Circadian rhythm and redox homeostasis candidate genes showed association with shallow elevation in Norway spruce.

The analysis of genetic variation underlying local adaptation in natural populations, together with the response to different external stimuli, is currently a hot topic in forest sciences, with the aim of identifying genetic markers controlling key phenotypic traits of interest for their inclusion in restoration and breeding programs. In Europe, one of the main tree species is Norway spruce (Picea abies (L.) H.Karst.). Using the MassARRAY® platform, 568 trees from North Rhine-Westphalia (Germany) were genotyped with 94 single nucleotide polymorphisms (SNPs) related to circadian and growth rhythms, and to stress response. The association analysis of the selected markers with health status and elevation was performed using three different methods, and those identified by at least two of these were considered as high confidence associated SNPs. While just five markers showed a weak association with health condition, 32 SNPs were correlated with elevation, six of which were considered as high confidence associated SNPs, as indicated by at least two different association methods. Among these genes, thioredoxin and pseudo response regulator 1 (PRR1) are involved in redox homeostasis and ROS detoxification, APETALA2-like 3 (AP2L3), a transcription factor, is involved in seasonal apical growth, and a RPS2-like is a disease resistance gene. The function of some of these genes in controlling light-dependent reactions and metabolic processes suggests signatures of adaptation to local photoperiod and the synchronization of the circadian rhythm. This work provides new insights into the genetic basis of local adaptation over a shallow elevation gradient in Norway spruce.

Mortality and pathology of hybrid catfish, Clarias macrocephalus (Günther) × Clarias gariepinus (Burchell), associated with Edwardsiella ictaluri infection in southern Thailand.

Enteric septicaemia of catfish (ESC) caused by Edwardsiella ictaluri is becoming an increasing problem in aquaculture and has been reported worldwide in a variety of fish species. This study reports ESC in hybrid catfish, Clarias macrocephalus (Günther) × Clarias gariepinus (Burchell), cultured in southern Thailand. The bacteria were identified as E. ictaluri by conventional and rapid identification systems, as well as by genetic and phylogenetic characterization. Analysis of 16S rRNA indicated 100% homology to the 16S rRNA sequence of several E. ictaluri strains in GenBank. Plasmid profiles demonstrated 4.0- and 5.6-kb plasmids, compared with the 4.8- and 5.6-kb plasmids in the US isolates, and representative genes of three of the four known pathogenicity islands of US isolates were present. Serologically, lipopolysaccharide (LPS) purified from the Thai isolates was not recognized by a monoclonal antibody against the LPS of US isolates. Fish experimentally infected with E. ictaluri showed 23-100% mortality within 14 days with a 168-h LD50 of 6.92 × 10(7)  CFU mL(-1) by immersion and a 96-h LD50 of 1.58 × 10(6)  CFU fish(-1) by intraperitoneal injection. Examination of tissue sections obtained from both naturally and experimentally infected fish indicated that infection of hybrid catfish with E. ictaluri produced lesions in several organs including liver, kidney, spleen, heart and brain. Histopathology findings included cellular necrosis, focal haemorrhage, infiltration of lymphocytes and multifocal granulomatous inflammation in the infected organs.

Molecular and quantitative signatures of biparental inbreeding depression in the self-incompatible tree species Prunus avium.

Genetic diversity strongly influences populations' adaptability to changing environments and therefore survival. Sustainable forest management practices have multiple roles including conservation of genetic resources and timber production. In this study, we aimed at better understanding the variation in genetic diversity among adult and offspring individuals, and the effects of mating system on offspring survival and growth in wild cherry, Prunus avium. We analysed adult trees and open pollinated seed-families from three stands in Germany at eight microsatellite loci and one incompatibility system locus and conducted paternity analyses. Seed viability testing and seed sowing in a nursery allowed further testing for the effects of pollen donor diversity and genetic similarity between mates on the offspring performance at the seed and seedling stages. Our results were contrasting across stands. Loss of genetic diversity from adult to seedling stages and positive effect of mate diversity on offspring performance occurred in one stand only, whereas biparental inbreeding depression and significant decrease in fixation index from adults to seedlings was detected in two stands. We discussed the effects of stand genetic diversity on the magnitude of biparental inbreeding depression at several life-stages and its consequences on the management of genetic resources in P. avium.

