INFLUENCE OF HYDRATED SODIUM CALCIUM ALUMINOSILICATE AND ACTIVATED CHARCOAL ON THE PHARMACOKINETICS OF SINGLE PULSE DOSING OF ENROFLOXACIN IN BROILER CHICKEN

ABSTRACTObjective: The present study was undertaken to evaluate the interaction kinetics of enrofloxacin, the commonly used antibacterial in poultry withmycotoxin binders namely hydrated sodium calcium aluminosilicate (HSCAS) and activated charcoal (AC), which have become inevitable componentsof poultry feed.Methods: Control group received normal feed free of toxin binder, whereas HSCAS and AC group were supplemented with HSCAS and AC at 0.5% infeed, respectively. Enrofloxacin was administered as single pulse dose (at 10 mg/kg) through drinking water to all the groups. Blood samples werecollected at predetermined time intervals after drug administration, and plasma was separated and analyzed for enrofloxacin concentrations usinghigh-performance liquid chromatography.Results: Significant decrease in area under the plasma concentration-time curve (AUC0-∞)was noticed in AC group when compared to control group(13.90±1.15 vs. 19.67±1.68 mg.h/ml), whereas HSCAS group (16.42±1.24 mg.h/ml) neither differed significantly from AC nor control group. Thevolume of distribution and clearance were significantly high in AC group when compared to control group (8.31±0.89 vs. 6.39±0.13 l/kg; 0.77±0.07 vs.0.53±0.05 l/h/kg). HSCAS group was intermediate and did not differ significantly from the other two groups (8.13±0.45 l/kg; 0.63±0.04 l/h/kg).However, volume of distribution at steady state was significantly high in both AC (10.42±1.09 l/kg) and HSCAS group (9.45±0.48 l/kg) when comparedto control group (7.21±0.20 l/kg). Maximum plasma concentration was significantly low (0.99±0.04, 0.97±0.06, 1.38±0.04 mg/ml) and time to reachmaximum plasma concentration was significantly delayed (7.33±0.42, 6.67±0.67, 4.33±0.67 h) in AC and HSCAS group when compared to controlgroup, respectively. The relative bioavailability was significantly low in both AC and HSCAS group (74.95±10.70, 88.88±15.03%) when comparedto control group. Pharmacokinetic/pharmacodynamic integration revealed that the dose of enrofloxacin (10 mg/kg) was capable of treating onlymoderately sensitive organisms (minimum inhibitory concentration ≤0.125 mg/ml) both in the presence and absence of toxin binder and higherdosage is needed for the less sensitive organism.Conclusion: The study revealed that the administration of enrofloxacin to HSCAS and AC supplemented broilers would lead to decrease in clinicalefficacy and promote the development of antimicrobial resistance. AC was found to interact more with enrofloxacin than HSCAS as observed fromthe PK parameters. Hence, careful adjustment of dosage or withdrawal of the usage of toxin binder containing either HSCAS or AC in feed duringenrofloxacin treatment is recommended.Keywords: Enrofloxacin, Pulse dosing, Hydrated sodium calcium aluminosilicate, Activated charcoal, Interaction kinetics

Analysis of Power Optimization Using SVL Techniques

Abstract--- Flip-flops are the major storage elements in all SOC's of digital design. They accommodate most of the power that has been applied to the chip. Flip-flop is one of the most power consumption components. It is important to reduce the power dissipation in both clock distribution networks and flip-flops. The power delay is mainly due to the clock delays. The delay of the flip-flops should be minimized for efficient implementation. The concept of this project is to reduce the power consumption and to increase the speed and functionality of the chip. This project moves around in replacing conventional master-slave based flip flop to a pulse triggered flip flop which acts as a tribute alternate for low power applications. Initially in the critical path the pulse generation controls logic along with SVL function. A simple transistor SVL design is used to reduce the circuit complexity. In this scheme transistor sizes and pulse generation circuit can be further reduce for power saving. Here UMC CMOS 180nm technology is use in SPICE tool to design the proposed structure. This would bring up the result in power saving approximately to 38.4%

Unique and conserved MicroRNAs in wheat chromosome 5D revealed by next-generation sequencing

