With massive amounts of data being generated in electronic format, there is a need in basic science laboratories to adopt new methods for tracking and analyzing data. An electronic laboratory notebook (ELN) is not just a replacement for a paper lab notebook, it is a new method of storing and organizing data while maintaining the data entry flexibility and legal recording functions of paper notebooks. Paper notebooks are regarded as highly flexible since the user can configure it to store almost anything that can be written or physically pasted onto the pages. However, data retrieval and data sharing from paper notebooks are labor intensive processes and notebooks can be misplaced, a single point of failure that loses all entries in the volume. Additional features provided by electronic notebooks include searchable indices, data sharing, automatic archiving for security against loss and ease of data duplication. Furthermore, ELNs can be tasked with additional functions not commonly found in paper notebooks such as inventory control. While ELNs have been on the market for some time now, adoption of an ELN in academic basic science laboratories has been lagging. Issues that have restrained development and adoption of ELN in research laboratories are the sheer variety and frequency of changes in protocols with a need for the user to control notebook configuration outside the framework of professional IT staff support. In this commentary, we will look at some of the issues and experiences in academic laboratories that have proved challenging in implementing an electronic lab notebook.
The electronic laboratory notebook (ELN) has the potential to replace the paper notebook with a marked-up digital record that can be searched and shared. However, it is a challenge to achieve these benefits without losing the usability and flexibility of traditional paper notebooks. We investigate a blog-based platform that addresses the issues associated with the development of a flexible system for recording scientific research.
We chose a blog-based approach with the journal characteristics of traditional notebooks in mind, recognizing the potential for linking together procedures, materials, samples, observations, data, and analysis reports. We implemented the LabTrove blog system as a server process written in PHP, using a MySQL database to persist posts and other research objects. We incorporated a metadata framework that is both extensible and flexible while promoting consistency and structure where appropriate. Our experience thus far is that LabTrove is capable of providing a successful electronic laboratory recording system.
LabTrove implements a one-item one-post system, which enables us to uniquely identify each element of the research record, such as data, samples, and protocols. This unique association between a post and a research element affords advantages for monitoring the use of materials and samples and for inspecting research processes. The combination of the one-item one-post system, consistent metadata, and full-text search provides us with a much more effective record than a paper notebook. The LabTrove approach provides a route towards reconciling the tensions and challenges that lie ahead in working towards the long-term goals for ELNs. LabTrove, an electronic laboratory notebook (ELN) system from the Smart Research Framework, based on a blog-type framework with full access control, facilitates the scientific experimental recording requirements for reproducibility, reuse, repurposing, and redeployment.
Currently most biomedical labs in universities and government funded research institutions use paper lab notebooks for recording experimental data and spreadsheets for managing sample data. One consequence is that sample management and documenting experiments are viewed as separate and distinct activities, notwithstanding that samples and aliquots are an integral part of a majority of the experiments carried out by these labs.
Various drivers are pushing labs towards integrated management of sample data and experimental data. These include the ever increasing amounts of both kinds of data, the increasing adoption of online collaborative tools, changing expectations about online communication, and the increasing affordability of electronic lab notebooks and sample management software. There is now an opportunity for smaller labs, which have been slow to move from paper to electronic record keeping, to leapfrog better resourced commercial labs and lead the way in adopting the new generation of tools which permit integrated management of samples and experimental data and a range of tangible benefits to conducting research, including:
1. Fewer lost and mislabelled samples
2. Clearer visualization of relationships between samples and experiments
3. Reduction of experimental error
4. More effective search
5. Productivity gains
6. More efficient use of freezers, leading to cost reduction and enhanced sustainability
7. Improved archiving and enhanced memory at the lab and institutional levels
In order to exploit the vast body of currently inaccessible chemical information held in Electronic Laboratory Notebooks (ELNs) it is necessary not only to make it available but also to develop protocols for discovery, access and ultimately automatic processing. An aim of the Dial-a-Molecule Grand Challenge Network is to be able to draw on the body of accumulated chemical knowledge in order to predict or optimize the outcome of reactions. Accordingly the Network drew up a working group comprising informaticians, software developers and stakeholders from industry and academia to develop protocols and mechanisms to access and process ELN records. The work presented here constitutes the first stage of this process by proposing a tiered metadata system of knowledge, information and processing where each in turn addresses a) discovery, indexing and citation b) context and access to additional information and c) content access and manipulation. A compact set of metadata terms, called the elnItemManifest, has been derived and caters for the knowledge layer of this model. The elnItemManifest has been encoded as an XML schema and some use cases are presented to demonstrate the potential of this approach.
