Activation of the Tumor Suppressor PP2A Emerges as a Potential Therapeutic Strategy for Treating Prostate Cancer

Protein phosphatase 2A (PP2A) is a tumor suppressor complex that has recently been reported as a novel and highly relevant molecular target in prostate cancer (PCa). However, its potential therapeutic value remains to be fully clarified. We treated PC-3 and LNCaP cell lines with the PP2A activators forskolin and FTY720 alone or combined with the PP2A inhibitor okadaic acid. We examined PP2A activity, cell growth, prostasphere formation, levels of PP2A phosphorylation, CIP2A and SET expression, and AKT and ERK activation. Interestingly, both forskolin and FTY720 dephosphorylated and activated PP2A, impairing proliferation and prostasphere formation and inducing changes in AKT and ERK phosphorylation. Moreover, FTY720 led to reduced CIP2A levels. Treatment with okadaic acid impaired PP2A activation thus demonstrating the antitumoral PP2A-dependent mechanism of action of both forskolin and FTY720. Levels of PP2A phosphorylation together with SET and CIP2A protein expression were studied in 24 PCa patients and both were associated with high Gleason scores and presence of metastatic disease. Altogether, our results suggest that PP2A inhibition could be involved in PCa progression, and the use of PP2A-activating drugs might represent a novel alternative therapeutic strategy for treating PCa patients.     

Non-lethal loop-mediated isothermal amplification assay as a point-of-care diagnostics tool for Neoparamoeba perurans,the causative agent of amoebic gill disease

Neoparamoeba perurans is the causative agent of amoebic gill disease (AGD). Two loop-mediated isothermal amplification (LAMP) assays targeting the parasite 18S rRNA and the Atlantic salmon EF1α, used as internal control, were designed. The N. perurans LAMP assay did not amplify close relatives N. pemaquidensis and N. branchiphila, or the host DNA. This assay detected 10⁶ copies of the parasite 18S rRNA gene under 13 min and 10³ copies under 35 min. Five “fast-and-dirty” DNA extraction methods were compared with a reference method and further validated by TaqMan™ qPCR. Of those, the QuickExtract buffer was selected for field tests. Seventy-one non-lethal gill swabs were analysed from AGD-clinically infected Atlantic salmon. The pathogen was detected under 23 min in fish of gill score >2 and under 39 min for lower gill scores. About 1.6% of the tests were invalid (no amplification of the internal control). 100% of positives were obtained from swabs taken from fish showing gill score ˃3, but only ~50% of positives for lower gill scores. The present LAMP assay could be implemented as a point-of-care test for the on-site identification of N. perurans; however, further work is required to improve its performance for lower scores.     

Pharmacokinetics of acetaminophen after intravenous and oral administration in fasted and fed Labrador Retriever dogs

Acetaminophen (paracetamol) is used in dogs to manage fever and mild pain. The aim of this study was to assess the pharmacokinetics of acetaminophen in both fed and fasted Labrador Retrievers after a single intravenous and oral administration (20 mg/kg). Six healthy dogs underwent three treatments in a randomized block study (a, n = 2; b, n = 2; c, n = 2). In phase one, group a received acetaminophen intravenously, group b and c orally after being fasted and fed, respectively. In phase two and three, groups were swapped, and the experiment was repeated. At the end of the trial, each dog received the same treatment. Acetaminophen plasma concentrations were detected using a validated HPLC‐UV method. The pharmacokinetic analysis was performed using a noncompartmental model. Clearance, volume at steady state and half‐life of acetaminophen in Labrador Retrievers were 0.42 L/kg hr, 0.87 L/kg and 1.35 hr, respectively. No significant statistical differences were found between fasted and fed dogs regarding maximum plasma concentration, time at maximum concentration and bioavailability as measured by the AUC. Feeding does not significantly affect the acetaminophen oral pharmacokinetics.     

A case study of biofilter activation and microbial nitrification in a marine recirculation aquaculture system for rearing Atlantic salmon (Salmo salar L.)

