (n = 400) and subsequent embryonic development (n = 1340). Spermatozoa were evaluated for motility, viability, and membrane lipid peroxidation levels at 0, 2, 4, 6, 8, and 10 h after thawing. Embryos were evaluated for IVF kinetics 12 h after IVF (penetration, polyspermy, male pronucleus formation, IVF efficiency) and cleavage and blastocyst formation at 48 h and 144 h after IVF, respectively. Spermatozoa supplemented with 0.25 mM quercetin had significantly higher (P < 0.05) motility (51.67±8.50 %) and percent of viable cells (61.21 ± 2.44 %) compared to all other treatments at 10 h after thawing, in addition to having significantly (P < 0.05) lower levels of hydroperoxide (3.38 ± 0.88 μM/10⁷cells). There were no differences in penetration rates and male pronucleus formation between treatment groups. Supplementation of quercetin significantly decreased (P < 0.05) polyspermy and significantly increased (P < 0.05) the percentage of embryos reaching blastocyst stage of development by 144 h after IVF compared to no supplementation. Results indicated that supplementing frozen-thawed boar semen with 0.25 mM quercetin improves sperm characteristics up to 10 h after thawing and decreases polyspermy while improving early embryonic development in pigs.     

Environmental effects on nutrient and energy metabolism in pigs

Besides the genetic predisposition and the dietary supply of nutrients, environmental factors affect the performance of the pig. Environment can affect the nutrient composition and availability of the feed. It can also influence appetite and therefore the feed intake. Furthermore, nutrient and energy utilization on the level of digestion and intermediate metabolism can depend on environmental factors. Especially the energy requirements of the pig are determined by such factors like physical activity or climate. Finally, stress or disease are interacting with the environment, nutrient and energy utilization as well as performance of the pigs.     

ATP binding to solubilized (Na+ + K+)-ATPase: The abolition of subunit-subunit interaction and the maximum weight of the nucleotide-binding unit

Membrane-bound (Na⁺ + K⁺)-ATPase from pig kidney outer medulla shows apparent heterogeneity in its ATP-binding site population when assays are carried out in the presence of K⁺. This finding has been interpreted as being due to interaction between (at least) two subunits, each containing an ATP-binding site. Treating the membrane-bound enzyme with the detergent, C₁₂E₈, has been shown to solubilize enzymatically active αβ-protomers. We show that in the dissolved enzyme all ATP-binding sites in the population are identical both in the absence and in the presence of K⁺, which would be consistent with an abolition of subunit-subunit interaction. This supports previous suggestions that enzyme solubilized by C₁₂E₈ is monomeric and that the membrane-bound enzyme is not. Differential extraction of enzyme-containing membranes with C₁₂E₈ yielded preparations with an ATP-binding capacity of up to 5.8 nmol per mg protein, measured by the method of Lowry et al. (Lowry, O.H., Rosebrough, N.J., Farr, A.L., and Randall, R.J. (1951) J. Biol. Chem. 193, 265–275), with bovine serum albumin as standard. Evidence is presented that makes it likely that preparations with an ATP-binding capacity of 7.5 nmol per mg protein (as determined by the above-mentioned assay) will be obtainable. This corresponds to an αβ-protomer molecular weight of 133 000 which approximates closely to the minimum value found in the literature for an αβ-protomer (i.e., 126 000).     

Optimizing the protein nutrition of growing-finishing pigs

Growing-finishing pigs should consume each day the minimum amounts of energy and amino acids needed for maximum lean deposition. This should optimize performance traits, carcass leanness, and N excretion. These ideal conditions are difficult to achieve under experimental or farm conditions due to the factors affecting amino acid requirements and feed intake on a daily basis. Lean deposition rate and sex are two of the major factors affecting amino acid needs. If possible, maximum lean deposition rates should be determined for each herd in order to customize feeding programs, and split-sex feeding will improve N utilization.

Amino acid requirements have been determined empirically and by the factorial method. The latter is preferred if the efficiency of use of absorbed amino acids can be accurately determined. Development of computer models will likely be needed to accomplish this. Apparent ileal digestibility of amino acids is the most practical means of estimating amino acid absorption at present, although it likely overestimates amino acid availability for some amino acids.