Research in organic chemistry generates samples of novel chemicals together with their properties and other related data. The involved scientists must be able to store this data and search it by chemical structure. There are commercial solutions for common needs like chemical registration systems or electronic lab notebooks. However for specific requirements of in-house databases and processes no such solutions exist. Another issue is that commercial solutions have the risk of vendor lock-in and may require an expensive license of a proprietary relational database management system. To speed up and simplify the development for applications that require chemical structure search capabilities, I have developed Molecule Database Framework. The framework abstracts the storing and searching of chemical structures into method calls. Therefore software developers do not require extensive knowledge about chemistry and the underlying database cartridge. This decreases application development time.
Molecule Database Framework is written in Java and I created it by integrating existing free and open-source tools and frameworks. The core functionality includes:
• Support for multi-component compounds (mixtures)
• Import and export of SD-files
• Optional security (authorization)
For chemical structure searching Molecule Database Framework leverages the capabilities of the Bingo Cartridge for PostgreSQL and provides type-safe searching, caching, transactions and optional method level security. Molecule Database Framework supports multi-component chemical compounds (mixtures).
Furthermore the design of entity classes and the reasoning behind it are explained. By means of a simple web application I describe how the framework could be used. I then benchmarked this example application to create some basic performance expectations for chemical structure searches and import and export of SD-files.
By using a simple web application it was shown that Molecule Database Framework successfully abstracts chemical structure searches and SD-File import and export to simple method calls. The framework offers good search performance on a standard laptop without any database tuning. This is also due to the fact that chemical structure searches are paged and cached. Molecule Database Framework is available for download on the projects web page on bitbucket: https://bitbucket.org/kienerj/moleculedatabaseframework.
Chemical structure search; Database; Framework; Open-source
Elastic blood vessels provide capacitance and pulse-wave dampening, which are critically important in a pulsatile circulatory system. By studying newborn mice with reduced (Eln+/−) or no (Eln−/−) elastin, we determined the effects of altered vessel elasticity on cardiovascular development and function. Eln−/− mice die within 72 hours of birth but are viable throughout fetal development when dramatic cardiovascular structural and hemodynamic changes occur. Thus, newborn Eln−/− mice provide unique insight into how a closed circulatory system develops when the arteries cannot provide the elastic recoil required for normal heart function. Compared with wild type, the Eln−/− aorta has a smaller unloaded diameter and thicker wall because of smooth muscle cell overproliferation and has greatly reduced compliance. Arteries in Eln−/− mice are also tortuous with stenoses and dilations. Left ventricular pressure is 2-fold higher than wild type, and heart function is impaired. Newborn Eln+/− mice, in contrast, have normal heart function despite left ventricular pressures 25% higher than wild type. The major vessels have smaller unloaded diameters and longer lengths. The Eln+/− aorta has additional smooth muscle cell layers that appear in the adventitia at or just before birth. These results show that the major adaptive changes in cardiovascular hemodynamics and in vessel wall structure seen in the adult Eln+/− mouse are defined in late fetal development. Together, these results show that reduced elastin in mice leads to adaptive remodeling, whereas the complete lack of elastin leads to pathological remodeling and death.
