Prolonged expression of CRISPR/Cas9 raises concerns about off-target cleavage, cytotoxicity, and immune responses. To address these issues, we have developed a system to produce helper-dependent ...adenoviruses that express CRISPR/Cas9 to direct cleavage of the vectors’ own genome after transduction of target cells. To prevent self-cleavage during vector production, it was necessary to downregulate Cas9 mRNA as well as inhibit Cas9 protein activity. Cas9 mRNA downregulation was achieved by inserting the target sequences for the helper-virus-encoded miRNA, mivaRNAI, and producer-cell-encoded miRNAs, hsa-miR183-5p, and hsa-miR218-5p, into the 3′ UTR of the HDAd-encoded Cas9 expression cassette. Cas9 protein activity was inhibited by expressing anti-CRISPR proteins AcrIIA2 and AcrAII4 from both the producer cells and the helper virus. After purification, these helper-dependent adenoviruses will perform CRISPR/Cas9-mediated self-cleavage in the transduced target cells, thereby limiting the duration of Cas9 expression and thus represent an important platform for improving the safety of gene editing by CRISPR/Cas9.
Previously unstudied dibenzoindolo3,2-bcarbazoles have been prepared by two-directional, phase tag-assisted synthesis utilizing a connective-Pummerer cyclization and a SmI2-mediated tag ...cleavage–cyclization cascade. The use of a phase tag allows us to exploit unstable intermediates that would otherwise need to be avoided. The novel materials were characterized by X-ray, cyclic voltammetry, UV–vis spectroscopy, TGA, and DSC. Preliminary studies on the performance of OFET devices are also described.
Depsipeptide, FR901228, has demonstrated potent in vitro and in vivo cytotoxic activity against murine and human tumor cell lines. In the laboratory, it has been shown to be a histone deacetylase ...(HDAC) inhibitor. In a phase I trial of depsipeptide conducted at the National Cancer Institute, 3 patients with cutaneous T-cell lymphoma had a partial response, and 1 patient with peripheral T-cell lymphoma, unspecified, had a complete response. Sézary cells isolated from patients after treatment had increased histone acetylation. These results suggest that inhibition of HDAC is a novel and potentially effective therapy for patients with T-cell lymphoma.
Despite the high number of children living with neurodevelopmental disabilities in sub-Saharan Africa, access to early intervention is almost non-existent. It is therefore important to develop ...feasible, scalable early autism intervention that can be integrated into systems of care. While Naturalistic Developmental Behavioural Intervention (NDBI) has emerged as an evidence-based intervention approach, implementation gaps exist globally, and task-sharing approaches may address access gaps. In this South African proof-of-principle pilot study, we set out to answer two questions about a 12-session cascaded task-sharing NDBI-whether the approach could be delivered with fidelity, and whether we could identify signals of change in child and caregiver outcomes.
We utilized a single-arm pre-post design. Fidelity (non-specialists, caregivers), caregiver outcomes (stress, sense of competence), and child outcomes (developmental, adaptive) were measured at baseline (T1) and follow-up (T2). Ten caregiver-child dyads and four non-specialists participated. Pre-to-post summary statistics were presented alongside individual trajectories. Non-parametric Wilcoxon signed rank test for paired samples was used to compare group medians between T1 and T2.
Caregiver implementation fidelity increased in 10/10 participants. Non-specialists demonstrated a significant increase in coaching fidelity (increases in 7/10 dyads). Significant gains were seen on two Griffiths-III subscales (Language/Communication-9/10 improved, Foundations of Learning-10/10 improved) and on the General Developmental Quotient (9/10 improved). Significant gains were also seen on two Vineland Adaptive Behaviour Scales (Third Edition) subscales (Communication-9/10 improved, Socialization-6/10 improved) and in the Adaptive Behaviour Standard Score (9/10 improved). Caregiver sense of competence improved in 7/10 caregivers and caregiver stress in 6/10 caregivers.
This proof-of-principle pilot study of the first cascaded task-sharing NDBI in Sub-Saharan Africa provided fidelity and intervention outcome data which supported the potential of such approaches in low-resource contexts. Larger studies are needed to expand on the evidence-base and answer questions on intervention effectiveness and implementation outcomes.