Interaction-induced spin polarization in quantum dots.

The electronic states of lateral many-electron quantum dots in high magnetic fields are analyzed in terms of energy and spin. In a regime with two Landau levels in the dot, several Coulomb-blockade peaks are measured. A zigzag pattern is found as it is known from the Fock-Darwin spectrum. However, only data from Landau level 0 show the typical spin-induced bimodality, whereas features from Landau level 1 cannot be explained with the Fock-Darwin picture. Instead, by including the interaction effects within spin-density-functional theory a good agreement between experiment and theory is obtained. The absence of bimodality on Landau level 1 is found to be due to strong spin polarization.

Multiple transitions of the spin configuration in quantum dots.

Single electron tunneling is studied in a many electron quantum dot in high magnetic fields. For such a system multiple transitions of the spin configuration are theoretically predicted. With a combination of spin blockade and Kondo effect we are able to detect five regions with different spin configurations. Transitions are induced with changing electron numbers.

Clinical experience with the 3F stentless aortic bioprosthesis: one-year follow up.

BACKGROUND AND AIM OF THE STUDY: The 3F equine aortic bioprosthesis is a new stentless valve which is currently undergoing evaluation in a multicenter clinical trial and is considered to have superior hemodynamic performance. Herein is reported the authors' initial experience with the 3F valve, focusing on the hemodynamic performance of the device. METHODS: Between March 2002 and October 2003, 35 patients (age range 61-86 years) underwent aortic valve replacement with the 3F valve implanted in the subcoronary position. Evaluation of valve hemodynamic performance was assessed at discharge (postoperative day 5 +/- 2), at 3-6 months, and at 11-14 months' follow up by means of transthoracic echocardiography. The mean preoperative transvalvular pressure gradient was 63 +/- 14 mmHg. Before surgery, all patients were in NYHA class III or IV, despite aggressive medical treatment. Five patients underwent concomitant coronary artery bypass grafting. RESULTS: The implanted valve sizes ranged from 21 to 29 mm. There were no operative deaths and no major perioperative complications. After 12 months, mean pressure gradients for the 23-, 25-, 27- and 29-mm valves were 13, 13, 12 and 9 mmHg, respectively. Mean systolic pressure gradients and NYHA class were improved significantly after surgery. Mild signs of hemolysis and minimal central regurgitation were detected in some patients, but were of no clinical importance. Valve dysfunction or mechanical failure did not occur during the follow up period. CONCLUSION: Because of its flexible structure, the 3F aortic bioprosthesis is simple to implant, and no major adverse effects have been associated with such valve implantation at the authors' center. Transprosthetic gradients appeared to regress at 6-12 months' follow up. The durability of the device is yet to be established in ongoing long-term trials.

Bioavailability and relative tissue distribution of [125I]-recombinant human thrombin following intravenous or subcutaneous administration to non-human primates.