blood pressure; cardiovascular physiology; development; extracellular matrix; large artery stiffness
Obstructive vascular disease is an important health problem in the industrialized world. Through a series of molecular genetic studies, we demonstrated that loss-of-function mutations in one elastin allele cause an inherited obstructive arterial disease, supravalvular aortic stenosis (SVAS). To define the mechanism of elastin's effect, we generated mice hemizygous for the elastin gene (ELN +/-). Although ELN mRNA and protein were reduced by 50% in ELN +/- mice, arterial compliance at physiologic pressures was nearly normal. This discrepancy was explained by a paradoxical increase of 35% in the number of elastic lamellae and smooth muscle in ELN +/- arteries. Examination of humans with ELN hemizygosity revealed a 2. 5-fold increase in elastic lamellae and smooth muscle. Thus, ELN hemizygosity in mice and humans induces a compensatory increase in the number of rings of elastic lamellae and smooth muscle during arterial development. Humans are exquisitely sensitive to reduced ELN expression, developing profound arterial thickening and markedly increased risk of obstructive vascular disease.
Many transcription factors and DNA binding proteins play essential roles in the development of organs in which they are highly and/or specifically expressed. Embryonic stem cell (ESC)-associated transcript 15-1 (ECAT15-1) and ECAT15-2, also known as developmental pluripotency-associated 4 (Dppa4) and Dppa2, respectively, are enriched in mouse ESCs and preimplantation embryos, and their genes encode homologous proteins with a common DNA binding domain known as the SAP motif. Previously, ECAT15-1 was shown to be important in lung development, while it is dispensable in early development. In this study, we generated ECAT15-2 single and ECAT15-1 ECAT15-2 double knockout (double KO) mice and found that almost all mutants, like ECAT15-1 mutants, died around birth with respiratory defects. Paradoxically, the expression of neither ECAT15-1 nor ECAT15-2 was detected in lung organogenesis. Several genes, such as Nkx2-5, Gata4, and Pitx2, were downregulated in the ECAT15-2-null lung. On the other hand, genomic DNA of these genes showed inactive chromatin statuses in ECAT15-2-null ESCs, but not in wild-type ESCs. The chromatin immunoprecipitation (ChIP) assay revealed that ECAT15-2 binds to the regulatory region of Nkx2-5 in ESCs. These data suggest that ECAT15-2 has important roles in lung development, where it is no longer expressed, by leaving epigenetic marks from earlier developmental stages.
Elastin, the main component of elastic fibers, is synthesized only in early life and provides the blood vessels with their elastic properties. With aging, elastin is progressively degraded, leading to arterial enlargement, stiffening, and dysfunction. Also, elastin is a key regulator of vascular smooth muscle cell proliferation and migration during development since heterozygous mutations in its gene (Eln) are responsible for a severe obstructive vascular disease, supravalvular aortic stenosis, isolated or associated to Williams syndrome. Here, we have studied whether early elastin synthesis could also influence the aging processes, by comparing the structure and function of ascending aorta from 6- and 24-month-old Eln+/− and Eln+/+ mice. Eln+/− animals have high blood pressure and arteries with smaller diameters and more rigid walls containing additional although thinner elastic lamellas. Nevertheless, longevity of these animals is unaffected. In young adult Eln+/− mice, some features resemble vascular aging of wild-type animals: cardiac hypertrophy, loss of elasticity of the arterial wall through enhanced fragmentation of the elastic fibers, and extracellular matrix accumulation in the aortic wall, in particular in the intima. In Eln+/− animals, we also observed an age-dependent alteration of endothelial vasorelaxant function. On the contrary, Eln+/− mice were protected from several classical consequences of aging visible in aged Eln+/+ mice, such as arterial wall thickening and alteration of α1-adrenoceptor-mediated vasoconstriction. Our results suggest that early elastin expression and organization modify arterial aging through their impact on both vascular cell physiology and structure and mechanics of blood vessels.
