Identification of Schistosoma mansoni microRNAs

Background: MicroRNAs (miRNAs) constitute a class of single-stranded RNAs which play a crucial role in regulating development and controlling gene expression by targeting mRNAs and triggering either translation repression or messenger RNA (mRNA) degradation. miRNAs are widespread in eukaryotes and to date over 14,000 miRNAs have been identified by computational and experimental approaches. Several miRNAs are highly conserved across species. In Schistosoma, the full set of miRNAs and their expression patterns during development remain poorly understood. Here we report on the development and implementation of a homology-based detection strategy to search for miRNA genes in Schistosoma mansoni. In addition, we report results on the experimental detection of miRNAs by means of cDNA cloning and sequencing of size-fractionated RNA samples. Results: Homology search using the high-throughput pipeline was performed with all known miRNAs in miRBase. A total of 6,211 mature miRNAs were used as reference sequences and 110 unique S. mansoni sequences were returned by BLASTn analysis. The existing mature miRNAs that produced these hits are reported, as well as the locations of the homologous sequences in the S. mansoni genome. All BLAST hits aligned with at least 95% of the miRNA sequence, resulting in alignment lengths of 19-24 nt. Following several filtering steps, 15 potential miRNA candidates were identified using this approach. By sequencing small RNA cDNA libraries from adult worm pairs, we identified 211 novel miRNA candidates in the S. mansoni genome. Northern blot analysis was used to detect the expression of the 30 most frequent sequenced miRNAs and to compare the expression level of these miRNAs between the lung stage schistosomula and adult worm stages. Expression of 11 novel miRNAs was confirmed by northern blot analysis and some presented a stage-regulated expression pattern. Three miRNAs previously identified from S. japonicum were also present in S. mansoni. Conclusion: Evidence for the presence of miRNAs in S. mansoni is presented. The number of miRNAs detected by homology-based computational methods in S. mansoni is limited due to the lack of close relatives in the miRNA repository. In spite of this, the computational approach described here can likely be applied to the identification of pre-miRNA hairpins in other organisms. Construction and analysis of a small RNA library led to the experimental identification of 14 novel miRNAs from S. mansoni through a combination of molecular cloning, DNA sequencing and expression studies. Our results significantly expand the set of known miRNAs in multicellular parasites and provide a basis for understanding the structural and functional evolution of miRNAs in these metazoan parasites

South African dyslipidaemia guideline consensus statement

The European Society of Cardiology together with the European Atherosclerosis Society published updated dyslipidaemia guidelines in 2011. SA Heart and the Lipid and Atherosclerosis Society of Southern Africa officially adopt these guidelines. This statement adapts aspects of the guidelines to the South African situation. Using the updated Framingham risk charts, interventional strategies are based according to the cardiovascular risk score and low-density lipoprotein cholesterol (LDL-C) levels. The Framingham risk score refers to the 10-year risk of any cardiovascular event, and includes four categories of risk. Treatment targets are those of the European guidelines. The LDL-C goal is 1.8mmol/l for the very high-risk group (>30%), 2.5mmol/l for the high-risk group (15 - 30%), and 3mmol/l for those below 15% risk. Intensive management of dyslipidaemia in South Africa will significantly reduce the cardiovascular disease health burden

South African Dyslipidaemia Guideline Consensus Statement: A joint statement from the South African Heart Association (SA Heart) and the Lipid and Atherosclerosis Society of Southern Africa (LASSA)

The European Society of Cardiology together with the European Atherosclerosis Society published updated dyslipidaemia guidelines in 2011. SA Heart and the Lipid and Atherosclerosis Society of Southern Africa officially adopt these guidelines. This statement adapts aspects of the guidelines to the South African situation. Using the updated Framingham risk charts, interventional strategies are based according to the cardiovascular risk score and low-density lipoprotein cholesterol (LDL-C) levels. The Framingham risk score refers to the 10-year risk of any cardiovascular event, and includes four categories of risk. Treatment targets are those of the European guidelines. The LDL-C goal is 1.8 mmol/l for the very high-risk group (>30%), 2.5 mmol/l for the high-risk group (15 - 30%), and 3 mmol/l for those below 15% risk. Intensive management of dyslipidaemia in South Africa will significantly reduce the cardiovascular disease health burden

Role of surface nickel content on human cell cytoskeleton formation on Nitinol

Cell activity on an implant surface can be modulated by cues such as topography, chemistry or stiffness(1,2). For Ni-Ti alloy this is achieved mainly by alteration in chemistry. However, high nickel concentrations may be a concern in the use Nitinol on a larger scale. Current reports on Nitinol bring contradictory data(3-5) suggesting that high nickel content is not particularly dangerous and nickel-titanium alloys are safe to be used. On the other hand it was shown that nickel has a toxic effects on cells(6). Nevertheless, shape memory effects and pseudo-elasticity could support different treatments (e.g. scoliosis) and currently, Nitinol is used to produce porous foams and coatings (Actipore™), pins, clamps and intramedullary nails. In this paper authors investigated a role for nickel surface concentration on influencing cell behaviour e.g. cytoskeleton formation and organization in vitro

Prior Coronary Artery Bypass Graft Surgery and Outcome After Percutaneous Coronary Intervention: An Observational Study From the Pan-London Percutaneous Coronary Intervention Registry.

