3rd Annual CDI User Group Meeting
The 3rd Annual CDI User Group Meeting brought together thought leaders, customers, and commercial providers of assays & platforms to discuss ideas, showcase customer research, and share experiences to advance the use of iPS cell technology for drug discovery, disease research, and regenerative medicine.
Dates: Aug 11 – 13, 2014
Location: Sheraton Boston Hotel, 39 Dalton St., Boston, MA
- Disease Research
- Regenerative Medicine
- Ulrich Broeckel, PhD, Medical College of Wisconsin
- Eric Chiao, PhD, Biogen Idec, Inc.
- M. Eileen Dolan, PhD, University of Chicago
- Linda Griffith, PhD, Massachusetts Institute of Technology
- James Hickman, PhD, University of Central Florida
- Roberto Iacone, PhD, Hoffmann-La Roche
- Salman Khetani, PhD, Colorado State University
- Scott Nyberg, MD, Mayo Clinic
- Matt Peters, PhD, AstraZeneca
- Amogh Sivarapatna, PhD, Yale University
- Marija Usenovic, PhD, Merck & Co., Inc.
- Jason Wertheim, MD, PhD, Northwestern University
- Jordy Whisler, PhD, Massachusetts Institute of Technology
Monday, August 11
6:00 pm – 8:00 pm
Tuesday, August 12
7:00 am – 8:00 am
Breakfast, Exhibit Setup & Poster Setup
8:00 am – 8:30 am
Chris Parker, Chief Commercial Officer, CDI
8:30 – 3:10 pm
Session 1: Disease Modeling & Drug Discovery
Chairperson: Eric Chiao, PhD, Biogen Idec, Inc.
8:30 am – 9:00 am
Modeling Neurodegenerative Diseases with Human iPS Cells
Eric Chiao, PhD, Biogen Idec, Inc.
Abstract: The combined economic burden of neurodegenerative diseases such as ALS, Parkinson’s disease and dementia are estimated to be in excess of $200 billion dollars per year in the USA alone. These figures are projected to rise significantly as our population ages. Although much is understood with regards to the clinical progression of these diseases, few therapies are available to treat patients with neurodegenerative diseases. Challenges in discovering efficacious new drugs are due in part to the lack of appropriate platforms for modeling the complexity of the human CNS. Human pluripotent stem cells offer a promising new approach to create informative models of neurodegenerative diseases. This presentation will outline efforts at Biogen to utilize iPSC-derived cells to create models of ALS and Parkinson’s disease that could be used for drug screening, new target identification and validation, and for examining potentially clinically relevant biomarkers. A description of the differentiated cell types we are examining, tools we are employing to assess cellular function and an assessment of challenges and gaps in the field will be provided.
9:05 am – 9:35 am
Model of Tau Pathology in Induced Pluripotent Stem Cell-derived Human Neurons
Marija Usenovic , PhD, Merck & Co., Inc.
Abstract: The pathology of neurodegenerative disorders, like Alzheimer’s disease (AD), may spread throughout the brain via transcellular propagation of misfolded aggregated proteins. The principle of tau transmission in AD has emerged due to spatial and temporal progression of tau pathology in patients’ brains, and evidence showing that tau misfolded proteins propagate from one brain region to another in animal models. However, the molecular and cellular mechanisms of tau transmission remain unknown. We developed a clinically relevant cellular model to study tau aggregation and transmission using iCell® Neurons. We seeded these neurons with full-length human tau monomers and oligomers and examined the long term effects. Tau oligomer-treated neurons exhibited an increase in aggregated and phosphorylated pathological tau. These effects were associated with progressive changes in neuronal morphology. We did not observe any changes after tau monomer treatment. This model of human neurons supports the hypothesis of tau propagation and provides a platform for studying mechanisms involved in tau pathology.