MicroRNAs are a class of short, non-coding, single-stranded RNAs that act as post-transcriptional regulators in gene expression. miRNA analysis of Triticum aestivum chromosome 5D was performed on 454 GS FLX Titanium sequences of flow sorted chromosome 5D with a total of 3,208,630 good quality reads representing 1.34x and 1.61x coverage of the short (5DS) and long (5DL) arms of the chromosome respectively. In silico and structural analyses revealed a total of 55 miRNAs; 48 and 42 miRNAs were found to be present on 5DL and 5DS respectively, of which 35 were common to both chromosome arms, while 13 miRNAs were specific to 5DL and 7 miRNAs were specific to 5DS. In total, 14 of the predicted miRNAs were identified in wheat for the first time. Representation (the copy number of each miRNA) was also found to be higher in 5DL (1,949) compared to 5DS (1,191). Targets were predicted for each miRNA, while expression analysis gave evidence of expression for 6 out of 55 miRNAs. Occurrences of the same miRNAs were also found in Brachypodium distachyon and Oryza sativa genome sequences to identify syntenic miRNA coding sequences. Based on this analysis, two other miRNAs: miR1133 and miR167 were detected in B. distachyon syntenic region of wheat 5DS. Five of the predicted miRNA coding regions (miR6220, miR5070, miR169, miR5085, miR2118) were experimentally verified to be located to the 5D chromosome and three of them : miR2118, miR169 and miR5085, were shown to be 5D specific. Furthermore miR2118 was shown to be expressed in Chinese Spring adult leaves. miRNA genes identified in this study will expand our understanding of gene regulation in bread wheat

A Loss of Function Screen of Identified Genome-Wide Association Study Loci Reveals New Genes Controlling Hematopoiesis

The formation of mature cells by blood stem cells is very well understood at the cellular level and we know many of the key transcription factors that control fate decisions. However, many upstream signalling and downstream effector processes are only partially understood. Genome wide association studies (GWAS) have been particularly useful in providing new directions to dissect these pathways. A GWAS meta-analysis identified 68 genetic loci controlling platelet size and number. Only a quarter of those genes, however, are known regulators of hematopoiesis. To determine function of the remaining genes we performed a medium-throughput genetic screen in zebrafish using antisense morpholino oligonucleotides (MOs) to knock down protein expression, followed by histological analysis of selected genes using a wide panel of different hematopoietic markers. The information generated by the initial knockdown was used to profile phenotypes and to position candidate genes hierarchically in hematopoiesis. Further analysis of brd3a revealed its essential role in differentiation but not maintenance and survival of thrombocytes. Using the from-GWAS-to-function strategy we have not only identified a series of genes that represent novel regulators of thrombopoiesis and hematopoiesis, but this work also represents, to our knowledge, the first example of a functional genetic screening strategy that is a critical step toward obtaining biologically relevant functional data from GWA study for blood cell traits

Downstream signalling and specific inhibition of c-MET/HGF pathway in small cell lung cancer: implications for tumour invasion

The c-MET receptor can be overexpressed, amplified, or mutated in solid tumours including small cell lung cancer (SCLC). In c-MET-overexpressing SCLC cell line NCI-H69, hepatocyte growth factor (HGF) dramatically induced c-MET phosphorylation at phosphoepitopes pY1230/1234/1235 (catalytic tyrosine kinase), pY1003 (juxtamembrane), and also of paxillin at pY31 (CRKL-binding site). We utilised a global proteomics phosphoantibody array approach to identify further c-MET/HGF signal transduction intermediates in SCLC. Strong HGF induction of specific phosphorylation sites in phosphoproteins involved in c-MET/HGF signal transduction was detected, namely adducin-α [S724], adducin-γ [S662], CREB [S133], ERK1 [T185/Y187], ERK1/2 [T202/Y204], ERK2 [T185/Y187], MAPKK (MEK) 1/2 [S221/S225], MAPKK (MEK) 3/6 [S189/S207], RB [S612], RB1 [S780], JNK [T183/Y185], STAT3 [S727], focal adhesion kinase (FAK) [Y576/S722/S910], p38α-MAPK [T180/Y182], and AKT1[S473] and [T308]. Conversely, inhibition of phosphorylation by HGF in protein kinase C (PKC), protein kinase R (PKR), and also CDK1 was identified. Phosphoantibody-based immunohistochemical analysis of SCLC tumour tissue and microarray established the role of c-MET in SCLC biology. This supports a role of c-MET activation in tumour invasive front in the tumour progression and invasion involving FAK and AKT downstream. The c-MET serves as an attractive therapeutic target in SCLC, as shown through small interfering RNA (siRNA) and selective prototype c-MET inhibitor SU11274, inhibiting the phosphorylation of c-MET itself and its downstream molecules such as AKT, S6 kinase, and ERK1/2. Investigation of mechanisms of invasion and, ultimately, metastasis in SCLC would be very useful with these signal transduction molecules

