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https://www.readbyqxmd.com/read/28549865/application-of-crispr-cas9-to-the-study-of-brain-development-and-neuropsychiatric-disease
#1
REVIEW
S K Powell, J Gregory, S Akbarian, K J Brennand
CRISPR/Cas9 technology has transformed our abilities to manipulate the genome and epigenome, as applications have expanded from efficient genomic editing to include the targeted localization of effectors to specific genomic loci. By facilitating the manipulation of DNA- and histone-modifying enzyme activities, activation or repression of gene expression, and targeting of transcriptional regulators to defined loci, it is now possible to directly probe the role of gene-regulatory and epigenetic pathways in order to examine their roles in basic biology and disease processes...
May 23, 2017: Molecular and Cellular Neurosciences
https://www.readbyqxmd.com/read/28549786/action-potential-based-mea-platform-for-in-vitro-screening-of-drug-induced-cardiotoxicity-using-human-ipscs-and-rat-neonatal-myocytes
#2
Danny Jans, Geert Callewaert, Olga Krylychkina, Luis Hoffman, Francesco Gullo, Dimiter Prodanov, Dries Braeken
Drug-induced cardiotoxicity poses a negative impact on public health and drug development. Cardiac safety pharmacology issues urged for the preclinical assessment of drug-induced ventricular arrhythmia leading to the design of several in vitro electrophysiological screening assays. In general, patch clamp systems allow for intracellular recordings, while multi-electrode array (MEA) technology detect extracellular activity. Here, we demonstrate a complementary metal oxide semiconductor (CMOS)-based MEA system as a reliable platform for non-invasive, long-term intracellular recording of cardiac action potentials at high resolution...
May 23, 2017: Journal of Pharmacological and Toxicological Methods
https://www.readbyqxmd.com/read/28546047/an-impedance-based-approach-using-human-ipsc-derived-cardiomyocytes-significantly-improves-in-vitro-prediction-of-in-vivo-cardiotox-liabilities
#3
Bryan Koci, Gregory Luerman, Anika Duenbostell, Ralf Kettenhofen, Heribert Bohlen, Luke Coyle, Brian Knight, Warren Ku, Walter Volberg, Joseph R Woska, Martha P Brown
Current in vitro approaches to cardiac safety testing typically focus on mechanistic ion channel testing to predict in vivo proarrhythmic potential. Outside of the Comprehensive in vitro Proarrhythmia Assay (CiPA) initiative, structural and functional cardiotoxicity related to chronic dosing effects are of great concern as these effects can impact compound attrition. Development and implementation of an in vitro cardiotoxicity screening platform that effectively identifies these liabilities early in the discovery process should reduce costly attrition and decrease preclinical development time...
May 22, 2017: Toxicology and Applied Pharmacology
https://www.readbyqxmd.com/read/28539859/human-induced-pluripotent-stem-cells-generated-neural-cells-behaving-like-brain-and-spinal-cord-cells-an-insight-into-the-involvement-of-retinoic-acid-and-sonic-hedgehog-proteins
#4
Akinlolu Abdulazeez Adelaja
OBJECTIVES: The previous studies generated neural progenitor cells (NPCs) from human induced pluripotent stem cells (hiPSCs) using different protocols. However, the nature of the temporal or regional specificity of NPCs derived using these protocols is not well defined. Therefore, this study aimed to generate age- and region-specific NPCs from hiPSCs, which mimic in vivo fetal brain (FNPC-B) or spinal cord (FNPC-SC) tissues, in the absence or presence of retinoic acid (RA) and sonic hedgehog (SHH)...
April 2017: International Journal of Health Sciences
https://www.readbyqxmd.com/read/28537488/capturing-human-na%C3%A3-ve-pluripotency-in-the-embryo-and-in-the-dish
#5
Ludovic Zimmerlin, Tea Soon Park, Elias T Zambidis
Although human embryonic stem cells (hESC) were first derived almost 20 years ago, it was only recently acknowledged that they share closer molecular and functional identity to post-implantation lineage-primed murine epiblast stem cells (EpiSC) than to naïve pre-implantation ICM-derived mouse embryonic stem cells (ESC). A myriad of transcriptional, epigenetic, biochemical, and metabolic attributes have now been described that distinguish naïve and primed pluripotent states in both rodents and humans. Conventional hESC and human induced pluripotent stem cells (hiPSC) appear to lack many of the defining hallmarks of naïve mouse ESC...
