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In JoVE (1)
Other Publications (10)
Articles by Shannon Meredith in JoVE
JoVE Bioengineering
Hydrophobic Salt-modified Nafion for Enzyme Immobilization and Stabilization
Departments of Chemistry and Materials Science and Engineering, University of Utah
This article will describe the procedure for synthesizing a hydrophobically modified Nafion enzyme immobilization membrane and how to immobilize proteins and/or enzymes within the membrane and test their specific activity.
Other articles by Shannon Meredith on PubMed
Flt3L Controls the Development of Radiosensitive Dendritic Cells in the Meninges and Choroid Plexus of the Steady-state Mouse Brain
Antigen-presenting cells in the disease-free brain have been identified primarily by expression of antigens such as CD11b, CD11c, and MHC II, which can be shared by dendritic cells (DCs), microglia, and monocytes. In this study, starting with the criterion of Flt3 (FMS-like receptor tyrosine kinase 3)-dependent development, we characterize the features of authentic DCs within the meninges and choroid plexus in healthy mouse brains. Analyses of morphology, gene expression, and antigen-presenting function established a close relationship between meningeal and choroid plexus DCs (m/chDCs) and spleen DCs. DCs in both sites shared an intrinsic requirement for Flt3 ligand. Microarrays revealed differences in expression of transcripts encoding surface molecules, transcription factors, pattern recognition receptors, and other genes in m/chDCs compared with monocytes and microglia. Migrating pre-DC progenitors from bone marrow gave rise to m/chDCs that had a 5-7-d half-life. In contrast to microglia, DCs actively present self-antigens and stimulate T cells. Therefore, the meninges and choroid plexus of a steady-state brain contain DCs that derive from local precursors and exhibit a differentiation and antigen-presenting program similar to spleen DCs and distinct from microglia.
Open Innovation for Phenotypic Drug Discovery: The PD2 Assay Panel
Phenotypic lead generation strategies seek to identify compounds that modulate complex, physiologically relevant systems, an approach that is complementary to traditional, target-directed strategies. Unlike gene-specific assays, phenotypic assays interrogate multiple molecular targets and signaling pathways in a target "agnostic" fashion, which may reveal novel functions for well-studied proteins and discover new pathways of therapeutic value. Significantly, existing compound libraries may not have sufficient chemical diversity to fully leverage a phenotypic strategy. To address this issue, Eli Lilly and Company launched the Phenotypic Drug Discovery Initiative (PD(2)), a model of open innovation whereby external research groups can submit compounds for testing in a panel of Lilly phenotypic assays. This communication describes the statistical validation, operations, and initial screening results from the first PD(2) assay panel. Analysis of PD(2) submissions indicates that chemical diversity from open source collaborations complements internal sources. Screening results for the first 4691 compounds submitted to PD(2) have confirmed hit rates from 1.6% to 10%, with the majority of active compounds exhibiting acceptable potency and selectivity. Phenotypic lead generation strategies, in conjunction with novel chemical diversity obtained via open-source initiatives such as PD(2), may provide a means to identify compounds that modulate biology by novel mechanisms and expand the innovation potential of drug discovery.
Proteomics, Ultrastructure, and Physiology of Hippocampal Synapses in a Fragile X Syndrome Mouse Model Reveal Presynaptic Phenotype
Fragile X syndrome (FXS), the most common form of hereditary mental retardation, is caused by a loss-of-function mutation of the Fmr1 gene, which encodes fragile X mental retardation protein (FMRP). FMRP affects dendritic protein synthesis, thereby causing synaptic abnormalities. Here, we used a quantitative proteomics approach in an FXS mouse model to reveal changes in levels of hippocampal synapse proteins. Sixteen independent pools of Fmr1 knock-out mice and wild type mice were analyzed using two sets of 8-plex iTRAQ experiments. Of 205 proteins quantified with at least three distinct peptides in both iTRAQ series, the abundance of 23 proteins differed between Fmr1 knock-out and wild type synapses with a false discovery rate (q-value) <5%. Significant differences were confirmed by quantitative immunoblotting. A group of proteins that are known to be involved in cell differentiation and neurite outgrowth was regulated; they included Basp1 and Gap43, known PKC substrates, and Cend1. Basp1 and Gap43 are predominantly expressed in growth cones and presynaptic terminals. In line with this, ultrastructural analysis in developing hippocampal FXS synapses revealed smaller active zones with corresponding postsynaptic densities and smaller pools of clustered vesicles, indicative of immature presynaptic maturation. A second group of proteins involved in synaptic vesicle release was up-regulated in the FXS mouse model. In accordance, paired-pulse and short-term facilitation were significantly affected in these hippocampal synapses. Together, the altered regulation of presynaptically expressed proteins, immature synaptic ultrastructure, and compromised short-term plasticity points to presynaptic changes underlying glutamatergic transmission in FXS at this stage of development.