Background Limited information exists regarding procedural success and clinical outcomes in patients with previous coronary artery bypass grafting (CABG) undergoing percutaneous coronary intervention (PCI). We sought to compare outcomes in patients undergoing PCI with or without CABG. Methods and Results This was an observational cohort study of 123 780 consecutive PCI procedures from the Pan-London (UK) PCI registry from 2005 to 2015. The primary end point was all-cause mortality at a median follow-up of 3.0 years (interquartile range, 1.2-4.6 years). A total of 12 641(10.2%) patients had a history of previous CABG, of whom 29.3% (n=3703) underwent PCI to native vessels and 70.7% (n=8938) to bypass grafts. There were significant differences in the demographic, clinical, and procedural characteristics of these groups. The risk of mortality during follow-up was significantly higher in patients with prior CABG (23.2%; P=0.0005) compared with patients with no prior CABG (12.1%) and was seen for patients who underwent either native vessel (20.1%) or bypass graft PCI (24.2%; P<0.0001). However, after adjustment for baseline characteristics, there was no significant difference in outcomes seen between the groups when PCI was performed in native vessels in patients with previous CABG (hazard ratio [HR],1.02; 95%CI, 0.77-1.34; P=0.89), but a significantly higher mortality was seen among patients with PCI to bypass grafts (HR,1.33; 95% CI, 1.03-1.71; P=0.026). This was seen after multivariate adjustment and propensity matching. Conclusions Patients with prior CABG were older with greater comorbidities and more complex procedural characteristics, but after adjustment for these differences, the clinical outcomes were similar to the patients undergoing PCI without prior CABG. In these patients, native-vessel PCI was associated with better outcomes compared with the treatment of vein grafts

Controlling cell behavior through the design of polymer surfaces

Polymers have gained a remarkable place in the biomedical fi eld as materials for the fabrication of various devices and for tissue engineering applications. The initial acceptance or rejection of an implantable device is dictated by the crosstalk of the material surface with the bioentities present in the physiological environment. Advances in microfabrication and nanotechnology offer new tools to investigate the complex signaling cascade induced by the components of the extracellular matrix and consequently allow cellular responses to be tailored through the mimicking of some elements of the signaling paths. Patterning methods and selective chemical modifi cation schemes at different length scales can provide biocompatible surfaces that control cellular interactions on the micrometer and sub-micrometer scales on which cells are organized. In this review, the potential of chemically and topographically structured micro- and nanopolymer surfaces are discussed in hopes of a better understanding of cell–biomaterial interactions, including the recent use of biomimetic approaches or stimuli-responsive macromolecules. Additionally, the focus will be on how the knowledge obtained using these surfaces can be incorporated to design biocompatible materials for various biomedical applications, such as tissue engineering, implants, cell-based biosensors, diagnostic systems, and basic cell biology. The review focusses on the research carried out during the last decade.The research leading to these results has received partial funding from the European Union Seventh Framework Programme (FP7/2007-2013) under grant agreement no. NMP4-SL-2009-229292 and by the FCT projects PTDC/FIS/68517/2006, PTDC/QUI/69263/2006, PTDC/FIS/68209/2006, and PTDC/QUI/68804/2006

microRNA 1307 Is a Potential Target for SARS-CoV-2 Infection: An in Vitro Model

microRNAs (miRs) are proposed as critical molecular targets in SARS-CoV-2 infection. Our recent in silico studies identified seven SARS-CoV-2 specific miR-like sequences, which are highly conserved with humans, including miR-1307-3p, with critical roles in COVID-19. In this current study, Vero cells were infected with SARS-CoV-2, and miR expression profiles were thereafter confirmed by qRT-PCR. miR-1307-3p was the most highly expressed miR in the infected cells; we, therefore, transiently inhibited its expression in both infected and uninfected cells. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) cell proliferation assay assessed cell viability following SARS-CoV-2 infection, identifying that miR-1307 expression is inversely correlated with cell viability. Lastly, changes in miR-1307-dependent pathways were analyzed through a detailed miRNOME and associated in silico analysis. In addition to our previously identified miRs, including miR-1307-3p, the upregulation of miR-193a-5p, miR-5100, and miR-23a-5p and downregulation of miR-130b-5p, miR34a-5p, miR-505-3p, miR181a-2-3p, miR-1271-5p, miR-598-3p, miR-34c-3p, and miR-129-5p were also established in Vero cells related to general lung disease-related genes following SARS-CoV-2 infection. Targeted anti-miR-1307-3p treatment rescued cell viability in infection when compared to SARS CoV-2 mediated cell cytotoxicity only. We furthermore identified by in silico analysis that miR-1307-3p is conserved in all SARS-CoV-2 sequences/strains, except in the BA.2 variant, possibly contributing to the lower disease severity of this variant, which warrants further investigation. Small RNA seq analysis was next used to evaluate alterations in the miRNOME, following miR-1307-3p manipulation, identifying critical pathobiological pathways linked to SARS-CoV-2 infection-mediated upregulation of this miR. On the basis of our findings, miRNAs like miR-1307-3p play a critical role in SARS-CoV-2 infection, including via effects on disease progression and severity

