Novel Protein Kinase Signaling Systems Regulating Lifespan Identified by Small Molecule Library Screening Using Drosophila

Protein kinase signaling cascades control most aspects of cellular function. The ATP binding domains of signaling protein kinases are the targets of most available inhibitors. These domains are highly conserved from mammals to flies. Herein we describe screening of a library of small molecule inhibitors of protein kinases for their ability to increase Drosophila lifespan. We developed an assay system which allowed screening using the small amounts of materials normally present in commercial chemical libraries. The studies identified 17 inhibitors, the majority of which targeted tyrosine kinases associated with the epidermal growth factor receptor (EGFR), platelet-derived growth factor (PDGF)/vascular endothelial growth factor (VEGF) receptors, G-protein coupled receptor (GPCR), Janus kinase (JAK)/signal transducer and activator of transcription (STAT), the insulin and insulin-like growth factor (IGFI) receptors. Comparison of the protein kinase signaling effects of the inhibitors in vitro defined a consensus intracellular signaling profile which included decreased signaling by p38MAPK (p38), c-Jun N-terminal kinase (JNK) and protein kinase C (PKC). If confirmed, many of these kinases will be novel additions to the signaling cascades known to regulate metazoan longevity

Ab-Externo AAV-Mediated Gene Delivery to the Suprachoroidal Space Using a 250 Micron Flexible Microcatheter

The current method of delivering gene replacement to the posterior segment of the eye involves a three-port pars plana vitrectomy followed by injection of the agent through a 37-gauge cannula, which is potentially wrought with retinal complications. In this paper we investigate the safety and efficacy of delivering adeno-associated viral (AAV) vector to the suprachoroidal space using an ab externo approach that utilizes an illuminated microcatheter.6 New Zealand White rabbits and 2 Dutch Belted rabbits were used to evaluate the ab externo delivery method. sc-AAV5-smCBA-hGFP vector was delivered into the suprachoroidal space using an illuminated iTrackTM 250A microcatheter. Six weeks after surgery, the rabbits were sacrificed and their eyes evaluated for AAV transfection using immunofluorescent antibody staining of GFP.Immunostaining of sectioned and whole-mounted eyes demonstrated robust transfection in all treated eyes, with no fluorescence in untreated control eyes. Transfection occurred diffusely and involved both the choroid and the retina. No apparent adverse effects caused by either the viral vector or the procedure itself could be seen either clinically or histologically.The ab externo method of delivery using a microcatheter was successful in safely and effectively delivering a gene therapy agent to the suprachoroidal space. This method presents a less invasive alternative to the current method of virally vectored gene delivery

Tailoring communications to the evolving needs of patients throughout the cancer care trajectory: a qualitative exploration with breast cancer patients

Background: Doctor-patient communication is a crucial aspect of patient care. This study explored the communication experience of patients in a cancer consultation over the course of the cancer continuum. Methods: In-depth interviews with seven breast cancer patients were carried out. Results: Themes related to communication experiences across the five phases of cancer consultation, from diagnosis to recurrence, were identified. The most salient issue is that patients also perceived cancer as 'a disease of the mind', which is not adequately cared for in consultation. This highlights the notion that cancer care providers should provide appropriate care for the psychological dimensions of the cancer experience with an empathic and sincere attitude during consultations. To this end, non-verbal aspects of communication that convey caring, support, and respect seem important. Furthermore, patients perceived that the consultation time was far shorter then they needed and reported that they felt pressured for time. Moreover, the stance taken by patients and the needs and preferences of patients varied across the phases of the cancer trajectory. As patients progressed through the phases of their treatment, they assumed more active roles in the course of their care and the need for more detailed information and questioning increased. Thus, ensuring that patients have opportunities to ask questions in the consultation is important. Conclusion: Current findings suggest that the efficacy of communication varies depending on which phase patients are in and that effective communication should be tailored to these evolving needs and preferences of breast cancer patients. Also, patients perceived that the consultation did not adequately address their need for information related to their care or their emotional issues associated with the cancer experience. It is therefore important to address their needs by paying particular attention to non-verbal aspects of communication that convey empathy and respect toward patients, as well as allowing patients to ask questionsope

A Simple and Effective Method for Construction of Escherichia coli Strains Proficient for Genome Engineering

Multiplex genome engineering is a standalone recombineering tool for large-scale programming and accelerated evolution of cells. However, this advanced genome engineering technique has been limited to use in selected bacterial strains. We developed a simple and effective strain-independent method for effective genome engineering in Escherichia coli. The method involves introducing a suicide plasmid carrying the l Red recombination system into the mutS gene. The suicide plasmid can be excised from the chromosome via selection in the absence of antibiotics, thus allowing transient inactivation of the mismatch repair system during genome engineering. In addition, we developed another suicide plasmid that enables integration of large DNA fragments into the lacZ genomic locus. These features enable this system to be applied in the exploitation of the benefits of genome engineering in synthetic biology, as well as the metabolic engineering of different strains of E. coli.open7

Combining active learning and semi-supervised learning techniques to extract protein interaction sentences