Marine recirculation aquaculture system (RAS) is a prominent technology within fish farming. However, the nitrifying bacteria in the biofilter have low growth rates, which can make the biofilter activation a long and delicate process with periods of low nitrification rates and variations in water quality. More knowledge on the microbial development in biofilters is therefore needed in order to understand the rearing conditions that favour optimal activation of the biofilters. In this case study, we investigated the activation of two biofilters in a marine RAS for Atlantic salmon post‐smolt associated with either high or low stocking densities of fish by monitoring the microbial communities and chemical composition. The results showed that the microbial communities in both biofilters were similar during the first rearing cycle, despite variations in the water quality. Nitrifying bacteria were established in both biofilters; however, the biofilter associated with low stocking density had the highest relative abundance of ammonia‐oxidizing Nitrosococcus (1.0%) and nitrite‐oxidizing Nitrospira (2.1%) at the end of the first rearing cycle, while the relative abundance of ammonia‐oxidizing Nitrosomonas (2.3%–2.9%) was similar in both biofilters. Our study showed that low fish stocking density during the first rearing cycle provided low and steady concentrations of ammonium, nitrite and organic load, which can stimulate rapid development of a nitrifying population in new marine RAS biofilters.     

High levels of spatial heterogeneity in the biodiversity of soil prokaryotes on Signy Island, Antarctica

In a previous study, soil bacterial diversity at environmentally distinct locations on Signy Island was examined using denaturing gradient gel electrophoresis (DGGE) profiling, and a range of chemical variables in soils was determined in order to describe variations between them. The dominant bacterial communities of all locations were found to be significantly different, although higher levels of similarity were observed between locations with similar physico-chemical characteristics, such as at penguin rookeries, seal wallows and vegetated soils. Extending this study, here soil prokaryote biodiversity was compared between 15 distinct locations in order to elucidate any interaction between four general habitat types on Signy Island (South Orkney Islands, maritime Antarctic) and any influence of previous human impacts at these sites. Specific sites were selected to represent the range of different soil environments present and to cover a range of environmental factors present in the maritime Antarctic which are known to influence bacterial community composition in soils elsewhere. A diverse prokaryote community is described, again with the majority of excised and sequenced bands belonging to the Bacteroidetes. Although DGGE profiling identified significant differences in prokaryotic biodiversity between all sampling sites, aggregations of banding patterns were also apparent across the different soil environments examined. Correlations between specific DGGE profiles and 10 selected soil parameters suggested that much of this variation could be explained by differences in the levels of environmental disturbance and soil pH. In particular, a greater proportion of variation in soil bacterial diversity was explained by differences in soil properties at human-disturbed locations than at undisturbed locations, with higher explanatory values by edaphic factors in the former and soil metal content in the later. In general, our data indicate that small-scale variation is an important factor in understanding patterns of prokaryotic distributions in soil habitats in the maritime Antarctic environment.     

Infection of cotton byfusarium oxysporum f.sp.vasinfectum as affected by water stress

Since virulence ofFusarium oxysporum f.sp.vasinfectum (FOV) on cotton (Gossypium hirsutum) is enhanced when the fungus is cultivated in a saline environment, excessively saline water must not be used for the irrigation of cotton. However, the limitations thus placed on the available water resources may lead to conditions of enforced water stress for the plant. The present study investigated whether water stress affects the susceptibility of cotton to FOV. Groups of 2-month-old cotton plants of theFusarium-susceptible Coker 304 and the moderately resistant GSC 20 varieties were maintained without watering for varying periods immediately before or after being inoculated with FOV (15 plants per group, two replications). Watering was suspended for 3, 6, 12 or 24 days before inoculation, and for 3, 6, 12 or 15 days after inoculation. After inoculation the plants were maintained in a controlled environment with a 15,000 lux, 12-h photoperiod, at 28°/24°C D/N, 20% r.h. Xylem water potential was determined in a pressure chamber. Percent infected leaf area and date of onset of wilt were the parameters used to define severity of FOV infection. There was a consistent relation between low water potential in the xylem (-7 and -20 MPa) and severity of infection, particularly when the dry period occurred after inoculation. After exposure to the lowest post-inoculation water potentials, even variety GSC 20, which is normally moderately resistant, exhibited a fairly high percent infected leaf area. This should be taken into account when the cotton grower is faced with water shortages, especially during the period from branching to flower bud break.     

