Long-Term Safety of Repeated Blood-Brain Barrier Opening via Focused Ultrasound with Microbubbles in Non-Human Primates Performing a Cognitive Task

Focused Ultrasound (FUS) coupled with intravenous administration of microbubbles (MB) is a non-invasive technique that has been shown to reliably open (increase the permeability of) the blood-brain barrier (BBB) in multiple in vivo models including non-human primates (NHP). This procedure has shown promise for clinical and basic science applications, yet the safety and potential neurological effects of long term application in NHP requires further investigation under parameters shown to be efficacious in that species (500kHz, 200–400 kPa, 4–5μm MB, 2 minute sonication). In this study, we repeatedly opened the BBB in the caudate and putamen regions of the basal ganglia of 4 NHP using FUS with systemically-administered MB over 4–20 months. We assessed the safety of the FUS with MB procedure using MRI to detect edema or hemorrhaging in the brain. Contrast enhanced T1-weighted MRI sequences showed a 98% success rate for openings in the targeted regions. T2-weighted and SWI sequences indicated a lack edema in the majority of the cases. We investigated potential neurological effects of the FUS with MB procedure through quantitative cognitive testing of’ visual, cognitive, motivational, and motor function using a random dot motion task with reward magnitude bias presented on a touchpanel display. Reaction times during the task significantly increased on the day of the FUS with MB procedure. This increase returned to baseline within 4–5 days after the procedure. Visual motion discrimination thresholds were unaffected. Our results indicate FUS with MB can be a safe method for repeated opening of the BBB at the basal ganglia in NHP for up to 20 months without any long-term negative physiological or neurological effects with the parameters used.     

Caracterización morfológica y molecular del antagonismo entre el endofito Diaporthe sp. aislado de frailejón (Espeletia sp.) y el fitopatógeno Phytophthora infestans

Endophytic fungi produce a great variety of secondary metabolites both in vivo and in vitro. In this study, we characterized the ability of a sterile-mycelium endophytic fungus isolated from Espeletia sp. to control the growth of Phytophthora infestans in Petri dishes. Sequence from the ITS regions (internal transcribed spacer) of the endophyte showed 94% similarity to Diaporthe phaseolorum's. The antagonistic interaction between Diaporthe sp. and P. infestans was evaluated in three different culture media. Diaporthe sp. showed an antagonistic effect towards P. infestans, with some variation depending on which medium was used. In an attempt to identify possible genes involved in this antagonism, we detected a gene from the endophyte encoding an amylase, which was differentially expressed during this biotic interaction.     

Serum T-kininogen levels increase two to four months before death.

We have reported an accumulation of T-kininogen mRNA in the liver of aging Sprague-Dawley rats. T-kininogen is a cysteine proteinase inhibitor. Since a disruption of the intracellular protein degradation machinery is known to occur during senescence, we wished to further define the role of this protein in the aging process. As a first step, we have measured T-kininogen levels both in serum and within the liver. We have found that serum protein levels are indeed augmented during senescence, although not as dramatically as the mRNA (2.5-fold versus 8.3-fold). Immunocytochemistry, as well as Western blot analysis suggests that this is due to the presence of T-kininogen within hepatic cells in aged rats. Life-long dietary restriction, a known age-prolonging treatment, decreases the overexpression of the protein in 24-month-old rats. Later, diet-restricted animals still show an increased expression from the gene, the effect being delayed but not abolished by dietary manipulation. Interestingly, a longitudinal study indicated the existence of a positive correlation between the time of increase of serum T-kininogen and the time of death of the animal. Serum T-kininogen was found to increase 2.5-4 months before death.     

Validated predictive modelling of the environmental resistome

Multi-drug-resistant bacteria pose a significant threat to public health. The role of the environment in the overall rise in antibiotic-resistant infections and risk to humans is largely unknown. This study aimed to evaluate drivers of antibiotic-resistance levels across the River Thames catchment, model key biotic, spatial and chemical variables and produce predictive models for future risk assessment. Sediment samples from 13 sites across the River Thames basin were taken at four time points across 2011 and 2012. Samples were analysed for class 1 integron prevalence and enumeration of third-generation cephalosporin-resistant bacteria. Class 1 integron prevalence was validated as a molecular marker of antibiotic resistance; levels of resistance showed significant geospatial and temporal variation. The main explanatory variables of resistance levels at each sample site were the number, proximity, size and type of surrounding wastewater-treatment plants. Model 1 revealed treatment plants accounted for 49.5% of the variance in resistance levels. Other contributing factors were extent of different surrounding land cover types (for example, Neutral Grassland), temporal patterns and prior rainfall; when modelling all variables the resulting model (Model 2) could explain 82.9% of variations in resistance levels in the whole catchment. Chemical analyses correlated with key indicators of treatment plant effluent and a model (Model 3) was generated based on water quality parameters (contaminant and macro- and micro-nutrient levels). Model 2 was beta tested on independent sites and explained over 78% of the variation in integron prevalence showing a significant predictive ability. We believe all models in this study are highly useful tools for informing and prioritising mitigation strategies to reduce the environmental resistome.     