A Normal Platelet Count May Not Be Enough: the Impact of Admission Platelet Count on Mortality and Transfusion in Severely Injured Trauma Patients
Platelets play a central role in hemostasis after trauma. However, the platelet count of most trauma patients does not fall below the normal range (100-450 × 10(9)/L), and as a result, admission platelet count has not been adequately investigated as a predictor of outcome. The purpose of this study was to examine the relationship between admission platelet count and outcomes after trauma.
The Association of Blood Component Use Ratios with the Survival of Massively Transfused Trauma Patients with and Without Severe Brain Injury
The effect of blood component ratios on the survival of patients with traumatic brain injury (TBI) has not been studied.
Hyperconnectivity and Slow Synapses During Early Development of Medial Prefrontal Cortex in a Mouse Model for Mental Retardation and Autism
Neuronal theories of neurodevelopmental disorders (NDDs) of autism and mental retardation propose that abnormal connectivity underlies deficits in attentional processing. We tested this theory by studying unitary synaptic connections between layer 5 pyramidal neurons within medial prefrontal cortex (mPFC) networks in the Fmr1-KO mouse model for mental retardation and autism. In line with predictions from neurocognitive theory, we found that neighboring pyramidal neurons were hyperconnected during a critical period in early mPFC development. Surprisingly, excitatory synaptic connections between Fmr1-KO pyramidal neurons were significantly slower and failed to recover from short-term depression as quickly as wild type (WT) synapses. By 4-5 weeks of mPFC development, connectivity rates were identical for both KO and WT pyramidal neurons and synapse dynamics changed from depressing to facilitating responses with similar properties in both groups. We propose that the early alteration in connectivity and synaptic recovery are tightly linked: using a network model, we show that slower synapses are essential to counterbalance hyperconnectivity in order to maintain a dynamic range of excitatory activity. However, the slow synaptic time constants induce decreased responsiveness to low-frequency stimulation, which may explain deficits in integration and early information processing in attentional neuronal networks in NDDs.
A Prospective, Randomized Evaluation of a Novel Everolimus-eluting Coronary Stent: the PLATINUM (a Prospective, Randomized, Multicenter Trial to Assess an Everolimus-Eluting Coronary Stent System [PROMUS Element] for the Treatment of Up to Two De Novo Coronary Artery Lesions) Trial
We sought to evaluate the clinical outcomes with a novel platinum chromium everolimus-eluting stent (PtCr-EES) compared with a predicate cobalt chromium everolimus-eluting stent (CoCr-EES) in patients undergoing percutaneous coronary intervention (PCI).
Measurements of Higher Order Flow Harmonics in Au+Au Collisions at Sqrt[s_{NN}]=200 GeV
Flow coefficients v_{n} for n=2, 3, 4, characterizing the anisotropic collective flow in Au+Au collisions at sqrt[s_{NN}]=200 GeV, are measured relative to event planes Ψ_{n}, determined at large rapidity. We report v_{n} as a function of transverse momentum and collision centrality, and study the correlations among the event planes of different order n. The v_{n} are well described by hydrodynamic models which employ a Glauber Monte Carlo initial state geometry with fluctuations, providing additional constraining power on the interplay between initial conditions and the effects of viscosity as the system evolves. This new constraint can serve to improve the precision of the extracted shear viscosity to entropy density ratio η/s.
Cold Nuclear Matter Effects on J/ψ Yields As a Function of Rapidity and Nuclear Geometry in D+A Collisions at Sqrt[s_{NN}]=200 GeV
We present measurements of J/ψ yields in d+Au collisions at sqrt[s_{NN}]=200 GeV recorded by the PHENIX experiment and compare them with yields in p+p collisions at the same energy per nucleon-nucleon collision. The measurements cover a large kinematic range in J/ψ rapidity (-2.2
Suppression of Back-to-Back Hadron Pairs at Forward Rapidity in D+Au Collisions at Sqrt[s_{NN}]=200 GeV
Back-to-back hadron pair yields in d+Au and p+p collisions at sqrt[s_{NN}]=200 GeV were measured with the PHENIX detector at the Relativistic Heavy Ion Collider. Rapidity separated hadron pairs were detected with the trigger hadron at pseudorapidity |η|<0.35 and the associated hadron at forward rapidity (deuteron direction, 3.0<η<3.8). Pairs were also detected with both hadrons measured at forward rapidity; in this case, the yield of back-to-back hadron pairs in d+Au collisions with small impact parameters is observed to be suppressed by a factor of 10 relative to p+p collisions. The kinematics of these pairs is expected to probe partons in the Au nucleus with a low fraction x of the nucleon momenta, where the gluon densities rise sharply. The observed suppression as a function of nuclear thickness, p_{T}, and η points to cold nuclear matter effects arising at high parton densities.