The effect of pH on the glucose metabolism of non-growing cells of L. lactis MG1363 was studied by in vivo NMR in the range 4.8 to 6.5. Immediate pH effects on glucose transporters and/or enzyme ...activities were distinguished from transcriptional/translational effects by using cells grown at the optimal pH of 6.5 or pre-adjusted to low pH by growth at 5.1. In cells grown at pH 5.1, glucose metabolism proceeds at a rate 35% higher than in non-adjusted cells at the same pH. Besides the upregulation of stress-related genes (such as dnaK and groEL), cells adjusted to low pH overexpressed H(+)-ATPase subunits as well as glycolytic genes. At sub-optimal pHs, the total intracellular pool of lactic acid reached approximately 500 mM in cells grown at optimal pH and about 700 mM in cells grown at pH 5.1. These high levels, together with good pH homeostasis (internal pH always above 6), imply intracellular accumulation of the ionized form of lactic acid (lactate anion), and the concomitant export of the equivalent protons. The average number, n, of protons exported with each lactate anion was determined directly from the kinetics of accumulation of intra- and extracellular lactic acid as monitored online by (13)C-NMR. In cells non-adjusted to low pH, n varies between 2 and 1 during glucose consumption, suggesting an inhibitory effect of intracellular lactate on proton export. We confirmed that extracellular lactate did not affect the lactate: proton stoichiometry. In adjusted cells, n was lower and varied less, indicating a different mix of lactic acid exporters less affected by the high level of intracellular lactate. A qualitative model for pH effects and acid stress adaptation is proposed on the basis of these results.
During the first months of observations, the Magnetospheric Multiscale Fly's Eye Energetic Particle Spectrometer instrument has observed several instances of electron acceleration up to greater than ...100 keV while in the vicinity of the dayside reconnection region. While particle acceleration associated with magnetic reconnection has been seen to occur up to these energies in the tail region, it had not yet been reported at the magnetopause. This study reports on observations of electron acceleration up to hundreds of keV that were recorded on 19 September 2015 around 1000 UT, in the midst of an X-line crossing. In the region surrounding the X-line, whistler-mode and broadband electrostatic waves were observed simultaneously with the appearance of highly energetic electrons which exhibited significant energization in the perpendicular direction. The mechanisms by which particles may be accelerated via reconnection-related processes are intrinsic to understanding particle dynamics among a wide range of spatial scales and plasma environments.
The Haast chronosequence on the west coast of the South Island of New Zealand consists of a series of coastal dune ridges formed by periodic earthquake disturbance over the last 6500years. It ...approximates an ideal chronosequence, because soils are under the same climate and vegetation, have similar topography, and are developed in the same parent material. We assessed soil development and the changes in nutrients at eleven sites along the sequence to determine rates of nutrient transformations and the potential for limitation of biological activity by nitrogen and phosphorus availability. Soils develop rapidly to podzols (Spodosols) under the super-humid climate, involving acidification and depletion of base cations in the first few hundred years, development of a bleached eluvial horizon under a thick organic horizon within 2000years, and formation of a continuous cemented iron pan within 4000years. Soil phosphorus concentrations decline markedly in the first few hundred years in both the organic and upper mineral soil horizons, with rapid depletion of primary mineral phosphate and the accumulation of organic phosphorus to ~80% of the total phosphorus in the upper mineral soil. These changes lead to an increasing degree of phosphorus limitation along the sequence, as indicated by an increase in C:P and N:P ratios and a decline in available phosphate. The rates of soil development and phosphorus depletion are more rapid than at the nearby Franz Josef post-glacial chronosequence, where glacial moraine derived from graywacke contains a relatively high phosphorus concentration and weathers into fine-textured soils. The Haast chronosequence therefore provides an important additional example of soil development linked to long-term depletion of soil phosphorus under a perudic moisture regime.
► Rapid podzol formation in coastal dunes under the super-humid climate of Westland, New Zealand. ► Phosphorus in surface soil present predominantly in organic forms for the majority of the sequence.. ► Increasing phosphorus limitation indicated by changes in N:P ratio and available phosphate. ► Decline in total phosphorus consistent with that at the nearby Franz Josef post-glacial sequence. ► Possibility of reconciling phosphorus depletion rates among chronosequences worldwide.
Does bioethics exist? Turner, L
Journal of medical ethics,
12/2009, Letnik:
35, Številka:
12
Journal Article
Recenzirano
Bioethicists disagree over methods, theories, decision-making guides, case analyses and public policies. Thirty years ago, the thinking of many scholars coalesced around a principlist approach to ...bioethics. That mid-level mode of moral reasoning is now one of many approaches to moral deliberation. Significant variation in contemporary approaches to the study of ethical issues related to medicine, biotechnology and health care raises the question of whether bioethics exists as widely shared method, theory, normative framework or mode of moral reasoning.
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