Employing machine learning for reliable miRNA target identification in plants

<p>Abstract</p> <p>Background</p> <p>miRNAs are ~21 nucleotide long small noncoding RNA molecules, formed endogenously in most of the eukaryotes, which mainly control their target genes post transcriptionally by interacting and silencing them. While a lot of tools has been developed for animal miRNA target system, plant miRNA target identification system has witnessed limited development. Most of them have been centered around exact complementarity match. Very few of them considered other factors like multiple target sites and role of flanking regions.</p> <p>Result</p> <p>In the present work, a Support Vector Regression (SVR) approach has been implemented for plant miRNA target identification, utilizing position specific dinucleotide density variation information around the target sites, to yield highly reliable result. It has been named as p-TAREF (plant-Target Refiner). Performance comparison for p-TAREF was done with other prediction tools for plants with utmost rigor and where p-TAREF was found better performing in several aspects. Further, p-TAREF was run over the experimentally validated miRNA targets from species like <it>Arabidopsis</it>, <it>Medicago</it>, Rice and Tomato, and detected them accurately, suggesting gross usability of p-TAREF for plant species. Using p-TAREF, target identification was done for the complete Rice transcriptome, supported by expression and degradome based data. miR156 was found as an important component of the Rice regulatory system, where control of genes associated with growth and transcription looked predominant. The entire methodology has been implemented in a multi-threaded parallel architecture in Java, to enable fast processing for web-server version as well as standalone version. This also makes it to run even on a simple desktop computer in concurrent mode. It also provides a facility to gather experimental support for predictions made, through on the spot expression data analysis, in its web-server version.</p> <p>Conclusion</p> <p>A machine learning multivariate feature tool has been implemented in parallel and locally installable form, for plant miRNA target identification. The performance was assessed and compared through comprehensive testing and benchmarking, suggesting a reliable performance and gross usability for transcriptome wide plant miRNA target identification.</p

Perifosine as a Potential Novel Anti-Cancer Agent Inhibits EGFR/MET-AKT Axis in Malignant Pleural Mesothelioma

PI3K/AKT signalling pathway is aberrantly active and plays a critical role for cell cycle progression of human malignant pleural mesothelioma (MMe) cells. AKT is one of the important cellular targets of perifosine, a novel bio-available alkylphospholipid that has displayed significant anti-proliferative activity in vitro and in vivo in several human tumour model systems and is currently being tested in clinical trials.We tested Perifosine activity on human mesothelial cells and different mesothelioma cell lines, in order to provide evidence of its efficacy as single agent and combined therapy.We demonstrate here that perifosine, currently being evaluated as an anti-cancer agent in phase 1 and 2 clinical trials, caused a dose-dependent reduction of AKT activation, at concentrations causing MMe cell growth arrest. In this study we firstly describe that MMe cells express aside from AKT1 also AKT3 and that either the myristoylated, constitutively active, forms of the two proteins, abrogated perifosine-mediated cell growth inhibition. Moreover, we describe here a novel mechanism of perifosine that interferes, upstream of AKT, affecting EGFR and MET phosphorylation. Finally, we demonstrate a significant increase in cell toxicity when MMe cells were treated with perifosine in combination with cisplatin.This study provides a novel mechanism of action of perifosine, directly inhibiting EGFR/MET-AKT1/3 axis, providing a rationale for a novel translational approach to the treatment of MMe