9:40 am – 10:10 am
Poster Teasers (Even Numbers)
10:10 am – 10:55 am
Refreshments, Exhibits & Poster Session
10:55 am – 11:25 am
Growth of iCell Hepatocytes in Three Dimensions
Jason Wertheim, MD, PhD
Abstract: Induced pluripotent stem cells (iPSCs) are a new cell source to model diseases and evaluate the toxicology profile of pharmaceutical agents ex vivo. This study investigates the influence of different 3D microenvironments on iCell Hepatocyte viability, proliferation, and expression of critical proteins that define hepatocyte functionality in comparison to cells grown in standard 2D culture. We developed an extracellular matrix (ECM scaffold) and a second hybrid scaffold comprised of bio-plotted poly-L-lactide acid coated and infused with Type I collagen (PLLA-collagen scaffold). iCell Hepatocytes cultured on these scaffolds were compared to 2D cultures (collagen coated with matrigel overlay). High cell density and good cell viability were found on both scaffolds throughout the 14-day culture period. Both ECM and PLLA-collagen scaffolds enhanced NDUFA3 and albumin expression compared to 2D culture. mRNA and activity of CYP3A4, CYP2C9 and CYP1A2 and protein expression of HMGCR was higher in iCell Hepatocytes grown on ECM scaffolds compared to cells grown on 2D or PLLA-collagen scaffolds. Conclusion: our results demonstrate good cell viability, ingrowth, and proliferation of iCell Hepatocytes in 3D micro-scaffolds with enhanced function on natural ECM cultures.
11:30 am – 12:00 pm
Disease Modeling and Phenotypic Drug Screening for Diabetic Cardiomyopathy Using Human Inducible Pluripotent Stem Cells
Roberto Iacone, PhD, Hoffmann-La Roche, Inc.
Abstract: The development of new medicines for Diabetes Type 2 complications has proven to be challenging due to the use of cellular models which recapitulate some subsets of the specific features of the human disease. Herewith, we develop environmentally and genetically driven in vitro models of the condition using human induced pluripotent stem cell-derived cardiomyocytes. First, we mimic diabetic clinical chemistry to induce a phenotypic surrogate of diabetic cardiomyopathy, observing structural and functional disarray. Next, we consider genetic effects by deriving cardiomyocytes from diabetic patients with variable disease progression. In this work, we present the first patient-specific iPSC model of a complex metabolic condition, showing the power of this technique for discovery and testing of new therapeutic strategies. We conclude presenting a new approach using chemical biology to ultimately elucidate novel mechanisms activating cardiomyocytes repair and regeneration.
12:00 pm – 1:30 pm
Lunch – Roundtable Discussions
Roundtable Discussion Topics
- Commercialization models for cell-based drug screening assays
- iPSCs in pharma & biotech: opportunities & challenges
- Aligning with the CiPA initiative and FDA cardiac safety guidelines
- Organotypic 3D culture / co-culture models
- Tissue engineering using iPSC-derived cells
- iPSCs and disease modeling: strategies, advantages, limitations
- Pharmacogenomic application of iPSC-derived cells
- Considerations for high-throughput screening using iPSC-derived cells
- If I could bank my iPSCs, I would/would not because…
1:30 pm – 2:00 pm
2:05 pm – 2:20 pm
2:25 pm – 2:40 pm
Jacqueline Bowlin, PhD,
University of Colorado School of Medicine
2:45 pm – 3:15 pm
Poster Teasers (Odd Numbers)
3:15 pm – 4:00 pm
Refreshments, Exhibits & Poster Session
4:00 pm – 4:30 pm
Sponsor Talk – Life Technologies
TAL-based Editing of iPSCs for the Study of Neurodegenerative Diseases
Kurt Vogel, PhD
Abstract: Due to the inherent biological variability between cell lines of different backgrounds, the study of subtle phenotypic changes associated with disease states when using iPSC derived materials requires isogenic control lines for proper comparison– that is, cell lines that differ in a defined location but that are otherwise identical at the genetic level. This presentation will describe the approaches we’ve taken to modifying genes involved in Parkinson’s Disease and related disorders. For example, modifying the glucocerebrosidase gene in iPSCs using TAL-based engineering technology presented several challenges, including the presence of an adjacent and highly homologous pseudogene. Initial phenotypic comparisons between the patient derived, TAL-edited lines that we have generated will be presented.
