The humoral immune response to recombinant nucleocapsid antigen of canine distemper virus in dogs vaccinated with attenuated distemper virus or DNA encoding the nucleocapsid of wild-type virus.

This study compared the humoral immune response against the nucleocapsid-(N) protein of canine distemper virus (CDV) of dogs vaccinated with a multivalent vaccine against parvo-, adeno-, and parainfluenza virus and leptospira combined with either the attenuated CDV Onderstepoort strain (n = 15) or an expression plasmid containing the N-gene of CDV (n = 30). The vaccinations were applied intramuscularly three times at 2-week intervals beginning at the age of 6 weeks. None of the pre-immune sera recognized the recombinant N-protein, confirming the lack of maternal antibodies at this age. Immunization with DNA vaccine for CDV resulted in positive serum N-specific IgG response. However, their IgG (and IgA) titres were lower than those of CDV-vaccinated dogs. Likewise, DNA-vaccinated dogs did not show an IgM peak. There was no increase in N-specific serum IgE titres in either group. Serum titres to the other multivalent vaccine components were similar in both groups.

Introduction of Trojan sex chromosomes to boost population growth.

Conservation programs that deal with small or declining populations often aim at a rapid increase of population size to above-critical levels in order to avoid the negative effects of demographic stochasticity and genetic problems like inbreeding depression, fixation of deleterious alleles, or a general loss of genetic variability and hence of evolutionary potential. In some situations, population growth is determined by the number of females available for reproduction, and manipulation of family sex ratios towards more daughters has beneficial effects. If sex determination is predominantly genetic but environmentally reversible, as is the case in many amphibia, reptiles, and fish, Trojan sex chromosomes could be introduced into populations in order to change sex ratios towards more females. We analyse the possible consequences for the introduction of XX-males (XX individuals that have been changed to phenotypic males in a XY/XX sex determination system) and ZW males, WW males, or WW females (in a ZZ/ZW sex determination system). We find that the introduction of WW individuals can be most effective for an increase of population growth, especially if the induced sex change has little or no effect on viability.

Does a shift in host plants trigger speciation in the Alpine leaf beetle Oreina speciosissima (Coleoptera, Chrysomelidae)?

Background: Within the Coleoptera, the largest order in the animal kingdom, the exclusively herbivorous Chrysomelidae are recognized as one of the most species rich beetle families. The evolutionary processes that have fueled radiation into the more than thirty-five thousand currently recognized leaf beetle species remain partly unresolved. The prominent role of leaf beetles in the insect world, their omnipresence across all terrestrial biomes and their economic importance as common agricultural pest organisms make this family particularly interesting for studying the mechanisms that drive diversification. Here we specifically focus on two ecotypes of the alpine leaf beetle Oreina speciosissima (Scop.), which have been shown to exhibit morphological differences in male genitalia roughly corresponding to the subspecies Oreina speciosissima sensu stricto and Oreina speciosissima troglodytes. In general the two ecotypes segregate along an elevation gradient and by host plants: Oreina speciosissima sensu stricto colonizes high forb vegetation at low altitude and Oreina speciosissima troglodytes is found in stone run vegetation at higher elevations. Both host plants and leaf beetles have a patchy geographical distribution. Through use of gene sequencing and genome fingerprinting (AFLP) we analyzed the genetic structure and habitat use of Oreina speciosissima populations from the Swiss Alps to examine whether the two ecotypes have a genetic basis. By investigating a wide range of altitudes and focusing on the structuring effect of habitat types, we aim to provide answers regarding the factors that drive adaptive radiation in this phytophagous leaf beetle.Results: While little phylogenetic resolution was observed based on the sequencing of four DNA regions, the topology and clustering resulting from AFLP genotyping grouped specimens according to their habitat, mostly defined by plant associations. A few specimens with intermediate morphologies clustered with one of the two ecotypes or formed separate clusters consistent with habitat differences. These results were discussed in an ecological speciation framework.Conclusions: The question of whether this case of ecological differentiation occurred in sympatry or allopatry remains open. Still, the observed pattern points towards ongoing divergence between the two ecotypes which is likely driven by a recent shift in host plant use.