Decreased expression of elastin results in smaller, less compliant arteries and high blood pressure. In mice, these differences become more significant with postnatal development. It is known that arterial size and compliance directly affect cardiac function, but the temporal changes in cardiac function have not been investigated in elastin insufficient mice. The aim of this study is to correlate changes in arterial size and compliance with cardiac function in wildtype (WT) and elastin haploinsufficient (Eln+/−) mice from birth to adulthood.
Ultrasound scans were performed at the ages of 3, 7, 14, 21, 30, 60, and 90 days on male and female WT and Eln+/− mice. 2-D ultrasound and pulse wave Doppler images were used to measure the dimensions and function of the left ventricle (LV), ascending aorta and carotid arteries.
Eln+/− arteries are smaller and less compliant at most ages, with significant differences from WT as early as 3 days old. Surprisingly, there are no correlations (R2 < 0.2) between arterial size and compliance with measures of LV hypertrophy or systolic function. There are weak correlations (0.2 < R2 < 0.5) between arterial size and compliance with measures of LV diastolic function.
Eln+/− mice have similar cardiac function to WT throughout postnatal development, demonstrating the remarkable ability of the developing cardiovascular system to adapt to mechanical and hemodynamic changes. Correlations between arterial size and compliance with diastolic function show that these measures may be useful indicators of early diastolic dysfunction.
Aorta; Carotid; Stiffness; Compliance; Diastolic dysfunction; Elastin
During the past several years, Baylor College of Medicine has made a substantial commitment to the use of information technology in support of its corporate and academic programs. The concept of an Integrated Academic Information Management System (IAIMS) has proved central in our planning, and the IAIMS activities that we have undertaken with funding from the National Library of Medicine have proved to be important extensions of our technology development. Here we describe our Virtual Notebook system, a conceptual and technologic framework for task coordination and information management in biomedical work groups. When fully developed and deployed, the Virtual Notebook will improve the functioning of basic and clinical research groups in the college, and it currently serves as a model for the longer-term development of our entire information management environment.
Elastolysis is central to progression of emphysema and aortic aneurysms. Characterization of steady-state enzyme kinetics of elastolysis is fettered by the insolubility of mature elastin and polydispersity of solubilized elastin. We prepared a fluor-tagged, 100 kDa fraction (fEln-100) from commercial α-elastin. It is soluble, less heterogenous in mass, cross-linked like mature elastin, and likely to retain the capacity of α-elastin to self assemble. fEln-100 has introduced the ability to compare quantitatively the apparent kcat and Km of elastases. For example, metalloelastase (MMP-12) displays higher apparent affinity for fEln-100, while MMP-2 displays faster catalytic turnover.
elastin degradation; fluorescence; enzyme kinetics; elastase; protease
The principal factors that lead to proliferation and pluripotency in embryonic stem cells (ESCs) have been vigorously investigated. However, the global network of factors and their full signaling cascade is still unclear. In this study, we found that ECAT11 (L1td1) is one of the ESC-associated transcripts harboring a truncated fragment of ORF-1, a component of theL1 retrotransposable element. We generated an ECAT11 knock-in mouse by replacing its coding region with green fluorescent protein. In the early stage of development, the fluorescence was observed at the inner cell mass of blastocysts and epiblasts. Despite this specific expression, ECAT11-null mice grow normally and are fertile. In addition, ECAT11 was dispensable for both the proliferation and pluripotency of ESCs.We found rapid and robust activation of ECAT11 in fibroblasts after the forced expression of transcription factors that can give rise pluripotency in somatic cells.However, iPS cells could be established from ECAT11-null fibroblasts. Our data demonstrate thedispensability of ECAT11/L1td1 in pluripotency, despite its specific expression.