BACKGROUND: Recombinant human thrombin (rhThrombin) is being developed as a general adjunct to hemostasis. Endogenous thrombin is rapidly inactivated by complex formation with antithrombin III and other inhibitors. It follows that these inhibitors will also inactivate any rhThrombin that reaches the systemic circulation. OBJECTIVES: Study goals were to determine the pharmacokinetic characteristics of [(125)I]-rhThrombin and [(125)I]-rhThrombin complexed to endogenous inhibitors, and the tissue distribution of rhThrombin-associated radioactivity in non-human primates. Hematology, serum chemistry and coagulation status were also monitored. METHODS: [(125)I]-rhThrombin was administered intravenously (i.v.; 3.5 U kg(-1)) or subcutaneously (s.c.; 350 U kg(-1)) to male cynomolgus monkeys. Plasma was analyzed for rhThrombin-associated radioactivity and non-compartmental analysis was used to determine the corresponding pharmacokinetic parameters. A size exclusion-high pressure liquid chromatography (SE-HPLC) method was used to quantitate rhThrombin complexes, non-complexed rhThrombin, and free [(125)I]. Whole-body gamma scintigraphy was used to follow radioactivity localization up to 72 h postdose. RESULTS: No adverse events were observed following [(125)I]-rhThrombin administration. The pharmacokinetic profile of rhThrombin-associated radioactivity following i.v. injection was multi-exponential with an initial half-life of approximately 10 min. Following both i.v. and s.c. dosing, the terminal half-life was approximately 15 h. SE-HPLC analysis revealed that rhThrombin was rapidly complexed to antithrombin III and other inhibitors in the systemic circulation following i.v. administration. Thus, rhThrombin-associated radioactivity in the blood was complexed and presumed inactive. [(125)I]-rhThrombin inhibitor complexes accumulated and were eliminated in the liver following both routes of administration. CONCLUSIONS: These data suggest that rhThrombin rapidly binds to endogenous inhibitors following either i.v. or s.c. administration.

Influenza virus host response of C57Bl/6 mice treated with TACI-Ig.

TACI-Ig is a soluble glycoprotein comprised of a human IgG1-Fc fused with the extracellular domain of the human TACI receptor. Chronic exposure to TACI-Ig is associated with reduced circulating B cells in mouse and non-human primates, and a concomitant decrease in circulating immunoglobulin. Because of these activities, TACI-Ig is in clinical evaluation for treatment of various autoimmune diseases and B cell malignancies. In this study, the effect of TACI-Ig treatment on the ability of C57Bl/6 mice to clear influenza virus was evaluated. C57Bl/6 mice were exposed to vehicle (negative control), dexamethasone (positive control), or TACI-Ig (0.05, 0.50, or 5.0 mg/kg, SC, thrice weekly) from within one week prior to viral exposure through 21 days thereafter. Dexamethasone treatment of influenza-infected mice prolonged the infection, and decreased survival, body weight, lymphoid organ weight, influenza-specific IgM and IgG, and viral clearance relative to control animals, consistent with its expected immunosuppressive activity. Animals treated with TACI-Ig (0.05, 0.50, and 5.0 mg/kg) demonstrated a dose-dependent decrease in spleen weight and influenza-specific IgG and IgM in both lung and serum relative to control animals. In addition, flow cytometric analyses showed a decrease in B cells, but not T cells, in peripheral blood in animals treated with TACI-Ig. However, neither viral clearance nor survival was affected by TACI-Ig treatment. These data demonstrate the expected B cell-specific pharmacological effects of TACI-Ig in influenza-challenged C57Bl/6 mice without apparent effect on influenza virus clearance. It is concluded that non-B cell related antiviral competence remains intact during TACI-Ig treatment.

Probing a Kondo-correlated quantum dot with spin spectroscopy.

We investigate the Kondo effect and spin blockade observed in a many-electron quantum dot and study the magnetic field dependence. At lower fields, a pronounced Kondo effect is found, which is replaced by the spin blockade at higher fields. In an intermediate regime, both effects are visible. We make use of this combined effect to gain information about the internal spin configuration of our quantum dot. We find that the data cannot be explained assuming regular filling of electronic orbitals. Instead, spin polarized filling seems to be probable.

Safety of recombinant human factor XIII in a cynomolgus monkey model of extracorporeal blood circulation.

