Cytoskeletal regulation by the Nedd8 ubiquitin-like protein modification pathway

The Nedd8 ubiquitin-like protein modification pathway regulates cell-cycle progression. Our analysis of Nedd8 requirements during Caenorhabditis elegans embryogenesis indicates that the cytoskeleton is another target. Nedd8 conjugation negatively regulated contractility of the microfilament-rich cell cortex during pronuclear migration and again during cytokinesis. The Nedd8 pathway also was required after meiosis to negatively regulate katanin, a microtubule-severing complex, permitting the assembly of a large mitotic spindle. We propose that Nedd8-modified cullin, as part of an E3 ubiquitin ligase complex, targets katanin for degradation during the transition from meiosis to mitosis.     

Phase I and pharmacokinetic evaluation of the combination of orally administered docetaxel and cyclosporin A in tumor-bearing dogs

OBJECTIVE: To determine the maximum tolerated dose and characterize the pharmacokinetic disposition of an orally administered combination of docetaxel and cyclosporin A (CSA) in dogs with tumors. ANIMALS: 16 client-owned dogs with metastatic or advanced-stage refractory tumors. PROCEDURES: An open-label, dose-escalation, single-dose, phase I study of docetaxel administered in combination with a fixed dose of CSA was conducted. Docetaxel (at doses of 1.5, 1.625, or 1.75 mg/kg) and CSA (5 mg/kg) were administered concurrently via gavage twice during a 3-week period. Plasma docetaxel concentrations were quantified by use of high-performance liquid chromatography, and pharmacokinetic disposition was characterized by use of noncompartmental analysis. Dogs' clinical signs and results of hematologic and biochemical analyses were monitored for evidence of toxicosis. RESULTS: No acute hypersensitivity reactions were observed after oral administration of docetaxel. Disposition of docetaxel was dose independent over the range evaluated, and pharmacokinetic variables were similar to those reported in previous studies involving healthy dogs, with the exception that values for clearance were significantly higher in the dogs reported here. The maximum tolerated dose of docetaxel was 1.625 mg/kg. Gastrointestinal signs of toxicosis were dose limiting. CONCLUSIONS AND CLINICAL RELEVANCE: The absence of myelosuppression suggested that the docetaxel-CSA combination may be administered more frequently than the schedule used. Further studies are warranted to evaluate combination treatment administered on a biweekly schedule in dogs with epithelial tumors.     

Comparison of four methods of extracting DNA from D. gallinae (Acari: Dermanyssidae)

Dermanyssus gallinae is one of the most serious ectoparasites of poultry and it has been implicated as a vector of several major pathogenic diseases. Molecular detection of such pathogens in mites is crucial and therefore, an important step is the extraction of their DNA from mites. So, we compared four DNA extraction protocols from engorged and unfed individual mites: a conventional method using a Cethyl Trimethyl Ammonium Bromide buffer (CTAB), a Chelex resin, a Qiamp DNA extraction kit and a more recent one filter-based technology (FTA). The DNA samples have been tested for their ability to be amplified by an amplification of a D. gallinae 16S rRNA gene region. The best results were obtained using CTAB and Qiagen methods at the same time with unfed and engorged mites (96% and 100% of amplified samples). FTA produced similar results when using unfed mites but not when processing engorged ones (96% and 70%). Finally, the Chelex method was the least efficient in terms of DNA amplification, especially when applied on engorged individuals (50%). The possible inhibitor role of these Chelex extracted DNA was demonstrated by the means of a PCR control on PUC plasmid. No difference was observed with CTAB, Qiamp DNA extraction kit or FTA methods using DNA extracted one year before.     

Variability of retention process of isoxaflutole and its diketonitrile metabolite in soil under conventional and conservation tillage

BACKGROUND: Sorption largely controls pesticide fate in soils because it influences its availability for biodegradation or transport in the soil water. In this study, variability of sorption and desorption of isoxaflutole (IFT) and its active metabolite diketonitrile (DKN) was investigated under conventional and conservation tillage. RESULTS: According to soil samples, IFT K_D values ranged from 1.4 to 3.2 L kg^?1 and DKN K_D values ranged from 0.02 to 0.17 L kg^?1. Positive correlations were found between organic carbon content and IFT and DKN sorption. IFT and DKN sorption was higher under conservation than under conventional tillage owing to higher organic carbon content. Under conservation tillage, measurements on maize and oat residues collected from the soil surface showed a greater sorption of IFT on plant residues than on soil samples, with the highest sorbed quantities measured on maize residues (K_D ≈ 45 L kg^?1). Desorption of IFT was hysteretic, and, after five consecutive desorptions, between 72 and 89% of the sorbed IFT was desorbed from soil samples. For maize residues, desorption was weak (<50% of the sorbed IFT), but, after two complementary desorptions allowing for IFT hydrolysis, DKN was released from maize residues. CONCLUSION: Owing to an increase in organic carbon in topsoil layers, sorption of IFT and DKN was enhanced under conservation tillage. Greater sorption capacities under conservation tillage could help in decreasing DKN leaching to groundwater. Copyright © 2012 Society of Chemical Industry     

Goat uterine epithelial cells are susceptible to infection with Caprine Arthritis Encephalitis Virus (CAEV) in vivo

ABSTRACT: The aim of this study was to determine, using immunofluorescence and in situ hybridization, whether CAEV is capable of infecting goat uterine epithelial cells in vivo. Five CAEV seropositive goats confirmed as infected using double nested polymerase chain reaction (dnPCR) on leucocytes and on vaginal secretions were used as CAEV positive goats. Five CAEV-free goats were used as controls. Samples from the uterine horn were prepared for dnPCR, in situ hybridization, and immunofluorescence. The results from dnPCR confirmed the presence of CAEV proviral DNA in the uterine horn samples of infected goats whereas no CAEV proviral DNA was detected in samples taken from the uninfected control goats. The in situ hybridization probe was complementary to part of the CAEV gag gene and confirmed the presence of CAEV nucleic acids in uterine samples. The positively staining cells were seen concentrated in the mucosa of the lamina propria of uterine sections. Finally, laser confocal analysis of double p28/cytokeratin immunolabelled transverse sections of CAEV infected goat uterus, demonstrated that the virus was localized in glandular and epithelial cells. This study clearly demonstrates that goat uterine epithelial cells are susceptible to CAEV infection in vivo. This finding could help to further our understanding of the epidemiology of CAEV, and in particular the possibility of vertical transmission.