Using either the principle of minimum energy or constant shear stress, a relation can be derived that predicts the diameters of branching vessels at a bifurcation. This relation, known as Murray's Law, has been shown to predict vessel diameters in a variety of cardiovascular systems from adult humans to developing chicks. The goal of this study is to investigate Murray's Law in vessels from mice that are haploinsufficient for the elastin protein (Eln+/−). Elastin is one of the major proteins in the blood vessel wall and is organized in concentric rings, known as lamellae, with smooth muscle cells (SMCs) around the vessel lumen. Eln+/− mice have an increased number of lamellae, as well as smaller, thinner vessels. It is possible that, due to decreased amounts of elastin available for vessel wall remodeling during development and in adulthood, Eln+/− vessels would not follow Murray's Law. We examined vessel bifurcations in six different physiologic regions, including the brain, heart, epidermis, ceocum (or cecum), testes, and intestines, in Eln+/− mice and wild-type (WT) littermates. All vessels were between 40 – 300 μm in diameter. We found that the diameters of both Eln+/− and WT vessels have an average of 13% error from the diameters predicted by Murray's Law, with no significant differences between genotypes or physiologic regions. The data suggest that vessels are optimized to follow Murray's Law, despite limitations on the proteins available for growth and remodeling of the vessel wall.
biomechanics; design and control of biological systems; physiological systems
The elastin gene (ELN) is implicated as a factor in both supravalvular aortic stenosis (SVAS) and Williams Beuren Syndrome (WBS), two diseases involving pronounced complications in mental or physical development. Although the complete spectrum of functional roles of the processed gene product remains to be established, these roles are inferred to be analogous in human and mouse. This view is supported by genomic sequence comparison, in which there are no large-scale differences in the ~1.8 Mb sequence block encompassing the common region deleted in WBS, with the exception of an overall reversed physical orientation between human and mouse.
Conserved synteny around ELN does not translate to a high level of conservation in the gene itself. In fact, ELN orthologs in mammals show more sequence divergence than expected for a gene with a critical role in development. The pattern of divergence is non-conventional due to an unusually high ratio of gaps to substitutions. Specifically, multi-sequence alignments of eight mammalian sequences reveal numerous non-aligning regions caused by species-specific insertions and deletions, in spite of the fact that the vast majority of aligning sites appear to be conserved and undergoing purifying selection.
The pattern of lineage-specific, in-frame insertions/deletions in the coding exons of ELN orthologous genes is unusual and has led to unique features of the gene in each lineage. These differences may indicate that the gene has a slightly different functional mechanism in mammalian lineages, or that the corresponding regions are functionally inert. Identified regions that undergo purifying selection reflect a functional importance associated with evolutionary pressure to retain those features.
To evaluate the prognostic significance of the international European LeukemiaNet (ELN) guidelines for reporting genetic alterations in acute myeloid leukemia (AML).
Patients and Methods
We analyzed 1,550 adults with primary AML, treated on Cancer and Leukemia Group B first-line trials, who had pretreatment cytogenetics and, for cytogenetically normal patients, mutational status of NPM1, CEBPA, and FLT3 available. We compared complete remission (CR) rates, disease-free survival (DFS), and overall survival (OS) among patients classified into the four ELN genetic groups (favorable, intermediate-I, intermediate-II, adverse) separately for 818 younger (age < 60 years) and 732 older (age ≥ 60 years) patients.
The percentages of younger versus older patients in the favorable (41% v 20%; P < .001), intermediate-II (19% v 30%; P < .001), and adverse (22% v 31%; P < .001) genetic groups differed. The favorable group had the best and the adverse group the worst CR rates, DFS, and OS in both age groups. Both intermediate groups had significantly worse outcomes than the favorable but better than the adverse group. Intermediate-I and intermediate-II groups in older patients had similar outcomes, whereas the intermediate-II group in younger patients had better OS but not better CR rates or DFS than the intermediate-I group. The prognostic significance of ELN classification was confirmed by multivariable analyses. For each ELN group, older patients had worse outcomes than younger patients.
The ELN classification clearly separates the genetic groups by outcome, supporting its use for risk stratification in clinical trials. Because they have different proportions of genetic alterations and outcomes, younger and older patients should be reported separately when using the ELN classification.