Factor XIII (FXIII) is a thrombin-activated plasma coagulation factor critical for blood clot stabilization and longevity. Administration of exogenous FXIII to replenish depleted stores after major surgery, including cardiopulmonary bypass, may reduce bleeding complications and transfusion requirements. Thus, a model of extracorporeal circulation (ECC) was developed in adult male cynomolgus monkeys (Macaca fascicularis) to evaluate the nonclinical safety of recombinant human FXIII (rFXIII). The hematological and coagulation profile in study animals during and after 2 h of ECC was similar to that reported for humans during and after cardiopulmonary bypass, including observations of anemia, thrombocytopenia, and activation of coagulation and platelets. Intravenous slow bolus injection of 300 U/kg (2.1 mg/kg) or 1000 U/kg (7 mg/kg) rFXIII after 2 h of ECC was well tolerated in study animals, and was associated with a dose-dependent increase in FXIII activity. No clinically significant effects in respiration, ECG, heart rate, blood pressure, body temperature, clinical chemistry, hematology (including platelet counts), or indicators of thrombosis (thrombin:anti-thrombin complex and D-Dimer) or platelet activation (platelet factor 4 and beta-thromboglobulin) were related to rFXIII administration. Specific examination of brain, heart, lung, liver, and kidney from rFXIII-treated animals provided no evidence of histopathological alterations suggestive of subclinical hemorrhage or thrombosis. Taken as a whole, the results demonstrate the ECC model suitably replicated the clinical presentation reported for humans during and after cardiopulmonary bypass surgery, and do not suggest significant concerns regarding use of rFXIII in replacement therapy after extracorporeal circulation.

Absence of a pharmacokinetic interaction between digoxin and levofloxacin.

BACKGROUND: Levofloxacin, a broad-spectrum fluoroquinolone, may enhance digoxin bioavailability by eliminating intestinal flora that metabolize digoxin. Moreover, levofloxacin, which is eliminated primarily by glomerular filtration and active tubular secretion, may alter the elimination rate of digoxin. Because of the narrow therapeutic index of digoxin, it is important to evaluate the potential for interaction with levofloxacin when administered concomitantly. METHODS: This was a placebo-controlled, randomized, double-blind, two-phase crossover study. Twelve healthy subjects (six males and six females) received 500 mg twice/day oral doses of levofloxacin or placebo for 6 days and a single oral dose of 0.4 mg digoxin on the morning of study day 5 along with levofloxacin or placebo. RESULTS: There was no significant effect of levofloxacin on the pharmacokinetics (Cmax, AUC, and other disposition parameters) of oral digoxin. Steady-state levofloxacin absorption and disposition kinetics were also similar in the presence or absence of digoxin. CONCLUSIONS: Results of this study suggest that an important pharmacokinetic interaction between levofloxacin and digoxin is unlikely to occur when administered concomitantly.

Transforming pathophysiology instruction through narrative pedagogy and Socratic questioning.

Pathophysiology, heavily content driven, has typically been taught through the use of traditional behavioral pedagogy and a reliance on the formal lecture. The author describes the limitations of this approach to teaching pathophysiology and describes the use of narrative pedagogy and Socratic questioning as alternative methods of instruction to augment lecture methods. Specific strategies for transforming traditional classroom teaching by using Socratic questions in a pathophysiology course for nurse practitioners are described. Student and faculty reactions to the initial efforts to transform pathophysiology instruction are also described.

Pharmacokinetics/dynamics of 5c8, a monoclonal antibody to CD154 (CD40 ligand) suppression of an immune response in monkeys.