4:35 pm – 5:05 pm
Using Patient Specific iPSC-Cardiomyocytes for Disease Modeling
Uli Broeckel, MD, Medical College of Wisconsin
Abstract: Patient specific iPSC lines offer unprecedented opportunities for disease modeling in particular for cardiovascular diseases. Utilizing this concept, we are currently establishing 250 iPSC cell lines and derive cardiomyocytes from a selected subset of participants in the NHLBI HyperGen study. This study represents a larger cohort of both African American and Caucasian families initially ascertained based on a positive family history of hypertension. Extensive phenotyping includes data on a broad set of risk factors for cardiovascular disease as well as longitudinal follow up data including echocardiography in addition to extensive genetic data such as GWAS and Whole Exome Sequence. Using this infrastructure, we developed assays to study the underlying cellular mechanisms related to the development of Left Ventricular Hypertrophy, a common complication in patients with hypertension, which represents a well established independent risk factor for cardiovascular disease, including heart failure. In this presentation, we will present data demonstrating our model and the use of this model for disease marker identification. In addition, we will discuss the use of this model for the development of novel treatment strategies with a special emphasis on drug development in the context of common complex disease genetics.
5:10 pm – 5:30 pm
Wrap-up – Day 1
6:30 pm – 10:00 pm
Dinner & Social Event
Sponsored by Life Technologies
Wednesday, August 13
7:00 am – 8:00 am
Breakfast, Exhibits & Poster Session
8:00 am – 8:15 am
Chris Parker, Chief Commercial Officer, CDI
8:15 am – 12:00 pm
Session 2: Toxicity
Chairperson: Matt Peters, PhD, AstraZeneca
8:15 am – 8:45 am
Using Stem Cell-derived Cardiomyocytes in Front Line Screens for Cardiotoxicity
Matt Peters, PhD, AstraZeneca
Abstract: Cardiovascular (CV) toxicity is a leading cause of drug attrition and withdrawal. Introducing in vitro assays with higher throughput should permit earlier CV hazard identification and enable medicinal chemists to design-out liabilities. Heretofore, development of in vitro CV assays has been limited by the challenge of replicating integrated cardiovascular physiology while achieving the throughput and consistency required for screening. These challenges appear to be met with a combination of human stem-cell derived cardiomyocytes (CM) which beat spontaneously and monitoring the response with technology that can assess drug induced changes in voltage dependent contraction such as cellular impedance which has been validated with excellent predictivity for drug-induced arrhythmia and contractility. Here, we review advances in cardiomyocyte impedance with emphasis on stem cell-derived cardiomyocyte models for toxicity screening. The combined approach will enhance testing for CV liabilities prior to traditional in vivo models.
8:50 am – 9:20 am
Microscale Engineering Approaches to Improve Liver Maturation of iPSC-derived Human Hepatocytes In Vitro
Salman Khetani, PhD, Colorado State University
Abstract: Primary human hepatocytes (PHHs) are the gold standard for constructing in vitro models of the human liver for drug screening. However, PHHs are a limited resource and their quality for in vitro use can vary considerably across lots. Induced pluripotent stem cell-derived human hepatocyte like cells (iHLCs) could potentially help mitigate this sourcing limitation with PHHs and also provide utility in personalized medicine and disease modeling. However, in conventional monolayers, iHLCs display fetal-like liver functions. We have used semiconductor-driven microfabrication and stromal co-culture approaches to significantly mature iHLC functions and stabilize phenotype for at least 4 weeks. Small molecules and extracellular matrix cues further augment iHLC maturity. Finally, the application of microscale iHLC co-cultures for prediction of drug-induced liver toxicity has been demonstrated with comparison to PHH cultures. In the future, microscale engineered iHLC co-cultures could be used in organs-on-a-chip platforms being designed to assess multi-tissue responses to drugs.
9:25 am – 9:55 am
Sponsor Talk – ACEA
The Applications of xCELLigence Cardio System and Stem Cell-derived Cardiomyocytes for Cardiac Risk Assessment
Yama Abassi, Ph.D.