Biochemical,biophysical, and genetic changes of porcine trophoblast‐derived stem‐like cells during differentiation as evaluated using Raman microspectroscopy,Atomic force microscopy,and quantitative polymerase chain reaction

Porcine trophoblast‐derived stem‐like cells grown into serum medium start to differentiate and become senescent within 30 days. However, trophoblast‐derived cells, cultured in vitro in a defined and non‐serum medium, have the regenerative properties, such as indefinite passage and foreign DNA receptivity, similar to stem cells. To evaluate the biochemical, biophysical, and genetic changes of the terminal differentiation of trophoblast derived cells, Raman microspectroscopy, atomic force microscopy, and qPCR were applied. It was found that Raman spectral intensities of characteristic peaks, cell morphology, and Young's modulus can be used to distinguish differentiated and undifferentiated trophoblast cells. In addition, 17 cytoskeleton and extracellular matrix‐related genes were significantly impacted by medium type (non‐serum versus serum). Our findings suggest that Raman microspectroscopy and atomic force microscopy—both considered as label‐free, non‐invasive techniques—can be applied to distinguish differentiated trophoblast cells, and cellular biochemical information and biophysical properties can be indicative of cellular differences during cell differentiation. In addition, most of cytoskeleton‐related genes exhibit similar pattern to that of Young's modulus during trophoblast cell differentiation, indicating the potential connection between cytoskeleton‐related genes and cellular stiffness. genesis 53:749–761, 2015. © 2015 Wiley Periodicals, Inc.     

Parameters Influencing Baseline HIV-1 Genotypic Tropism Testing Related to Clinical Outcome in Patients on Maraviroc

ObjectivesWe analysed the impact of different parameters on genotypic tropism testing related to clinical outcome prediction in 108 patients on maraviroc (MVC) treatment.Methods87 RNA and 60 DNA samples were used. The viral tropism was predicted using the geno2pheno_[coreceptor] and T-CUP tools with FPR cut-offs ranging from 1%-20%. Additionally, 27 RNA and 28 DNA samples were analysed in triplicate, 43 samples with the ESTA assay and 45 with next-generation sequencing. The influence of the genotypic susceptibility score (GSS) and 16 MVC-resistance mutations on clinical outcome was also studied.ResultsConcordance between single-amplification testing compared to ESTA and to NGS was in the order of 80%. Concordance with NGS was higher at lower FPR cut-offs. Detection of baseline R5 viruses in RNA and DNA samples by all methods significantly correlated with treatment success, even with FPR cut-offs of 3.75%-7.5%. Triple amplification did not improve the prediction value but reduced the number of patients eligible for MVC. No influence of the GSS or MVC-resistance mutations but adherence to treatment, on the clinical outcome was detected.ConclusionsProviral DNA is valid to select candidates for MVC treatment. FPR cut-offs of 5%-7.5% and single amplification from RNA or DNA would assure a safe administration of MVC without excluding many patients who could benefit from this drug. In addition, the new prediction system T-CUP produced reliable results.     

Relationship of aerobic and anaerobic parameters with 400 m front crawl swimming performance

The aims of the present study were to investigate the relationship of aerobic and anaerobic parameters with 400 m performance, and establish which variable better explains long distance performance in swimming. Twenty-two swimmers (19.1±1.5 years, height 173.9±10.0 cm, body mass 71.2±10.2 kg; 76.6±5.3% of 400 m world record) underwent a lactate minimum test to determine lactate minimum speed (LMS) (i.e., aerobic capacity index). Moreover, the swimmers performed a 400 m maximal effort to determine mean speed (S400m), peak oxygen uptake (V.O2PEAK) and total anaerobic contribution (C_ANA). The C_ANA was assumed as the sum of alactic and lactic contributions. Physiological parameters of 400 m were determined using the backward extrapolation technique (V.O2PEAK and alactic contributions of C_ANA) and blood lactate concentration analysis (lactic anaerobic contributions of C_ANA). The Pearson correlation test and backward multiple regression analysis were used to verify the possible correlations between the physiological indices (predictor factors) and S400m (independent variable) (p < 0.05). Values are presented as mean ± standard deviation. Significant correlations were observed between S400m (1.4±0.1 m·s^-1) and LMS (1.3±0.1 m·s^-1; r = 0.80), V.O2PEAK (4.5±3.9 L·min^-1; r = 0.72) and C_ANA (4.7±1.5 L·O₂; r= 0.44). The best model constructed using multiple regression analysis demonstrated that LMS and V.O2PEAK explained 85% of the 400 m performance variance. When backward multiple regression analysis was performed, C_ANA lost significance. Thus, the results demonstrated that both aerobic parameters (capacity and power) can be used to predict 400 m swimming performance.