A Model of a MAPK•Substrate Complex in an Active Conformation: A Computational and Experimental Approach

The mechanisms by which MAP kinases recognize and phosphorylate substrates are not completely understood. Efforts to understand the mechanisms have been compromised by the lack of MAPK-substrate structures. While MAPK-substrate docking is well established as a viable mechanism for bringing MAPKs and substrates into close proximity the molecular details of how such docking promotes phosphorylation is an unresolved issue. In the present study computer modeling approaches, with restraints derived from experimentally known interactions, were used to predict how the N-terminus of Ets-1 associates with ERK2. Interestingly, the N-terminus does not contain a consensus-docking site ((R/K)2-3-X2-6-ΦA-X-ΦB, where Φ is aliphatic hydrophobic) for ERK2. The modeling predicts that the N-terminus of Ets-1 makes important contributions to the stabilization of the complex, but remains largely disordered. The computer-generated model was used to guide mutagenesis experiments, which support the notion that Leu-11 and possibly Ile-13 and Ile-14 of Ets-1 1-138 (Ets) make contributions through binding to the hydrophobic groove of the ERK2 D-recruiting site (DRS). Based on the modeling, a consensus-docking site was introduced through the introduction of an arginine at residue 7, to give the consensus 7RK-X2-ΦA-X-ΦB13. This results in a 2-fold increase in kcat/Km for the phosphorylation of Ets by ERK2. Similarly, the substitution of the N-terminus for two different consensus docking sites derived from Elk-1 and MKK1 also improves kcat/Km by two-fold compared to Ets. Disruption of the N-terminal docking through deletion of residues 1-23 of Ets results in a 14-fold decrease in kcat/Km, with little apparent change in kcat. A peptide that binds to the DRS of ERK2 affects Km, but not kcat. Our kinetic analysis suggests that the unstructured N-terminus provides 10-fold uniform stabilization of the ground state ERK2•Ets•MgATP complex and intermediates of the enzymatic reaction

XplorSeq: A software environment for integrated management and phylogenetic analysis of metagenomic sequence data

<p>Abstract</p> <p>Background</p> <p>Advances in automated DNA sequencing technology have accelerated the generation of metagenomic DNA sequences, especially environmental ribosomal RNA gene (rDNA) sequences. As the scale of rDNA-based studies of microbial ecology has expanded, need has arisen for software that is capable of managing, annotating, and analyzing the plethora of diverse data accumulated in these projects.</p> <p>Results</p> <p>XplorSeq is a software package that facilitates the compilation, management and phylogenetic analysis of DNA sequences. XplorSeq was developed for, but is not limited to, high-throughput analysis of environmental rRNA gene sequences. XplorSeq integrates and extends several commonly used UNIX-based analysis tools by use of a Macintosh OS-X-based graphical user interface (GUI). Through this GUI, users may perform basic sequence import and assembly steps (base-calling, vector/primer trimming, contig assembly), perform BLAST (Basic Local Alignment and Search Tool; <abbrgrp><abbr bid="B1">1</abbr><abbr bid="B2">2</abbr><abbr bid="B3">3</abbr></abbrgrp>) searches of NCBI and local databases, create multiple sequence alignments, build phylogenetic trees, assemble Operational Taxonomic Units, estimate biodiversity indices, and summarize data in a variety of formats. Furthermore, sequences may be annotated with user-specified meta-data, which then can be used to sort data and organize analyses and reports. A document-based architecture permits parallel analysis of sequence data from multiple clones or amplicons, with sequences and other data stored in a single file.</p> <p>Conclusion</p> <p>XplorSeq should benefit researchers who are engaged in analyses of environmental sequence data, especially those with little experience using bioinformatics software. Although XplorSeq was developed for management of rDNA sequence data, it can be applied to most any sequencing project. The application is available free of charge for non-commercial use at <url>http://vent.colorado.edu/phyloware</url>.</p