Cardiac toxicity is a major concern in drug development and it is imperative that clinical candidates are thoroughly tested for adverse effects earlier in the drug discovery process. In this presentation we will discuss the utility of ACEA Biosciences xCELLigence RTCA Cardio System in conjunction with stem cell derived cardiomyocytes for assessment of compound risk in the drug discovery process. The system was validated using stem cell-derived cardiomyocytes and primary cardiomyocytes and by dose-response profiling of pharmacological compounds, including ion channel modulators, chronotropic/ionotropic agents, hERG trafficking inhibitors and drugs withdrawn due to TdP arrhythmia. Our results show this system can sensitively and quantitatively detect modulators of cardiac function, including compounds missed by electrophysiology. Our key finding is that pro-arrhythmic compounds produce signature profiles that reflect arrhythmia and can be used for identification of other pro-arrhythmic compounds. The time series data can be used to identify compounds which induce arrhythmia by complex mechanisms such as hERG trafficking inhibition. Microelectronic monitoring of stem cell derived cardiomyocyte beating provides a high throughput, quantitative and predictive assay system that can be used for assessment of cardiac liability earlier in the drug discovery process.
9:55 am – 10:55 am
Refreshments, Exhibits & Poster Session
10:55 am – 11:25 am
Establishing Stem Cell Technology as a Platform to Evaluate Chemotherapy-induced Peripheral Neuropathy
M. Eileen Dolan, PhD, University of Chicago
Abstract: With over 28 million cancer survivors worldwide, awareness of long-term toxicities and impact on quality of life in cancer survivors has heightened. Chemotherapy-induced peripheral neuropathy (CIPN) is one of the most common and potentially permanent side effects of modern chemotherapy. The mechanisms underlying CIPN have not been precisely determined and few human neuronal models to study CIPN exist.
This talk will highlight:
The use of human iPSC-derived neurons, iCell Neurons, as a model to study the effects of chemotherapy on neuronal health
In vitro assays used to assess chemotherapy-induced morphological changes of the neurons and genes involved in CIPN
The potential use of human iPSC-derived neurons for high throughput drug-induced neurotoxicity screening
11:30 am – 12:00 pm
Body-on-a-Chip Systems Utilizing CDI Cells
James Hickman, PhD,
University of Central Florida
Abstract: Body-on-a-chip or organs-on-a-chip systems that are composed of human cells are needed for drug discovery and toxicology research to replace animals due to new regulatory requirements. Our research focus is on the establishment of functional in vitro systems where we seek to create organs and subsystems to model motor control, myelination, blood brain barrier and cognitive function, as well as cardiac, liver and GI tract subsystems. The idea is to integrate microsystems fabrication technology and surface modifications with protein and cellular components, with the aim of initiating and maintaining self-assembly and growth into biologically, mechanically and electronically interactive functional multi-component systems. The ability to control the surface composition of an in vitro system, as well as controlling other variables, such as growth media and cell preparation, all play important roles in creating a defined system for hybrid device fabrication. Our advances in culturing human stem cell derived neurons, cardiomyocytes and liver cells in a defined serum-free medium, suggest outstanding potential for answering questions during the drug discovery process using a functional body-on-a-chip system composed of multiple organ systems.
12:00 pm – 1:00 pm
Lunch, Exhibits & Poster Session
1:00 pm – 4:15 pm
Session 3: Tissue Engineering & Regenerative Medicine
Chairperson: Linda Griffith, PhD, Massachusetts Institute of Technology
1:00 pm – 1:30 pm
Linda Griffith, PhD
Massachusetts Institute of Technology
1:35 pm – 2:05 pm
Production of Human Hepatocytes in FAH-deficient Pigs for Human Therapies
Scott Nyberg, MD, Mayo Clinic
Abstract: A large demand exists for an abundant, routinely available, high quality source of primary human hepatocytes for therapeutic applications. The overriding hypothesis of our research is that large scale production of human hepatocytes is possible in pigs deficient in the liver enzyme fumarylacetoacetate hydrolase (FAH-KO pig). FAH-deficiency provides a selective advantage to transplanted human hepatocytes over native mutant pig hepatocytes. Rejection of human hepatocytes is prevented if they are transplanted in utero before development of the pig’s immune system. We further hypothesize that the in utero environment will facilitate maturation of liver stem cells, such as induced pluripotent stem cells, into fully functional hepatocytes. The studies to be discussed will serve as a foundation for cellular therapies involving human hepatocytes such as bioartificial liver support of acute liver failure, hepatocyte transplantation for metabolic liver disease, and tissue engineered transplantable livers from human hepatocytes.