Crystalline amino acids can be used to improve amino acid balance and reduce excessive intake of protein which should improve feed efficiency. A portion of the high-quality protein feeds in pig diets can be replaced by synthetic amino acids without sacrificing performance, but the effects of these substitutions on carcass merit is uncertain.

Excretion of N, and the concomitant reduction of N in manure that has to be disposed of, can be manipulated nutritionally by increased use of crystalline amino acids to lower dietary protein, by use of highly digestible feedstuffs and by precise matching of amino acid needs to amino acid supply. Use of these factors could lead to a reduction in total N wastes of 20–30%.     

Genotype by environment interaction for litter size in pigs as quantified by reaction norms analysis

A Bayesian procedure was used to estimate linear reaction norms (i.e. individual G × E plots) on 297 518 litter size records of 121 104 sows, daughters of 2040 sires, recorded on 144 farms in North and Latin America, Europe, Asia and Australia. The method allowed for simultaneous estimation of all parameters involved. The analysis was carried out on three subsets, comprising (i) parity 1 records of 33 641 sows of line B, (ii) all parity records of 52 120 sows of line B and (iii) all parity records of 121 104 sows of lines A, B and A × B. Estimated heritabilities ranged from 0.09 to 0.10 (smallest to largest subset) for the intercept of the reaction norms, and were 0.15, 0.08 and 0.02 (ditto) for the slope. Estimated genetic correlations between intercept and slope were -0.09, +0.26 and +0.69 (ditto). The three subsets therefore showed a progressively lower genetic component to environmental sensitivity, and progressively less re-ranking of genotypes across the environmental (herd-year-season) range. In a genetic evaluation that does not include reaction norms in the statistical model, part of the G × E effect remains confounded with the additive genetic effect, which may lead to errors in the estimates of the additive genetic effect; the reaction norms model removes this confounding. The intercept estimates from the largest data subset show correlations with litter size estimated breeding values (EBV) from routine genetic evaluation (without reaction norms included) of 0.78 to 0.85 for sows with one to seven litter records, and 0.75 for sires. Hence, including reaction norms in genetic evaluation would increase the reliability of the EBV of young selection candidates without own performance or progeny data by considerably more than 100 × (1/0.75-1) = 33%. Reaction norm slope estimates turn out to be very demanding statistics; environmental sensitivity must therefore be classified as a 'hard-to-measure' trait.     

Unilaminar follicular cells transiently express galectin-3 during ovarian folliculogenesis in pigs

The localization of galectin-3, a β-galactoside-binding animal lectin, was immunohistochemically studied in the ovaries of pigs to determine its expression in ovarian folliculogenesis. Various stages of ovarian follicles were identified in the ovaries of adult pigs. Galectin-3 was immunostained in the squamous follicular cells surrounding oocytes in primordial follicles and in the unilaminar granulosa cells of primary follicles, but not in oocytes of multilaminar follicles (including primary, secondary, and tertiary Graafian follicles). As in adult ovaries, galectin-3 immunoreactivity was prominent in the unilaminar follicles in neonatal ovaries. Galectin-3 was also immunolocalized in the luteal cells in the corpus luteum and granulosa cells of atretic follicles as well as in interstitial macrophages in porcine ovaries. Collectively, these results suggest that galectin-3 is transiently expressed in follicular cells in the unilaminar ovarian follicles (primordial and primary) but not in multilaminar ovarian follicles (primary to tertiary), implying that galectin-3 is embryologically involved in ovum generation.     

血脑屏障上P-糖蛋白与耐药性癫痫关系的研究进展

耐药性癫痫是癫痫治疗的瓶颈。P-糖蛋白通过跨膜外排泵作用阻止抗癫痫药物由血脑屏障入脑发挥期望效应是耐药性癫痫产生的重要原因。本文介绍了耐药性癫痫与血脑屏障上P-糖蛋白的相互关系,对目前提出的调节P-糖蛋白功能,改善耐药性癫痫预后应注意的问题进行说明。安全地调节P-糖蛋白功能到适度可能为耐药性癫痫的治疗带来希望。     

Non-steroidal anti-inflammatory drugs react with two sites on platelet cyclo-oxygenase evidence from ‘in vivo’ drug interaction studies in rats