Background: Protein-protein interaction (PPI) extraction has been a focal point of many biomedical research and database curation tools. Both Active Learning and Semi-supervised SVMs have recently been applied to extract PPI automatically. In this paper, we explore combining the AL with the SSL to improve the performance of the PPI task. Methods: We propose a novel PPI extraction technique called PPISpotter by combining Deterministic Annealing-based SSL and an AL technique to extract protein-protein interaction. In addition, we extract a comprehensive set of features from MEDLINE records by Natural Language Processing (NLP) techniques, which further improve the SVM classifiers. In our feature selection technique, syntactic, semantic, and lexical properties of text are incorporated into feature selection that boosts the system performance significantly. Results: By conducting experiments with three different PPI corpuses, we show that PPISpotter is superior to the other techniques incorporated into semi-supervised SVMs such as Random Sampling, Clustering, and Transductive SVMs by precision, recall, and F-measure. Conclusions: Our system is a novel, state-of-the-art technique for efficiently extracting protein-protein interaction pairs.X116sciescopu

Fluorescence correlation spectroscopy of the binding of nucleotide excision repair protein XPC-hHr23B with DNA substrates

The interaction of the nucleotide excision repair (NER) protein dimeric complex XPC-hHR23B, which is implicated in the DNA damage recognition step, with three Cy3.5 labeled 90-bp double-stranded DNA substrates (unmodified, with a central unpaired region, and cholesterol modified) and a 90-mer single-strand DNA was investigated in solution by fluorescence correlation spectroscopy. Autocorrelation functions obtained in the presence of an excess of protein show larger diffusion times (τ d) than for free DNA, indicating the presence of DNA-protein bound complexes. The fraction of DNA bound (θ), as a way to describe the percentage of protein bound to DNA, was directly estimated from FCS data. A significantly stronger binding capability for the cholesterol modified substrate (78% DNA bound) than for other double-stranded DNA substrates was observed, while the lowest affinity was found for the single-stranded DNA (27%). This is in accordance with a damage recognition role of the XPC protein. The similar affinity of XPC for undamaged and 'bubble' DNA sub

Treatment adherence among sputum smear-positive pulmonary tuberculosis patients in mountainous areas in China

<p>Abstract</p> <p>Background</p> <p>We carried out an investigation in five provinces in China to assess treatment adherence and identify factors associated with insufficient treatment adherence in tuberculosis (TB) patients in mountainous, rural areas of China.</p> <p>Methods</p> <p>In each of the five provinces, all counties with > 80% mountainous area were stratified into three groups according to their gross domestic product. In each stratum, one county was randomly sampled. Study subjects were sampled from all smear positive TB cases registered in 2007 in the target counties. TB patients, village doctors, county doctors and directors of the TB prevention and control institutes were interviewed. Insufficient medication adherence was defined as taking less than 90% of anti-TB drug doses prescribed. Insufficient re-examination adherence was defined as having less than the recommended three sputum smear examinations during the treatment course.</p> <p>Results</p> <p>A minority of patients took drugs under direct observation: on average 29% during the intensive phase of treatment. In total, 524 TB patients were included, of whom 49 (9.4%) took less than 90% of all doses prescribed and 92 (17.6%) did not have all sputum smear examinations, with substantial variations between the provinces. In multivariable analysis, no direct observation of treatment during the intensive phase and the presence of adverse events were associated both with insufficient medication adherence and insufficient re-examination adherence. Overall, 79% of patients were adherent both to treatment and re-examinations.</p> <p>Conclusions</p> <p>In these remote and poor areas of China, the TB control program is not fully functioning according to the guidelines. The majority of patients are not treated under direct observation, while direct observation by health care staff was associated with better adherence, both to drug therapy and re-examinations. Insufficient adherence increases the risk of unsuccessful treatment outcomes and development of drug resistance. Measures should be taken urgently in these areas to strengthen implementation of the international Stop TB strategy.</p

Identification and Characterization of MicroRNAs in Asiatic Cotton (Gossypium arboreum L.)

To date, no miRNAs have been identified in the important diploid cotton species although there are several reports on miRNAs in upland cotton. In this study, we identified 73 miRNAs, belonging to 49 families, from Asiatic cotton using a well-developed comparative genome-based homologue search. Several of the predicted miRNAs were validated using quantitative real time PCR (qRT-PCR). The length of miRNAs varied from 18 to 22 nt with an average of 20 nt. The length of miRNA precursors also varied from 46 to 684 nt with an average of 138 ±120 nt. For a majority of Asiatic cotton miRNAs, there is only one member per family; however, multiple members were identified for miRNA 156, 414, 837, 838, 1044, 1533, 2902, 2868, 5021 and 5142 families. Nucleotides A and U were dominant, accounted for 62.95%, in the Asiatic cotton pre-miRNAs. The Asiatic cotton pre-miRNAs had high negative minimal folding free energy (MFE) and adjusted MFE (AMFE) and high MFE index (MFEI). Many miRNAs identified in Asiatic cotton suggest that miRNAs also play a similar regulatory mechanism in diploid cotton