The pharmacokinetics and pharmacodynamics (PK/PD) of chimeric (Ch5c8) and humanized (Hu5c8) 5c8, a monoclonal antibody that binds CD154 (CD40 ligand), thus blocking the interaction between CD40 and CD154, were investigated in cynomolgus monkeys. Single-dose groups (n = 3 animals per dose) received saline, 0.2, 1, 5 or 20 mg/kg i.v. doses of Hu5c8. The repeat-dose groups (n = 4 animals) received 0 or 5 mg/kg i.v. doses of Ch5c8 or Hu5c8 on days 1, 2, 3, 5, 7 and 9. The single-dose PK parameters showed dose proportionality, with a terminal half-life of 300 h, a volume of distribution at steady state of 73 ml/kg and clearance of 0.2 ml.h-1.kg-1. The repeat-dose regimen produced a longer terminal half-life (500 h) and lower clearance (0.13 ml.h-1.kg-1) than in the single-dose groups. The antibody titer to tetanus toxoid (ATT) challenge served as the immunodynamic marker. The primary ATT response consisted of a latent phase of approximately 10 days, during which the immune system was processing antigen but not yet producing antibody, a rise to an antibody maximum titer at approximately 18 days and a decline toward baseline by approximately 40 days in controls. The 5c8 produced a log(dose)-proportional reduction in the area under the curve of ATT. An indirect PK/PD model based on the kinetics of tetanus toxoid exposure and inhibition of ATT production in relation to 5c8 concentrations was developed. A median inhibitory concentration of 0.84 microg/ml and a efficacy of 0.84 reflected marked inhibition of ATT response by 5c8. The model provides quantitation of reduced ATT responses after 5c8 and was applicable to primary and secondary immune responses and to both single-dose and multiple-dose treatments. The monoclonal antibody 5c8 blocks the CD40 and CD154 interaction, producing consistent and substantive reduction in antibody formation after administration of tetanus toxoid, which can be characterized with PK/PD modeling. It is anticipated that 5c8 may have utility in the treatment of antibody-mediated autoimmune disease.

Impaired bioavailability of interferon beta-1a when administered intramuscularly by needle-free injection.

Intramuscular (IM) or subcutaneous (SC) drug administration of small molecules and protein has been demonstrated by use of needle-free jet injection methods. One device that achieves needle-free parenteral administration, BIOJECTOR, is commercially available and was evaluated for IM delivery of interferon beta-1a. Recombinant human interferon beta-1a (IFN beta-1a) is a glycosylated protein containing 166 amino acids and has a molecular weight of 22.5 kDa. Needle-free jet injection of IFN beta-1a with the BIOJECTOR was assessed in a human Phase I trial. The study was a randomized, open-label crossover in which 12 healthy subjects each received 60 microg of IFN beta-1a as an IM injection by standard needle administration and by needle-free jet injection. Blood samples for pharmacokinetic (serum activity, PK) and pharmacodynamic (serum neopterin, PD) determinations were collected through 144 hours post-dose. Mean serum antiviral activity AUC values for needle-free and standard needle injection were 218 and 531 U x h/ml, respectively; corresponding C(max) values were 19.7 and 29.0 U/ml. Median T(max) following both treatments was 12 hours. The relative bioavailability of IFN beta-1a, needle-free to standard needle injection, was 41.1% with 90% confidence limits of 24.4% to 69.3%. Mean serum neopterin E(AUC) values for needle-free and standard needle injection were 114 and 325 ng x h/mL, respectively; corresponding E(max) values were 2.3 and 5.6 ng/mL. The ratio of serum neopterin E(AUC), needle-free to standard needle, was 34.9% with 90% confidence limits of 23.4% to 52.1%. Injection site reactions were substantially more frequent following needle-free injection; however, systemic side effects were less frequent. Intramuscular needle-free jet injection and needle-based injection of a 22.5-kDa glycoprotein do not produce equivalent systemic PK or PD responses.

Pharmacokinetic profile of levofloxacin following once-daily 500-milligram oral or intravenous doses.