Non-steroidal anti-inflammatory drugs inhibit platelet cylco-oxygenase activity. This study shows that salicylate, diflunisal, sulphinpyrazone and indomethacine prevent in vivo aspirin inhibitory effect of cyclo-oxygenase activity as measured by the formation of malondialdehyde and thromboxane B₂, two products of platelet arachidonic acid metabolism. Salicylate also prevents the inhibitory effect of indomethacin. All these drugs therefore appear to interact with the same site on platelet cyclo-oxygenase. Since salicylate is inactive by itself on this platelet enzyme and diflunisal and sulphinpyrazone were used at ineffective doses, it is suggested that their interaction with aspirin (or indomethacin) occurs at the level of a supplementary site is necessary but not sufficient for the efficacy of these drugs as cyclo-oxygenase inhibitors. Acetylation by aspirin of the active site appears to be a phenomenon secondary to the binding of salicylate moiety to the supplementary site.     

P-glycoprotein antagonists confer synergistic sensitivity to short-chain ceramide in human multidrug-resistant cancer cells

P-glycoprotein (P-gp) antagonists inhibit ceramide metabolism at the juncture of glycosylation. The purpose of this study was to test whether targeting P-gp would be a viable alternative to targeting glucosylceramide synthase (GCS) for enhancing ceramide cytotoxicity. A2780 wild-type, and multidrug-resistant 2780AD and NCI/ADR-RES human ovarian cancer cell lines and the cell-permeable ceramide analog, C6-ceramide (C6-cer), were employed. Compared to P-gp-poor A2780 cells, P-gp-rich 2780AD cells converted 3.7-fold more C6-cer to nontoxic C6-glucosylceramide (C6-GC), whereas cell-free GCS activities were equal. 2780AD cells displayed resistance to C6-cer (10 μM) that was reversed by inclusion of the P-gp antagonist tamoxifen (5 μM) but not by inclusion of a GCS inhibitor. Co-administration of C6-cer and P-gp antagonists was also effective in NCI/ADR-RES cells. For example, C6-cer, VX-710 (Biricodar), and cyclosporin A (cyc A) exposure resulted in viabilities of ~ 90% of control; however, C6-cer/VX-710 and C6-cer/cyc A additions were synergistic and resulted in viabilities of 22% and 17%, respectively. Further, whereas C6-ceramide and cyc A imparted 1.5- and 0-fold increases in caspase 3/7 activity, the combination produced a 3.5-fold increase. Although the upstream elements of cell death have not been elucidated, the novel C6-ceramide/P-gp antagonist combination merits further study and assessment of clinical translational potential.     

Serum concentrations of free amino acids in growing pigs exposed to diurnal heat stress fluctuations

In areas where the ambient temperature (AT) is above the thermo neutral (TN) zone of pigs, significant changes within a 24-h period occur, differently affecting the availability of amino acids (AA) within the same day. An experiment was conducted to analyze the serum concentrations (SC) of free AA in pigs exposed to diurnal variations in AT. Six pigs (27.1 ±1.3 kg body weight) implanted with a thermometer to register the body temperature (BT) at 15-min intervals were used. Blood samples were collected on the last 3 d of the 14-d study, at 0700 h (lowest AT), 1200 h (mild HS), and 1600 h (severe HS). The pigs received 1.2 kg/d of an AA-supplemented, wheat-soybean meal diet, in two equal meals (0700 and 1900 h). The AT and BT, recorded at 0700, 1200, and 1600 h was: 30.6, 38.6, 41.1 °C, and 38.2, 39.5, 40.3 °C, respectively. The BT was significantly correlated (P < 0.001) with the AT. The SC (μM/mL) of Ile, Lys, Met, Val, Ala, Asn, and Pro were higher (P ≤ 0.01); Arg, Phe, Glu, and Tyr tended to be higher (P ≤ 0.10); but Cys was lower (P < 0.05) at 1200 h than at 0700 h. Lys was higher, Cys and Tyr were lower (P < 0.05), and Ile and Val tended to be higher (P ≤ 0.10) at 1600 h than at 0700 h. Serum Arg, Ile, Phe, Ala, Asn, Gln, Pro, Ser, and Tyr were lower (P < 0.05), and Leu and Val tended to be lower at 1600 h than at 1200 h. These data demonstrate that AT directly alters the BT of pigs, and that diurnal variations in AT differently affect their SC and availability of AA for growth.