Species differences in the response of liver drug-metabolizing enzymes to (S)-4-O-tolylsulfanyl-2-(4-trifluormethyl-phenoxy)-butyric acid (EMD 392949) in vivo and in vitro.

Induction of drug-metabolizing enzymes (DMEs) is highly species-specific and can lead to drug-drug interaction and toxicities. In this series of studies we tested the species specificity of the antidiabetic drug development candidate and mixed peroxisome proliferator-activated receptor (PPAR) alpha/gamma agonist (S)-4-O-tolylsulfanyl-2-(4-trifluormethyl-phenoxy)-butyric acid (EMD 392949, EMD) with regard to the induction of gene expression and activities of DMEs, their regulators, and typical PPAR target genes. EMD clearly induced PPARalpha target genes in rats in vivo and in rat hepatocytes but lacked significant induction of DMEs, except for cytochrome P450 (P450) 4A. CYP2C and CYP3A were consistently induced in livers of EMD-treated monkeys. Interestingly, classic rodent peroxisomal proliferation markers were induced in monkeys after 17 weeks but not after a 4-week treatment, a fact also observed in human hepatocytes after 72 h but not 24 h of EMD treatment. In human hepatocyte cultures, EMD showed similar gene expression profiles and induction of P450 activities as in monkeys, indicating that the monkey is predictive for human P450 induction by EMD. In addition, EMD induced a similar gene expression pattern as the PPARalpha agonist fenofibrate in primary rat and human hepatocyte cultures. In conclusion, these data showed an excellent correlation of in vivo data on DME gene expression and activity levels with results generated in hepatocyte monolayer cultures, enabling a solid estimation of human P450 induction. This study also clearly highlighted major differences between primates and rodents in the regulation of major inducible P450s, with evidence of CYP3A and CYP2C inducibility by PPARalpha agonists in monkeys and humans.

Functional late outgrowth endothelial progenitors isolated from peripheral blood of burned patients.

BACKGROUND: Bioengineered skin substitutes are increasingly considered as a useful option for the treatment of full thickness burn injury. Their viability following grafting can be enhanced by seeding the skin substitute with late outgrowth endothelial progenitor cells (EPCs). However, it is not known whether autologous EPCs can be obtained from burned patients shortly after injury. METHODS: Late outgrowth EPCs were isolated from peripheral blood sampled obtained from 10 burned patients (extent 19.6±10.3% TBSA) within the first 24h of hospital admission, and from 7 healthy subjects. Late outgrowth EPCs were phenotyped in vitro. RESULTS: In comparison with similar cells obtained from healthy subjects, growing colonies from burned patients yielded a higher percentage of EPC clones (46 versus 17%, p=0.013). Furthermore, EPCs from burned patients secreted more vascular endothelial growth factor (VEGF) into the culture medium than did their counterparts from healthy subjects (85.8±56.2 versus 17.6±14pg/mg protein, p=0.018). When injected to athymic nude mice 6h after unilateral ligation of the femoral artery, EPCs from both groups of subjects greatly accelerated the reperfusion of the ischaemic hindlimb and increased the number of vascular smooth muscle cells. CONCLUSIONS: The present study supports that, in patients with burns of moderate extension, it is feasible to obtain functional autologous late outgrowth EPCs from peripheral blood. These results constitute a strong incentive to pursue approaches based on using autotransplantation of these cells to improve the therapy of full thickness burns.

Reciprocal regulation of brain and muscle Arnt-like protein 1 and peroxisome proliferator-activated receptor alpha defines a novel positive feedback loop in the rodent liver circadian clock.