2:05 pm – 2:50 pm
Refreshments, Exhibits & Poster Session
2:50 pm – 3:20 pm
Endothelial Differentiation and Arterial Specification
Abstract: Vascular endothelial cells (ECs) are highly plastic in nature and hemodynamic fluid flow is a crucial regulator of arterio-venous fate determination. ECs derived from human iPSCs (hiPSC-ECs) are unique in that they have never been exposed to blood flow. We cultured hiPSC-ECs for 24 hours in a biomimetic shear stress bioreactor to demonstrate activation of Notch signaling, upregulation of several arterial markers including ephrinB2, conexin40 as well as anti-thrombotic and anti-inflammatory markers KLF2 and KLF4. Thus, we are able to modulate and specify the phenotype of hiPSC-ECs to enrich for a vasoprotective, arterial subtype with the de-novo application of shear stress, which is amenable to vascular tissue engineering applications.
3:25 pm – 3:55 pm
Engineered, Perfusable Networks on a Microfluidic Chip
Jordy Whisler, PhD
Massachusetts Institute of Technology
Abstract: The ability to engineer vascularized tissue will eventually enable us to overcome the current organ donor shortage. To achieve this, a reliable process is required for producing functional microvascular networks (MVNs) with specified morphological properties. We developed a multi-culture microfluidic platform to analyze and control the process of vasculogenesis in a three dimensional hydrogel. By manipulating the mechanical and biochemical microenvironment, we were able to control important morphological parameters of the resulting MVNs. To further probe the underlying mechanisms responsible for these observations, we compared our experimental results to an agent based computational model of microvascular network formation and hypothesize that a mechanically driven mechanism is at play. We further developed a method for extracting and sorting the individual cell populations (ECs and fibroblasts) from our microfluidic system in order to perform standard gene sequencing analysis. Specific applications of the platform will be discussed, including: a model for cancer metastasis, endothelial-MSC interactions, and human iPSC generated microvascular networks.
Poster # 101
The Cardiac Microwire Device – Assessment of Contractile Force in 3D Cardiac Microtissues
Abstract: The effective assessment of cardio-active drugs will require 3D microtissues that can recapitulate the in vivo environment. Devices currently exist that can determine both the frequency and rhythm of contraction but to date there are no practical means to measure the force of contraction. Combining CDI’s iCell Cardiomyocytes in co-culture with cardiac fibroblasts, using novel microengineering in a 96 well format, we have developed a medium throughput system for the measurement of the force of contraction. The design criteria identified with our platform should accelerate the development of predictive in vitro assays of human heart tissue function.
Poster # 201
Image-based Evaluation of Contractile Characteristics of hiPS-CMs
Abstract: To clarify the contractile characteristics and the correlation between contractile motion and electrical properties of human induced pluripotent stem cell-derived cardiomyocytes (hiPS-CMs), we performed video microscopy using high-resolution motion vector analysis and MEA measurements for hiPS-CMs (iCell Cardiomyocytes). Simultaneous measurements of contractile motion and field potential, in the presence of various cardioactive agents, revealed that the electrophysiological behaviors of hiPS-CMs are quantitatively reflected in the contractile motion detected by this image-based technique. The results of our present study offer insights into the interpretation of the motion kinetics of the hiPS-CMs, and are relevant for understanding electrical and mechanical relationship in hiPS-CMs.
Poster # 202
A Functional Phenotypic Screen for Synapse Formation in Human iPSC-derived Neurons
Abstract: We used iCell Neurons from CDI to develop a high content screen to assess synaptic foci in response to exposure to compounds. To detect synaptic foci, we used antibodies to the pre-synaptic protein, Synapsin -1, and post-synaptic protein, PSD95. To analyze synapse response to compound exposure, we analyzed images using algorithms to mask co-localized signals for the pre- and post-synaptic markers on neurite regions demarcated by β-III tubulin expression. Our screen detected hits among EPA ToxCAST libraries and known compounds. In addition to effects on synaptic foci, our assay detected effects on neurite outgrowth and cell death.