Elastin is an essential component of selected connective tissues that provides a unique physiological elasticity. Elastin may be considered a signature protein of lungs where matrix metalloprotease (MMP) -9-and -12, may be considered the signature proteases of the macrophages, which in part are responsible for tissue damage during disease progression. Thus, we hypothesized that a MMP-9/-12 generated fragment of elastin may be a relevant biochemical maker for lung diseases.
Elastin fragments were identified by mass-spectrometry and one sequence, generated by MMP-9 and -12 (ELN-441), was selected for monoclonal antibody generation and used in the development of an ELISA. Soluble and insoluble elastin from lung was cleaved in vitro and the time-dependent release of fragments was assessed in the ELN-441 assay. The release of ELN-441 in human serum from patients with chronic obstructive pulmonary disease (COPD) (n = 10) and idiopathic pulmonary fibrosis (IPF) (n = 29) were compared to healthy matched controls (n = 11).
The sequence ELN-441 was exclusively generated by MMP-9 and -12 and was time-dependently released from soluble lung elastin. ELN-441 levels were 287% higher in patients diagnosed with COPD (p < 0.001) and 124% higher in IPF patients (p < 0.0001) compared with controls. ELN-441 had better diagnostic value in COPD patients (AUC 97%, p = 0.001) than in IPF patients (AUC 90%, p = 0.0001). The odds ratios for differentiating controls from COPD or IPF were 24 [2.06–280] for COPD and 50 [2.64–934] for IPF.
MMP-9 and -12 time-dependently released the ELN-441 epitope from elastin. This fragment was elevated in serum from patients with the lung diseases IPF and COPD, however these data needs to be validated in larger clinical settings.
Elastin; Extracellular matrix remodeling; Biochemical marker; Neoepitope; COPD; IPF; MMP
Part diary, part scientific record, biological field notebooks often contain details necessary to understanding the location and environmental conditions existent during collecting events. Despite their clear value for (and recent use in) global change studies, the text-mining outputs from field notebooks have been idiosyncratic to specific research projects, and impossible to discover or re-use. Best practices and workflows for digitization, transcription, extraction, and integration with other sources are nascent or non-existent. In this paper, we demonstrate a workflow to generate structured outputs while also maintaining links to the original texts. The first step in this workflow was to place already digitized and transcribed field notebooks from the University of Colorado Museum of Natural History founder, Junius Henderson, on Wikisource, an open text transcription platform. Next, we created Wikisource templates to document places, dates, and taxa to facilitate annotation and wiki-linking. We then requested help from the public, through social media tools, to take advantage of volunteer efforts and energy. After three notebooks were fully annotated, content was converted into XML and annotations were extracted and cross-walked into Darwin Core compliant record sets. Finally, these recordsets were vetted, to provide valid taxon names, via a process we call “taxonomic referencing.” The result is identification and mobilization of 1,068 observations from three of Henderson’s thirteen notebooks and a publishable Darwin Core record set for use in other analyses. Although challenges remain, this work demonstrates a feasible approach to unlock observations from field notebooks that enhances their discovery and interoperability without losing the narrative context from which those observations are drawn.
“Compose your notes as if you were writing a letter to someone a century in the future.”
Perrine and Patton (2011)
Field notes; notebooks; crowd sourcing; digitization; biodiversity; transcription; text-mining; Darwin Core; Junius Henderson; annotation; taxonomic referencing; natural history; Wikisource; Colorado; species occurrence records
This article introduces a desktop application, named Concierge, for managing personal digital research resources. Using simple operations, it enables storage of various types of files and indexes them based on content descriptions. A key feature of the software is a high level of extensibility. By installing optional plug-ins, users can customize and extend the usability of the software based on their needs. In this paper, we also introduce a few optional plug-ins: literature management, electronic laboratory notebook, and XooNlps client plug-ins. XooNIps is a content management system developed to share digital research resources among neuroscience communities. It has been adopted as the standard database system in Japanese neuroinformatics projects. Concierge, therefore, offers comprehensive support from management of personal digital research resources to their sharing in open-access neuroinformatics databases such as XooNIps. This interaction between personal and open-access neuroinformatics databases is expected to enhance the dissemination of digital research resources. Concierge is developed as an open source project; Mac OS X and Windows XP versions have been released at the official site (http://concierge.sourceforge.jp).