Recent evidence has emerged that peroxisome proliferator-activated receptor alpha (PPARalpha), which is largely involved in lipid metabolism, can play an important role in connecting circadian biology and metabolism. In the present study, we investigated the mechanisms by which PPARalpha influences the pacemakers acting in the central clock located in the suprachiasmatic nucleus and in the peripheral oscillator of the liver. We demonstrate that PPARalpha plays a specific role in the peripheral circadian control because it is required to maintain the circadian rhythm of the master clock gene brain and muscle Arnt-like protein 1 (bmal1) in vivo. This regulation occurs via a direct binding of PPARalpha on a potential PPARalpha response element located in the bmal1 promoter. Reversely, BMAL1 is an upstream regulator of PPARalpha gene expression. We further demonstrate that fenofibrate induces circadian rhythm of clock gene expression in cell culture and up-regulates hepatic bmal1 in vivo. Together, these results provide evidence for an additional regulatory feedback loop involving BMAL1 and PPARalpha in peripheral clocks.

Effects of brood manipulation costs on optimal sex allocation in social hymenoptera.

In eusocial Hymenoptera, queens and workers are in conflict over optimal sex allocation. Sex ratio theory, while generating predictions on the extent of this conflict under a wide range of conditions, has largely neglected the fact that worker control of investment almost certainly requires the manipulation of brood sex ratio. This manipulation is likely to incur costs, for example, if workers eliminate male larvae or rear more females as sexuals rather than workers. In this article, we present a model of sex ratio evolution under worker control that incorporates costs of brood manipulation. We assume cost to be a continuous, increasing function of the magnitude of sex ratio manipulation. We demonstrate that costs counterselect sex ratio biasing, which leads to less female-biased population sex ratios than expected on the basis of relatedness asymmetry. Furthermore, differently shaped cost functions lead to different equilibria of manipulation at the colony level. While linear and accelerating cost functions generate monomorphic equilibria, decelerating costs lead to a process of evolutionary branching and hence split sex ratios.

Local delivery of glial cell line-derived neurotrophic factor improves facial nerve regeneration after late repair.

OBJECTIVES/HYPOTHESIS: Facial nerve regeneration is limited in some clinical situations: in long grafts, by aged patients, and when the delay between nerve lesion and repair is prolonged. This deficient regeneration is due to the limited number of regenerating nerve fibers, their immaturity and the unresponsiveness of Schwann cells after a long period of denervation. This study proposes to apply glial cell line-derived neurotrophic factor (GDNF) on facial nerve grafts via nerve guidance channels to improve the regeneration. METHODS: Two situations were evaluated: immediate and delayed grafts (repair 7 months after the lesion). Each group contained three subgroups: a) graft without channel, b) graft with a channel without neurotrophic factor; and c) graft with a GDNF-releasing channel. A functional analysis was performed with clinical observation of facial nerve function, and nerve conduction study at 6 weeks. Histological analysis was performed with the count of number of myelinated fibers within the graft, and distally to the graft. Central evaluation was assessed with Fluoro-Ruby retrograde labeling and Nissl staining. RESULTS: This study showed that GDNF allowed an increase in the number and the maturation of nerve fibers, as well as the number of retrogradely labeled neurons in delayed anastomoses. On the contrary, after immediate repair, the regenerated nerves in the presence of GDNF showed inferior results compared to the other groups. CONCLUSIONS: GDNF is a potent neurotrophic factor to improve facial nerve regeneration in grafts performed several months after the nerve lesion. However, GDNF should not be used for immediate repair, as it possibly inhibits the nerve regeneration.

Genetic male infertility and mutation of CATSPER ion channels.

A clinically significant proportion of couples experience difficulty in conceiving a child. In about half of these cases male infertility is the cause and often genetic factors are involved. Despite advances in clinical diagnostics ∼50% of male infertility cases remain idiopathic. Based on this, further analysis of infertile males is required to identify new genetic factors involved in male infertility. This review focuses on cation channel of sperm (CATSPER)-related male infertility. It is based on PubMed literature searches using the keywords 'CATSPER', 'male infertility', 'male contraception', 'immunocontraception' and 'pharmacologic contraception' (publication dates from January 1979 to December 2009). Previously, contiguous gene deletions including the CATSPER2 gene implicated the sperm-specific CATSPER channel in syndromic male infertility (SMI). Recently, we identified insertion mutations of the CATSPER1 gene in families with recessively inherited nonsyndromic male infertility (NSMI). The CATSPER channel therefore represents a novel human male fertility factor. In this review we summarize the genetic and clinical data showing the role of CATSPER mutation in human forms of NSMI and SMI. In addition, we discuss clinical management and therapeutic options for these patients. Finally, we describe how the CATSPER channel could be used as a target for development of a male contraceptive.