Poster # 203
The Use of iCell Cardiomyocytes with Cell-by-Cell Calcium Analysis Reveals Compound Induced Changes in Cardiac Repolarization and the Generation of Pro-arrhythmia Signals
Abstract: Cardiomyocytes derived from human stem cells (hSC-CM’s) provide researchers with a model that facilitates direct assessment of compound effects on cardiac excitation regardless of whether the effects are mediated through single ion channel agonist/antagonist action, through simultaneous multichannel effects or through altered signaling. In this study we analyzed 90 compounds using an automated cell-by-cell analysis of calcium transients to detect compound induced changes in repolarization and the generation of proarrhythmia signals. Most compound effects were found to occur at concentrations which match clinical outcomes. Additionally, many of the compounds which are known to induce arrhythmia also demonstrated pro-arrhythmia signals.
Poster # 205
Characterization of Synaptic Transmission Induced Synchronized Population Bursts of the iPSC-derived Neurons
Eisai Co., Ltd.
Abstract: Many drugs have been reported to cause seizures. Major reasons of the drug-induced seizures are from impaired inhibition of GABAA antagonism, GABAB agonism, adenosine antagonism, and also from enhanced excitation by NMDA and other excitatory amino acids. Seizures can also induced disordered conduction by sodium channel blockade and metabolic balance is disturbed in the neurons. Spontaneous neuron activity recordings by multi-electrode array (MEA) system from networks of cultured neurons could be a good risk evaluation system for such drug-inducing seizure events. It was reported that synapse maturation of human induced pluripotent stem cell (hiPSC)-derived neurons were accelerated by co-culture with rat astrocytes. In this study, we observed time course of generation of population burst spikes from iCell Neurons with humoral factors of mouse primary astrocytes by MEA system.
Pharmacological treatments suggest synaptic transmission induced the synchronized population bursts.
Disease Modeling & Drug Discovery
Poster # 301
VZV, a Neurotropic Human Alphaherpesvirus, Causes Varicella, Becomes latent in Ganglionic Neurons and Reactivates to Produce Zoster
University of Colorado, School of Medicine
Abstract: In VZV-infected iCell neurons, VZV capsids lack DNA and minimal infectious virus is produced. No significant differences in the VZV transcriptome were revealed by next-generation RNA-sequencing in infected neurons versus infected fibroblasts; however, compared to infected fibroblasts, VZV DNA did not accumulate in infected iCell neurons over 14 days. Together, the presence of incomplete virus particles in neurons and only minimal production of infectious virus and VZV DNA provide an explanation for the lack of a cytopathic effect in VZV-infected neurons within 2 weeks.
Poster # 302
The Characterization of Human UBE3A-ATS: A Transcript Implicated in Angelman Syndrome
Genzyme, A Sanofi Company
Abstract: Angelman Syndrome (AS) is a neurogenetic imprinting disorder that affects ~1/12,000 individuals. Mutations within the UBE3A gene are responsible for certain cases of AS.
Silencing of UBE3A may be mediated via UBE3A-ATS, a paternal lncRNA antisense transcript. We performed strand-specific RNA sequencing on primary and terminally differentiated iPSC iCell® Neurons to characterize UBE3A-ATS transcripts. De novo transcriptome assembly revealed three main groups of UBE3A-ATS that overlap with the UBE3A locus and could play a role in transcriptional interference. In silico reconstruction were verified by TaqMan qPCR. Our results will help us to evaluate UBE3A-ATS reduction as a therapeutic strategy
Poster # 303
Application of Human iPSC-derived Cardiomyocytes on Multielectrode Array System to Screen Small Molecules in Drug Discovery
Abstract: Recently, the Cardiac Safety Research Consortium proposed the consideration of a new paradigm to assess proarrhythmic risk in the preclinical space, specifically to utilize newer technologies including iPSC-derived cardiomyocytes and in silico predictive modelling. This has the potential to increase newer and safer chemical entities entering market to benefit patients.