Bacteria-responsive microRNAs regulate plant innate immunity by modulating plant hormone networks

MicroRNAs (miRNAs) are key regulators of gene expression in development and stress responses in most eukaryotes. We globally profiled plant miRNAs in response to infection of bacterial pathogen Pseudomonas syringae pv. tomato (Pst). We sequenced 13 small-RNA libraries constructed from Arabidopsis at 6 and 14 h post infection of non-pathogenic, virulent and avirulent strains of Pst. We identified 15, 27 and 20 miRNA families being differentially expressed upon Pst DC3000 hrcC, Pst DC3000 EV and Pst DC3000 avrRpt2 infections, respectively. In particular, a group of bacteria-regulated miRNAs targets protein-coding genes that are involved in plant hormone biosynthesis and signaling pathways, including those in auxin, abscisic acid, and jasmonic acid pathways. Our results suggest important roles of miRNAs in plant defense signaling by regulating and fine-tuning multiple plant hormone pathways. In addition, we compared the results from sequencing-based profiling of a small set of miRNAs with the results from small RNA Northern blot and that from miRNA quantitative RT-PCR. Our results showed that although the deep-sequencing profiling results are highly reproducible across technical and biological replicates, the results from deep sequencing may not always be consistent with the results from Northern blot or miRNA quantitative RT-PCR. We discussed the procedural differences between these techniques that may cause the inconsistency

Cross-Mapping Events in miRNAs Reveal Potential miRNA-Mimics and Evolutionary Implications

MicroRNAs (miRNAs) have important roles in various biological processes. miRNA cross-mapping is a prevalent phenomenon where miRNA sequence originating from one genomic region is mapped to another location. To have a better understanding of this phenomenon in the human genome, we performed a detailed analysis in this paper using public miRNA high-throughput sequencing data and all known human miRNAs. We observed widespread cross-mapping events between miRNA precursors (pre-miRNAs), other non-coding RNAs (ncRNAs) and the opposite strands of pre-miRNAs by analyzing the high-throughput sequencing data. Computational analysis on all known human miRNAs also confirmed that many of them could be involved in cross-mapping events. The processing or decay of both ncRNAs and pre-miRNA opposite strand transcripts may contribute to miRNA enrichment, although some might be miRNA-mimics due to miRNA mis-annotation. Comparing to canonical miRNAs, miRNAs involved in cross-mapping events between pre-miRNAs and other ncRNAs normally had shorter lengths (17–19 nt), lower prediction scores and were classified as pseudo miRNA precursors. Notably, 4.9% of all human miRNAs could be accurately mapped to the opposite strands of pre-miRNAs, which showed that both strands of the same genomic region had the potential to produce mature miRNAs and simultaneously implied some potential miRNA precursors. We proposed that the cross-mapping events are more complex than we previously thought. Sequence similarity between other ncRNAs and pre-miRNAs and the specific stem-loop structures of pre-miRNAs may provide evolutionary implications

Structural and electronic determinants of lytic polysaccharide monooxygenase reactivity on polysaccharide substrates

Lytic polysaccharide monooxygenases (LPMOs) are industrially important copper-dependent enzymes that oxidatively cleave polysaccharides. Here we present a functional and structural characterization of two closely related AA9-family LPMOs from Lentinus similis (LsAA9A) and Collariella virescens (CvAA9A). LsAA9A and CvAA9A cleave a range of polysaccharides, including cellulose, xyloglucan, mixed-linkage glucan and glucomannan. LsAA9A additionally cleaves isolated xylan substrates. The structures of CvAA9A and of LsAA9A bound to cellulosic and non-cellulosic oligosaccharides provide insight into the molecular determinants of their specificity. Spectroscopic measurements reveal differences in copper co-ordination upon the binding of xylan and glucans. LsAA9A activity is less sensitive to the reducing agent potential when cleaving xylan, suggesting that distinct catalytic mechanisms exist for xylan and glucan cleavage. Overall, these data show that AA9 LPMOs can display different apparent substrate specificities dependent upon both productive protein–carbohydrate interactions across a binding surface and also electronic considerations at the copper active site