Effects of racing on lymphocyte proliferation in horses

OBJECTIVE: To measure the lymphocyte proliferation response in horses 12 to 16 hours after completion of a race. ANIMALS: 8 Thoroughbreds that competed in 14 races and 3 control Thoroughbreds that did not race. PROCEDURE: Horses participated in races during the late afternoon or evening. Venous blood samples were collected on a morning before a race (1 or 2 days before the race or on the day of the race), on the afternoon of a race (40 to 60 minutes after the race), and on the morning of the day after a race (12 to 16 hours after the race). Lymphocyte proliferation responses and WBC count were measured in samples obtained in the mornings. Plasma cortisol was measured in all samples. RESULTS: Lymphocyte proliferation responses were significantly reduced and WBC counts significantly increased 12 to 16 hours after a race. Plasma cortisol concentrations were significantly increased 40 to 60 minutes after a race. In samples from the control horses, lymphocyte proliferation responses, WBC counts, or plasma cortisol concentrations did not differ significantly among time periods. CONCLUSIONS AND CLINICAL RELEVANCE: A decrease in proliferative responses of circulating lymphocytes can be found as late as 12 to 16 hours after a horse participates in a race. Although the clinical consequences of these exercise-related alterations of the immune response are not yet known, managers of horses should take into account that the immune system of a horse may be affected by racing.     

Variations in host defence mechanisms and blastogenesis in chickens and mice after rifamycin treatments

The immunobiological effect of rifamycins on different experimental models has been comparatively studied in chickens and mice using cyclophosphamide as a reference immunosuppressive agent. "In vivo" treatments with rifampicin and rifamycin SV diminished interferon levels in chickens and mice up to two hours after virus challenge. Mitogen induced blastogenesis was significantly depressed by either "in vivo" (5-50 mg/kg) or "in vitro" (5-20 micrograms/ml) drug treatments, although the activation of B lymphocytes was more easily inhibited than that of T lymphocytes. Primary anti-SRBC immune responses suffered a similar immune suppressive action by rifamycin treatments "in vivo". In every case, cyclophosphamide (10-20 micrograms/ml "in vitro" and 25 mg/kg "in vivo") proved to be the most active immunosuppressant. On the other hand, a remarkable parallelism in the effect of rifamycins in chickens and mice was observed in all the experimental systems used.     

Prevalence of tetracycline-resistant Campylobacter in organic broilers during a production cycle

Tetracycline (tet) resistance in Campylobacter isolated from organically raised broilers was investigated in this study. Two hundred forty-five samples from an organic broiler farm were collected weekly from the first week to the end of the production cycle, and they were cultured for thermophilic Campylobacter. Tetracycline resistance of these Campylobacter isolates was identified by the agar dilution method, whereas DNA fingerprinting profiles of tet-susceptible and tet-resistant strains were determined by pulsed-field gel electrophoresis (PFGE). None of the Campylobacter isolates from the third and the fourth week of the production period were resistant to tetracycline, whereas 66.7% of the isolates from the fifth week were resistant to this antibiotic. Although the prevalence of tetracycline resistance reached 100.0% during the sixth and seventh week, less than 34.0% of the isolates from the 10th week were resistant to this antimicrobial agent. In addition, only 13.8% of Campylobacter isolates from the intestinal tracts of these organically raised broilers were resistant to tetracycline. The presence of the tet(O) gene was detected in 98.9% of tet-resistant Campylobacter isolates, and tet-susceptible and tet-resistant Campylobacter strains showed distinct PFGE genotypes. The results suggest that the Campylobacter strains isolated from the early stage of the production were susceptible to tetracycline, but they were subsequently displaced by tet-resistant Campylobacter.     

In vitro studies of Lactobacillus delbrueckii subsp. lactis in Atlantic salmon (Salmo salar L.) foregut: tissue responses and evidence of protection against Aeromonas salmonicida subsp. salmonicida epithelial damage

Probiotic bacteria increase the host health status and protect mucosal tissue against pathogen-caused damage in mammalian models. Using an in vitro (intestinal sac) method this study aimed to address (a) the in vitro ability of Lactobacillus delbrueckii subsp. lactis to remain in the gastrointestinal tract of Atlantic salmon (Salmo salar L.) and (b) its ability to prevent cellular damage caused by successive incubation with Aeromonas salmonicida subsp. salmonicida the causative agent of furunculosis. Short in vitro incubation of salmon foregut with (TRITC)-labelled L. delbrueckii subsp. lactis showed that the probiont was able to colonize the enterocyte surface as studied by confocal microscopy. Furthermore, foregut incubated with the probiotic bacteria only, resulted in a healthy intestinal barrier whereas exposure to A. salmonicida disrupted its integrity. However, pre-treatment of salmon intestine with L. delbrueckii subsp. lactis prevented Aeromonas damaging effects. These results are promising in the context of the use of non-autochthonous probiotic bacteria as prophylactic agents against fish bacterial infections in the gastrointestinal tract.