software; resource management; resource sharing
Randomized, prospective trials involving multi-institutional collaboration have become a central part of clinical and translational research. However, data management and coordination of multi-center studies is a complex process that involves developing systems for data collection and quality control, tracking data queries and resolutions, as well as developing communication procedures. We describe DADOS-Prospective, an open-source Web-based application for collecting and managing prospective data on human subjects for clinical and translational trials. DADOS-Prospective not only permits users to create new clinical research forms (CRF) and supports electronic signatures, but also offers the advantage of containing, in a single environment, raw research data in downloadable spreadsheet format, source documentation and regulatory files stored in PDF format, and audit trails.
Feedback from formal and field usability tests was used to guide the design and development of DADOS-Prospective. To date, DADOS-Prospective has been implemented in five prospective clinical studies at our institution. Four of these studies are still in the CRF creation phase and one study has been entirely launched.
DADOS-Prospective has significant advantages over existing Web-based data collecting programs. At our institution, it has been demonstrated to be an efficient tool for prospective clinical studies.
Laboratories that produce protein reagents for research and development face the challenge of deciding whether to track batch-related data using simple file based storage mechanisms (e.g. spreadsheets and notebooks), or commit the time and effort to install, configure and maintain a more complex laboratory information management system (LIMS). Managing reagent data stored in files is challenging because files are often copied, moved, and reformatted. Furthermore, there is no simple way to query the data if/when questions arise. Commercial LIMS often include additional modules that may be paid for but not actually used, and often require software expertise to truly customize them for a given environment.
This web-application allows small to medium-sized protein production groups to track data related to plasmid DNA, conditioned media samples (supes), cell lines used for expression, and purified protein information, including method of purification and quality control results. In addition, a request system was added that includes a means of prioritizing requests to help manage the high demand of protein production resources at most organizations. ProteinTracker makes extensive use of existing open-source libraries and is designed to track essential data related to the production and purification of proteins.
ProteinTracker is an open-source web-based application that provides organizations with the ability to track key data involved in the production and purification of proteins and may be modified to meet the specific needs of an organization. The source code and database setup script can be downloaded from http://sourceforge.net/projects/proteintracker. This site also contains installation instructions and a user guide. A demonstration version of the application can be viewed at http://www.proteintracker.org.
Protein; Production; Purification; Reagent; Tracking; Prioritization; Web; Application
Supravalvular aortic stenosis (SVAS) is caused by mutations in the elastin (ELN) gene and is characterized by abnormal proliferation of vascular smooth muscle cells (SMCs) that can lead to narrowing or blockage of the ascending aorta and other arterial vessels. Availability of patient-specific SMCs may facilitate studying disease mechanisms and developing novel therapeutic interventions.
Methods and Results
Here, we report the development of a human induced pluripotent stem cell (iPSC) line from a patient with SVAS caused by the premature termination in exon 10 of the ELN gene due to an exon 9 4-nucleotide insertion. We showed that SVAS iPSC-derived SMCs (iPSC-SMCs) had significantly fewer organized networks of smooth muscle alpha actin (SM α-actin) filament bundles, a hallmark of mature contractile SMCs, compared to control iPSC-SMCs. Addition of elastin recombinant protein or enhancement of small GTPase RhoA signaling was able to rescue the formation of SM α-actin filament bundles in SVAS iPSC-SMCs. Cell counts and BrdU analysis revealed a significantly higher proliferation rate in SVAS iPSC-SMCs than control iPSC-SMCs. Furthermore, SVAS iPSC-SMCs migrated at a markedly higher rate to the chemotactic agent platelet-derived growth factor (PDGF) in comparison with the control iPSC-SMCs. We also provided evidence that elevated activity of extracellular signal-regulated kinase 1/2 (ERK1/2) is required for hyper-proliferation of SVAS iPSC-SMCs. The phenotype was confirmed in iPSC-SMCs generated from a patient with deletion of elastin due to Williams-Beuren syndrome (WBS).