The pharmacokinetics of once-daily oral levofloxacin (study A) or intravenous levofloxacin (study B) in 40 healthy male volunteers were investigated in two separate randomized, double-blind, parallel-design, placebo-controlled studies. Levofloxacin at 500 mg or placebo was administered orally or intravenously as a single dose on day 1; daily oral or intravenous dosing resumed on days 4 to 10. In a third study (study C), the comparability of the bioavailabilities of two oral and one intravenous levofloxacin formulations were investigated with 24 healthy male subjects in an open-label, randomized, three-way crossover study. Levofloxacin at 500 mg as a single tablet or an intravenous infusion was administered on day 1; following a 1-week washout period, subjects received the second regimen (i.e., the other oral formulation or the intravenous infusion); the third and final regimen was administered following a 1-week washout period. The concentrations of drug in plasma and urine were measured by validated high-pressure liquid chromatography methods. Pharmacokinetic parameters were estimated by noncompartmental methods. In both study A (oral levofloxacin) and study B (intravenous levofloxacin), steady state was attained within 48 h after the start of the multiple dosing on day 4. Levofloxacin pharmacokinetics were linear and predictable for the single and multiple 500-mg, once-daily oral and intravenous dosing regimens, and the values of the pharmacokinetic parameters for the oral and intravenous administrations were similar. Study C indicated that levofloxacin was rapidly and completely absorbed from the oral tablets, with mean times to the maximum concentration of drug in serum of approximately 1.5 h and mean absolute bioavailability of > or =99%. These results support the interchangeability of the oral and intravenous routes of levofloxacin administration.

Pharmacokinetics and safety of oral levofloxacin in human immunodeficiency virus-infected individuals receiving concomitant zidovudine.

This phase I, double-blind, randomized, placebo-controlled, parallel-design study was conducted to evaluate the safety and pharmacokinetics of levofloxacin in human immunodeficiency virus (HIV)-infected subjects concomitantly receiving a stable regimen of zidovudine (AZT). Sixteen HIV-infected males with CD4-cell counts ranging from 100 to 550 and not experiencing significant AZT intolerance were enrolled. Subjects received levofloxacin (350 mg of levofloxacin hemihydrate) or a placebo (eight subjects per treatment group) as a single oral dose on day 1, multiple doses every 8 h from days 3 to 9, and a single dose on day 10. On days 1 and 10, an AZT dose (100 mg) was administered concurrently with the study drug. In between these doses, AZT was administered according to the regimen used by the subject prior to entering the study up to a maximum of 500 mg/day. Plasma levofloxacin concentrations were monitored for 36 h after levofloxacin dosing on day 1, immediately prior to the morning doses on days 3 to 9, and for 72 h after dosing on day 10. Plasma AZT concentrations were monitored on day 0 for baseline (for 6 h after the AZT dose) and for 4 h after the AZT doses on days 1 and 10. Levofloxacin was rapidly absorbed (time to maximum plasma concentration, approximately 1.0 h) and extensively distributed in the body with an apparent volume of distribution of approximately 104 liters (approximately 1.34 liters/kg). Steady-state conditions on day 10 were confirmed. Pharmacokinetic profiles of levofloxacin from single doses and multiple (three-times-daily) doses were similar, with a moderate accumulation (observed day 10-to-day 1 ratio of the maximum plasma concentration, approximately 185% versus expected 169%; for the corresponding ratio of the area under the concentration-time curve from 0 to 8 h [AUC(0-8)], the values were observed 217% versus expected 169%) at steady state. Mean average steady-state peak plasma concentration, plasma levofloxacin concentration at the end of the dosing interval, AUC(0-8), terminal half-life, and total body clearance were 7.06 microg/ml, 3.62 microg/ml, 37.4 microg x h/ml, 7.2 h, and 9.4 liters/h (0.12 liters/h/kg), respectively. Pharmacokinetic profiles of levofloxacin in HIV-infected patients did not appear to be affected by the concomitant administration of AZT; nor were AZT pharmacokinetics altered by levofloxacin. Oral administration of 350 mg of levofloxacin hemihydrate every 8 h appeared to be well tolerated by the subjects. There were no apparent differences in adverse events between the two treatment groups. There were no clinically significant changes from baseline in any laboratory parameter or vital sign following treatments observed in this study. The study results suggest that there is no need for levofloxacin dosage adjustment in HIV-seropositive subjects who concomitantly receive AZT.