In addition, advancements in both biological models and electrophysiological technologies have allowed faster and higher throughput in vitro methodologies to assess beat rate and functional alterations in the most relevant species (human). Using iPSC-derived cardiomyocytes in the multielectrode array model, we have validated responses and reproducibility of reference compounds with known in vivo and clinical cardiac effects. Additionally, this model has proved useful in assessing structure-activity relationships and earlier rank ordering of compounds for small molecules in early drug discovery.
Poster # 304
Differential Pharmacology of Neurite Outgrowth in Human iPSC-derived Neurons Measured with Kinetic Live-cell Imaging
Abstract: Measurements of neurite outgrowth provide insight into neuropathological disorders and neuronal function. Long term changes in neurite outgrowth were quantified using a label-free live cell kinetic imaging platform, the IncuCyte™ Zoom.
High definition phase contrast images of human iPSC-derived neurons (iCell Neurons, Cellular Dynamics International, Inc.) were acquired for up to 12 days and analyzed for total neurite length, branching and cell body cluster number. Excitotoxic agents, kainic acid and L-glutamic acid, differentially disrupted neurite formation. Kainic acid yielded immediate neurite inhibition, while effects of L-glutamic acid were delayed, illustrating the importance of kinetic measurements for cell-based neurological disease models.
Poster # 305
An Evaluation of the Functional Properties of iCell Neurons Using Conventional Patch-clamp Electrophysiology
School of Pharmacy, University of the Pacific
Abstract: Studies in neuropharmacology have relied almost exclusively on animal neurons. Developments in stem cell science have provided new opportunities to generate human neurons from stem cells. Here, we have explored the neurophysiological and neuropharmacological properties of iCell neurons using single cell patch-clamp techniques. Whole cell recordings were obtained from iCell Neurons and cultured for up to 4 months in vitro. We determined the functional expression of sodium, potassium and calcium ion channels and GABA-A, glutamate, NMDA and glycine receptors; we also recorded spontaneous and evoked action potentials. iCell Neurons therefore appear to have important electrophysiological properties consistent with human neurons.
Poster # 306
Electric Field Stimulation (EFS) on iCell® Cardiomyocytes Using Hamamatsu FDSS/μCELL
Abstract: Hamamatsu has developed a 96-channel electrode array system that is mounted on the FDSS/μCELL. It adds electric field stimulations (EFS) to all 96 wells in a microplate simultaneously while fluorescence/luminescence signals are monitored. We measured oscillations of intracellular Ca2+ concentration, which occurs along with the beating of the cells, with a calcium sensitive fluorescent dye in human iPSC-derived cardiomyocytes (iCell® Cardiomyocytes, Cellular Dynamics). We observed that the Ca2+ oscillation was synchronized to the electric stimulation, which indicates that the EFS system is able to pace the beatings of cardiomyocytes. In the presence of some ion channel blockers, EFS was given to see the effects of blockers. Such intracellular Ca2+ kinetics measurements coupled with electric stimulation should be useful in the assessment of cardiac toxicity of pharmacological compounds, in particular in the toxicity screening at the early stages of drug development.
Poster # 307
Improving the Functional Analysis of Human Stem Cell Derived Cardiomyocytes for Use in Drug Discovery
Abstract: Cardiac ischemia is the world leading cause of death, with more than 7 million deaths per year. The process of restoring blood flow to the ischemic tissue can itself cause damage. This is known as a myocardial ischemia reperfusion injury (MIRI).
The aim is to study an in vitro model of MIRI on commercial hiPSC-cardiomyocytes (CDI) by means of engineered fluorescent reporter proteins. These proteins can be targeted specifically to report on diverse cellular properties such as membrane potential, calcium, pH, ATP, etc. Using specially designed microscopes, we can record from many fluorescent reporters simultaneously while maintaining a syncytium.
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Meeting Manager, Cellular Dynamics
Meeting Manager, Cellular Dynamics
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