Publications (5) View all
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Article: Genetic correction of Huntington's disease phenotypes in induced pluripotent stem cells.
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ABSTRACT: Huntington's disease (HD) is caused by a CAG expansion in the huntingtin gene. Expansion of the polyglutamine tract in the huntingtin protein results in massive cell death in the striatum of HD patients. We report that human induced pluripotent stem cells (iPSCs) derived from HD patient fibroblasts can be corrected by the replacement of the expanded CAG repeat with a normal repeat using homologous recombination, and that the correction persists in iPSC differentiation into DARPP-32-positive neurons in vitro and in vivo. Further, correction of the HD-iPSCs normalized pathogenic HD signaling pathways (cadherin, TGF-β, BDNF, and caspase activation) and reversed disease phenotypes such as susceptibility to cell death and altered mitochondrial bioenergetics in neural stem cells. The ability to make patient-specific, genetically corrected iPSCs from HD patients will provide relevant disease models in identical genetic backgrounds and is a critical step for the eventual use of these cells in cell replacement therapy.Cell stem cell 06/2012; 11(2):253-63. · 23.56 Impact Factor -
Article: Acetylcholine negatively regulates development of the neuromuscular junction through distinct cellular mechanisms.
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ABSTRACT: Emerging evidence suggests that the neurotransmitter acetylcholine (ACh) negatively regulates the development of the neuromuscular junction, but it is not clear if ACh exerts its effects exclusively through muscle ACh receptors (AChRs). Here, we used genetic methods to remove AChRs selectively from muscle. Similar to the effects of blocking ACh biosynthesis, eliminating postsynaptic AChRs increased motor axon branching and expanded innervation territory, suggesting that ACh negatively regulates synaptic growth through postsynaptic AChRs. However, in contrast to the effects of blocking ACh biosynthesis, eliminating postsynaptic AChRs in agrin-deficient mice failed to restore deficits in pre- and postsynaptic differentiation, suggesting that ACh negatively regulates synaptic differentiation through nonpostsynaptic receptors. Consistent with this idea, the ACh agonist carbachol inhibited presynaptic specialization of motorneurons in vitro. Together, these data suggest that ACh negatively regulates axon growth and presynaptic specialization at the neuromuscular junction through distinct cellular mechanisms.Proceedings of the National Academy of Sciences 06/2010; 107(23):10702-7. · 9.68 Impact Factor -
Article: Characterization of Human Huntington's Disease Cell Model from Induced Pluripotent Stem Cells.
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ABSTRACT: Huntington's disease (HD) is a dominantly inherited neurodegenerative disease caused by a CAG repeat expansion in the first exon of the gene Huntingtin (Htt). A dramatic pathological change in HD is the massive loss of striatal neurons as the disease progresses. A useful advance in HD would be the generation of a human-derived HD model to use for drug screening and understanding mechanisms of HD. We utilized the recently established human iPS cell line derived from HD patient fibroblasts to derive neuronal precursors and human striatal neurons. To achieve this goal, the differentiation of the HD-iPS cells into striatal fate required several steps. First, we generated nestin+/PAX6+/SOX1+/OCT4- neural stem cells (NSCs) from HD-iPS cells using the method of embryoid body formation. HD-NSCs were then subjected to a differentiation condition combining morphogens and neurotrophins to induce striatal lineage commitment. Striatal neuronal precursors/immature neurons stained with β-III tubulin, calbindin and GABA but not DARPP-32 (dopamine- and cyclic AMP-regulated phosphoprotein, Mr = 32,000) were produced in this step. Finally, maturation and terminal differentiation of the striatal neuronal precursors/immature neurons resulted in striatal neurons expressing markers like DARPP-32. The HD-iPS cells derived striatal neurons and neuronal precursors contain the same CAG expansion as the mutation in the HD patient from whom the iPS cell line was established. Moreover, the HD-NSCs showed enhanced caspase activity upon growth factor deprivation compared to normal NSCs (from iPS or H9 NSCs). Therefore, these differentiated cells may produce a human HD cell model useful in the study of HD mechanisms and drug screening.PLoS currents. 01/2010; 2:RRN1193. -
Article: Global impact of influenza virus on cellular pathways is mediated by both replication-dependent and -independent events.
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ABSTRACT: Influenza virus, the causative agent of the common flu, is a worldwide health problem with significant economic consequences. Studies of influenza virus biology have revealed elaborate mechanisms by which the virus interacts with its host cell as it inhibits the synthesis of cellular proteins, evades the innate antiviral response, and facilitates production of viral RNAs and proteins. With the advent of DNA array technology it is now possible to obtain a large-scale view of how viruses alter the environment within the host cell. In this study, the cellular response to influenza virus infection was examined by monitoring the steady-state mRNA levels for over 4,600 cellular genes. Infections with active and inactivated influenza viruses identified changes in cellular gene expression that were dependent on or independent of viral replication, respectively. Viral replication resulted in the downregulation of many cellular mRNAs, and the effect was enhanced with time postinfection. Interestingly, several genes involved in protein synthesis, transcriptional regulation, and cytokine signaling were induced by influenza virus replication, suggesting that some may play essential or accessory roles in the viral life cycle or the host cell's stress response. The gene expression pattern induced by inactivated viruses revealed induction of the cellular metallothionein genes that may represent a protective response to virus-induced oxidative stress. Genome-scale analyses of virus infections will help us to understand the complexities of virus-host interactions and may lead to the discovery of novel drug targets or antiviral therapies.Journal of Virology 06/2001; 75(9):4321-31. · 5.40 Impact Factor -
Article: Large-Scale Monitoring of Host Cell Gene Expression during HIV-1 Infection Using cDNA Microarrays
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ABSTRACT: Human immunodeficiency virus type 1 (HIV-1) infection alters the expression of host cell genes at both the mRNA and protein levels. To obtain a more comprehensive view of the global effects of HIV infection of CD4-positive T-cells at the mRNA level, we performed cDNA microarray analysis on approximately 1500 cellular cDNAs at 2 and 3 days postinfection (p.i.) with HIV-1. Host cell gene expression changed little at 2 days p.i., but at 3 days p.i. 20 cellular genes were identified as differentially expressed. Genes involved in T-cell signaling, subcellular trafficking, and transcriptional regulation, as well as several uncharacterized genes, were among those whose mRNAs were differentially regulated. These results support the hypothesis that HIV-1 infection alters expression of a broad array of cellular genes and provides a framework for future functional studies on the differentially expressed mRNA products.Virology 01/2000; · 3.35 Impact Factor
