Cloning and functional analysis of the rhesus macaque ABCG2 gene. Forced expression confers an SP phenotype among hematopoietic stem cell progeny in vivo

Hematopoietic cells can be highly enriched for repopulating ability based upon the efflux of the fluorescent Hoechst 33342 dye by sorting for SP (side population) cells, a phenotype attributed to expression of ABCG2, a member of the ABC transporter superfamily. Intriguingly, murine studies suggest that forced ABCG2 expression prevents hematopoietic differentiation. We cloned the full-length rhesus ABCG2 and introduced it into a retroviral vector. ABCG2-transduced human peripheral blood progenitor cells (PBPCs) acquired the SP phenotype but showed significantly reduced growth compared with control. Two rhesus macaques received autologous PBPCs split for transduction with the ABCG2 or control vectors. Marking levels were similar between fractions with no discrepancy between bone marrow and peripheral blood marking. Analysis for the SP phenotype among bone marrow and mature blood populations confirmed ABCG2 expression at levels predicted by vector copy number long term, demonstrating no block to differentiation in the large animal. In vitro studies showed selective protection against mitoxantrone among ABCG2-transduced rhesus PBPCs. Our results confirm the existence of rhesus ABCG2, establish its importance in conferring the SP phenotype, suggest no detrimental effect of its overexpression upon differentiation in vivo, and imply a potential role for its overexpression as an in vivo selection strategy for gene therapy applications.     

Immunocompetence increases with larval body size in a phytophagous moth

Despite the obvious benefit of an immune system, its efficacy against pathogens and parasites may show great variation among individuals, populations and species. Understanding the causes of this variation is becoming a central theme in ecology. Many biotic and abiotic factors are known to influence immunocompetence (temperature, age, etc.). However, for a given age, size among individuals varies, probably as a result of accumulated resources. Thus, these variable resources could be allocated to immune defence and, consequently, body size may explain part of the variation in immune responsiveness. However, the influence of body size on immune defence is often overlooked. The present study investigates variations in haemocyte count and phenoloxidase activity in larvae of the phytophagous vine moth Eupoecilia ambiguella Hübner of the same age, although differing in body size. The measurements of immune function are made both when the insects are immunologically naïve and 24 h after a bacterial immune challenge. The base levels of these immune parameters do not covary with body size in naïve larvae. After the bacterial immune challenge, more haemocytes and phenoloxidase enzyme are mobilized, and the mobilization of these immune effectors is correlated positively with individual body size. Thus, larger larvae exhibit higher immunocompetence than smaller ones, suggesting that smaller larvae might be more vulnerable to infection. These results suggest that body size is probably an underestimated variable, which nevertheless modulates the insect immune system and should thus be considered as a covariate in insect immune system measurement. It is recommended therefore, that body size should be taken into account in ecological immunity studies with insects. © 2013 The Royal Entomological Society     

LabKey Server: An open source platform for scientific data integration, analysis and collaboration

Background

This paper is devoted to distance measures for leaf-labelled trees on free leafset. A leaf-labelled tree is a data structure which is a special type of a tree where only leaves (terminal) nodes are labelled. This data structure is used in bioinformatics for modelling of evolution history of genes and species and also in linguistics for modelling of languages evolution history. Many domain specific problems occur and need to be solved with help of tree postprocessing techniques such as distance measures.

Results

Here we introduce the tree edit distance designed for leaf labelled trees on free leafset, which occurs to be a metric. It is presented together with tree edit consensus tree notion. We provide statistical evaluation of provided measure with respect to R-F, MAST and frequent subsplit based dissimilarity measures as the reference measures.

Conclusions

The tree edit distance was proven to be a metric and has the advantage of using different costs for contraction and pruning, therefore their properties can be tuned depending on the needs of the user. Two of the presented methods carry the most interesting properties. E(3,1) is very discriminative (having a wide range of values) and has a very regular distance distribution which is similar to a normal distribution in its shape and is good both for similar and non-similar trees. NFC(2,1) on the other hand is proportional or nearly proportional to the number of mutation operations used, irrespective of their type.     