Linezolid alone or combined with rifampin against methicillin-resistant Staphylococcus aureus in experimental foreign-body infection.

We investigated the activity of linezolid, alone and in combination with rifampin (rifampicin), against a methicillin-resistant Staphylococcus aureus (MRSA) strain in vitro and in a guinea pig model of foreign-body infection. The MIC, minimal bactericidal concentration (MBC) in logarithmic phase, and MBC in stationary growth phase were 2.5, >20, and >20 microg/ml, respectively, for linezolid; 0.01, 0.08, and 2.5 microg/ml, respectively, for rifampin; and 0.16, 0.63, >20 microg/ml, respectively, for levofloxacin. In time-kill studies, bacterial regrowth and the development of rifampin resistance were observed after 24 h with rifampin alone at 1x or 4x the MIC and were prevented by the addition of linezolid. After the administration of single intraperitoneal doses of 25, 50, and 75 mg/kg of body weight, linezolid peak concentrations of 6.8, 12.7, and 18.1 microg/ml, respectively, were achieved in sterile cage fluid at approximately 3 h. The linezolid concentration remained above the MIC of the test organism for 12 h with all doses. Antimicrobial treatments of animals with cage implant infections were given twice daily for 4 days. Linezolid alone at 25, 50, and 75 mg/kg reduced the planktonic bacteria in cage fluid during treatment by 1.2 to 1.7 log(10) CFU/ml; only linezolid at 75 mg/kg prevented bacterial regrowth 5 days after the end of treatment. Linezolid used in combination with rifampin (12.5 mg/kg) was more effective than linezolid used as monotherapy, reducing the planktonic bacteria by >or=3 log(10) CFU (P < 0.05). Efficacy in the eradication of cage-associated infection was achieved only when linezolid was combined with rifampin, with cure rates being between 50% and 60%, whereas the levofloxacin-rifampin combination demonstrated the highest cure rate (91%) against the strain tested. The linezolid-rifampin combination is a treatment option for implant-associated infections caused by quinolone-resistant MRSA.

Cyclooxygenase-2 in human and experimental ischemic proliferative retinopathy.

BACKGROUND: Intravitreal neovascular diseases, as in ischemic retinopathies, are a major cause of blindness. Because inflammatory mechanisms influence vitreal neovascularization and cyclooxygenase (COX)-2 promotes tumor angiogenesis, we investigated the role of COX-2 in ischemic proliferative retinopathy. METHODS AND RESULTS: We describe here that COX-2 is induced in retinal astrocytes in human diabetic retinopathy, in the murine and rat model of ischemic proliferative retinopathy in vivo, and in hypoxic astrocytes in vitro. Specific COX-2 but not COX-1 inhibitors prevented intravitreal neovascularization, whereas prostaglandin E2, mainly via its prostaglandin E receptor 3 (EP3), exacerbated neovascularization. COX-2 inhibition induced an upregulation of thrombospondin-1 and its CD36 receptor, consistent with the observed antiangiogenic effects of COX-2 inhibition; EP3 stimulation reversed effects of COX-2 inhibitors on thrombospondin-1 and CD36. CONCLUSIONS: These findings point to an important role for COX-2 in ischemic proliferative retinopathy, as in diabetes.

The nuclear receptor DHR3 modulates dS6 kinase-dependent growth in Drosophila.