Thus, SVAS iPSC-SMCs recapitulate key pathological features of patients with SVAS and may provide a promising strategy to study disease mechanisms and to develop novel therapies.
elastin; induced pluripotent stem cells; smooth muscle alpha actin filament bundle; smooth muscle cells; supravalvular aortic stenosis
Restenosis remains the main complication of balloon angioplasty and/or stent implantation. Preclinical testing of new pharmacologic agents preventing restenosis largely rely on porcine models, where restenosis is assessed after endothelial abrasion of the arterial wall or stent implantation. We combined endothelial cell denudation and implantation of stents to develop a new clinically relevant porcine model of restenosis, and used this model to determine the effects of an α4 integrin inhibitor, ELN 457946, on restenosis. Balloon-angioplasty endothelial cell denudation and subsequent implantation of bare metal stents in the left anterior descending coronary, iliac, and left common carotid arteries was performed in domestic pigs, treated with vehicle or ELN 457946, once weekly via subcutaneous injections, for four weeks. After 1 month, histopathology and morphometric analyses of the arteries showed complete healing and robust, consistent restenotic response in stented arteries. Treatment with ELN 457946 resulted in a reduction in the neointimal response, with decreases in area percent stenosis between 12% in coronary arteries and 30% in peripheral vessels. This is the first description of a successful pig model combining endothelial cell denudation and bare metal stent implantation. This new double injury model may prove particularly useful to assess pharmacological effects of drug candidates on restenosis, in coronary and/or peripheral arteries. Furthermore, the ELN 457946 α4 integrin inhibitor, administered subcutaneously, reduced inflammation and restenosis in stented coronary and peripheral arteries in pigs, therefore representing a promising systemic therapeutic approach in reducing restenosis in patients undergoing angioplasty and/or stent implantation.
Multiple sequence alignment (MSA) is a cornerstone of modern molecular biology and represents a unique means of investigating the patterns of conservation and diversity in complex biological systems. Many different algorithms have been developed to construct MSAs, but previous studies have shown that no single aligner consistently outperforms the rest. This has led to the development of a number of ‘meta-methods’ that systematically run several aligners and merge the output into one single solution. Although these methods generally produce more accurate alignments, they are inefficient because all the aligners need to be run first and the choice of the best solution is made a posteriori. Here, we describe the development of a new expert system, AlexSys, for the multiple alignment of protein sequences. AlexSys incorporates an intelligent inference engine to automatically select an appropriate aligner a priori, depending only on the nature of the input sequences. The inference engine was trained on a large set of reference multiple alignments, using a novel machine learning approach. Applying AlexSys to a test set of 178 alignments, we show that the expert system represents a good compromise between alignment quality and running time, making it suitable for high throughput projects. AlexSys is freely available from http://alnitak.u-strasbg.fr/∼aniba/alexsys.
To determine if antagonizing miR 29 enhances elastin (ELN) levels in cells and tissues lacking ELN. Methods and
miR-29 mimics reduced ELN levels in fibroblasts and smooth muscle cells, whereas miR-29 inhibition increased ELN levels. Antagonism of miR-29 also increased ELN levels in cells from patients haploinsufficient for ELN and in bioengineered human vessels.
miR-29 antagonism may promote increased ELN levels during conditions of ELN deficiencies.
microRNA; elastin; smooth muscle; fibroblasts; synthetic vessels