Changes in Sensitivity of Reward and Motor Behavior to Dopaminergic,Glutamatergic, and Cholinergic Drugs in a Mouse Model of Fragile X Syndrome

Fragile X syndrome (FXS) is a leading cause of intellectual disability. FXS is caused by loss of function of the FMR1 gene, and mice in which Fmr1 has been inactivated have been used extensively as a preclinical model for FXS. We investigated the behavioral pharmacology of drugs acting through dopaminergic, glutamatergic, and cholinergic systems in fragile X (Fmr1 ^-/Y) mice with intracranial self-stimulation (ICSS) and locomotor activity measurements. We also measured brain expression of tyrosine hydroxylase (TH), the rate-limiting enzyme in dopamine biosynthesis. Fmr1 ^-/Y mice were more sensitive than wild type mice to the rewarding effects of cocaine, but less sensitive to its locomotor stimulating effects. Anhedonic but not motor depressant effects of the atypical neuroleptic, aripiprazole, were reduced in Fmr1 ^-/Y mice. The mGluR5-selective antagonist, 6-methyl-2-(phenylethynyl)pyridine (MPEP), was more rewarding and the preferential M1 antagonist, trihexyphenidyl, was less rewarding in Fmr1 ^-/Y than wild type mice. Motor stimulation by MPEP was unchanged, but stimulation by trihexyphenidyl was markedly increased, in Fmr1 ^-/Y mice. Numbers of midbrain TH+ neurons in the ventral tegmental area were unchanged, but were lower in the substantia nigra of Fmr1 ^-/Y mice, although no changes in TH levels were found in their forebrain targets. The data are discussed in the context of known changes in the synaptic physiology and pharmacology of limbic motor systems in the Fmr1 ^-/Y mouse model. Preclinical findings suggest that drugs acting through multiple neurotransmitter systems may be necessary to fully address abnormal behaviors in individuals with FXS.     

Comparison of Worm Control Strategies in Grazing Sheep in Denmark

Control of nematode parasites with reduced reliance on the use of anthelmintics was studied in 16 ewes with suckling twin lambs on contaminated pasture in Denmark. Ewes and lambs were treated with albendazole at turn-out 3 May. Ewes were removed from the groups on 26 July, and lambs were slaughtered on 11 October. The animals were allocated to 4 groups of 8 lambs and their 4 ewes. Group TS was treated with albendazole at weeks 3, 6 and 8 after turnout and set-stocked; group TM was similarly treated but moved to clean pasture in conjunction with the last drenching; group US was untreated and set-stocked, and group UM was left untreated but moved to clean pasture week 8 after turn-out. Supplementary feed was offered in June and August due to scarcity of pasture. Strategic treatments of ewes and lambs weeks 3, 6 and 8 after turn-out, with or without a move to clean pasture, were highly effective in controlling nematode infections for most of the season. This was reflected in better weight gains and carcass characteristics in the treated compared to untreated lambs, resulting in an average increase in the value of the product by 36%. The effect of moving without treatment (UM) on faecal egg counts was limited but peak pasture infectivity was reduced to less than 10% compared to the set-stocked group and weight gains of lambs were significantly better despite poor feed availability in late season. The study showed that under set-stocked conditions repeated anthelmintic treatments of both ewes and lambs in early season may ensure sufficient nematode control whereas moving animals to clean pasture without dosing was less efficient. The latter may, however, still be a viable option in organic and other production systems where routine use of anthelmintics is banned, particularly if weaning and moving are combined or a second move is performed.     

Investigation of apoptosis in cultured cells infected with equine herpesvirus 1

Many viruses alter different stages of apoptosis of infected cells as a strategy for successful infection. Few studies have addressed mechanisms of equine herpesvirus 1 (EHV-1) strain-induced cell death. We investigated the effect of an abortigenic strain (AR8 strain) on heterologous Madin–Darby bovine kidney cells and homologous equine dermis (ED) cells cell lines. We compared morphologic and biochemical features of early and late apoptosis at different postinfection times. We investigated translocation of phosphatidylserine to the cell surface, nuclear fragmentation and changes in the cytoskeleton using flow cytometry and annexin V/propidium iodide staining, DNA laddering, terminal deoxynucleotidyl transferase UTP nick-end labeling assay and immunofluorescence staining of cytokeratin 18 cleavage. AR8 EVH-1 strain interfered with apoptosis in both cell lines, particularly during the middle stage of the replication cycle; this was more evident in ED cells. Although this antiapoptotic effect has been reported for other alpha herpesviruses, our findings may help elucidate how EHV-1 improves its infectivity during its cycle.