S6 kinases (S6Ks) act to integrate nutrient and insulin signaling pathways and, as such, function as positive effectors in cell growth and organismal development. However, they also have been shown to play a key role in limiting insulin signaling and in mediating the autophagic response. To identify novel regulators of S6K signaling, we have used a Drosophila-based, sensitized, gain-of-function genetic screen. Unexpectedly, one of the strongest enhancers to emerge from this screen was the nuclear receptor (NR), Drosophila hormone receptor 3 (DHR3), a critical constituent in the coordination of Drosophila metamorphosis. Here we demonstrate that DHR3, through dS6K, also acts to regulate cell-autonomous growth. Moreover, we show that the ligand-binding domain (LBD) of DHR3 is essential for mediating this response. Consistent with these findings, we have identified an endogenous DHR3 isoform that lacks the DBD. These results provide the first molecular link between the dS6K pathway, critical in controlling nutrient-dependent growth, and that of DHR3, a major mediator of ecdysone signaling, which, acting together, coordinate metamorphosis.

Selection on a eumelanic ornament is stronger in the tropics than in temperate zones in the worldwide-distributed barn owl.

Spatial variation in the pattern of natural selection can promote local adaptation and genetic differentiation between populations. Because heritable melanin-based ornaments can signal resistance to environmentally mediated elevation in glucocorticoids, to oxidative stress and parasites, populations may vary in the mean degree of melanic coloration if selection on these phenotypic aspects varies geographically. Within a population of Swiss barn owls (Tyto alba), the size of eumelanic spots is positively associated with survival, immunity and resistance to stress, but it is yet unknown whether Tyto species that face stressful environments evolved towards a darker eumelanic plumage. Because selection regimes vary along environmental gradients, we examined whether melanin-based traits vary clinally and are expressed to a larger extent in the tropics where parasites are more abundant than in temperate zones. To this end, we considered 39 barn owl species distributed worldwide. Barn owl species living in the tropics displayed larger eumelanic spots than those found in temperate zones. This was, however, verified in the northern hemisphere only. Parasites being particularly abundant in the tropics, they may promote the evolution of darker eumelanic ornaments.

In vivo quantification of neuro-glial metabolism and glial glutamate concentration using 1H-[13C] MRS at 14.1T.

Astrocytes have recently become a major center of interest in neurochemistry with the discoveries on their major role in brain energy metabolism. An interesting way to probe this glial contribution is given by in vivo (13) C NMR spectroscopy coupled with the infusion labeled glial-specific substrate, such as acetate. In this study, we infused alpha-chloralose anesthetized rats with [2-(13) C]acetate and followed the dynamics of the fractional enrichment (FE) in the positions C4 and C3 of glutamate and glutamine with high sensitivity, using (1) H-[(13) C] magnetic resonance spectroscopy (MRS) at 14.1T. Applying a two-compartment mathematical model to the measured time courses yielded a glial tricarboxylic acid (TCA) cycle rate (Vg ) of 0.27 ± 0.02 μmol/g/min and a glutamatergic neurotransmission rate (VNT ) of 0.15 ± 0.01 μmol/g/min. Glial oxidative ATP metabolism thus accounts for 38% of total oxidative metabolism measured by NMR. Pyruvate carboxylase (VPC ) was 0.09 ± 0.01 μmol/g/min, corresponding to 37% of the glial glutamine synthesis rate. The glial and neuronal transmitochondrial fluxes (Vx (g) and Vx (n) ) were of the same order of magnitude as the respective TCA cycle fluxes. In addition, we estimated a glial glutamate pool size of 0.6 ± 0.1 μmol/g. The effect of spectral data quality on the fluxes estimates was analyzed by Monte Carlo simulations. In this (13) C-acetate labeling study, we propose a refined two-compartment analysis of brain energy metabolism based on (13) C turnover curves of acetate, glutamate and glutamine measured with state of the art in vivo dynamic MRS at high magnetic field in rats, enabling a deeper understanding of the specific role of glial cells in brain oxidative metabolism. In addition, the robustness of the metabolic fluxes determination relative to MRS data quality was carefully studied.