May 24, 2017: Stem Cells and Development
https://www.readbyqxmd.com/read/28534195/identification-of-a-small-molecule-that-facilitates-the-differentiation-of-human-ipscs-escs-and-mouse-embryonic-pancreatic-explants-into-pancreatic-endocrine-cells
#6
Yasushi Kondo, Taro Toyoda, Ryo Ito, Michinori Funato, Yoshiya Hosokawa, Satoshi Matsui, Tomomi Sudo, Masahiro Nakamura, Chihiro Okada, Xiaotong Zhuang, Akira Watanabe, Akira Ohta, Nobuya Inagaki, Kenji Osafune
AIMS/HYPOTHESIS: Pancreatic beta-like cells generated from human induced pluripotent stem cells (hiPSCs) or human embryonic stem cells (hESCs) offer an appealing donor tissue source. However, differentiation protocols that mainly use growth factors are costly. Therefore, in this study, we aimed to establish efficient differentiation protocols to change hiPSCs/hESCs to insulin (INS)(+) cells using novel small-molecule inducers. METHODS: We screened small molecules that increased the induction rate of INS(+) cells from hESC-derived pancreatic and duodenal homeobox 1 (PDX1)(+) pancreatic progenitor cells...
May 22, 2017: Diabetologia
https://www.readbyqxmd.com/read/28526246/impaired-mitophagy-facilitates-mitochondrial-damage-in-danon-disease
#7
Sherin I Hashem, Anne N Murphy, Ajit S Divakaruni, Matthew L Klos, Bradley C Nelson, Emily C Gault, Teisha J Rowland, Cynthia N Perry, Yusu Gu, Nancy D Dalton, William H Bradford, Eric J Devaney, Kirk L Peterson, Kenneth L Jones, Matthew R G Taylor, Ju Chen, Neil C Chi, Eric D Adler
RATIONALE: Lysosomal associated membrane protein type-2 (LAMP-2) is a highly conserved, ubiquitous protein that is critical for autophagic flux. Loss of function mutations in the LAMP-2 gene cause Danon disease, a rare X-linked disorder characterized by developmental delay, skeletal muscle weakness, and severe cardiomyopathy. We previously found that human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) from Danon patients exhibited significant mitochondrial oxidative stress and apoptosis...
May 16, 2017: Journal of Molecular and Cellular Cardiology
https://www.readbyqxmd.com/read/28525643/synthetic-mrna-devices-that-detect-endogenous-proteins-and-distinguish-mammalian-cells
#8
Shunsuke Kawasaki, Yoshihiko Fujita, Takashi Nagaike, Kozo Tomita, Hirohide Saito
Synthetic biology has great potential for future therapeutic applications including autonomous cell programming through the detection of protein signals and the production of desired outputs. Synthetic RNA devices are promising for this purpose. However, the number of available devices is limited due to the difficulty in the detection of endogenous proteins within a cell. Here, we show a strategy to construct synthetic mRNA devices that detect endogenous proteins in living cells, control translation and distinguish cell types...
May 19, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/28522326/gene-editing-and-clonal-isolation-of-human-induced-pluripotent-stem-cells-using-crispr-cas9
#9
Saniye Yumlu, Jürgen Stumm, Sanum Bashir, Anne-Kathrin Dreyer, Pawel Lisowski, Eric Danner, Ralf Kühn
Human induced pluripotent stem cells (hiPSCs) represent an ideal in vitro platform to study human genetics and biology. The recent advent of programmable nucleases makes also the human genome amenable to experimental genetics through either the correction of mutations in patient-derived iPSC lines or the de novo introduction of mutations into otherwise healthy iPSCs. The production of specific and sometimes complex genotypes in multiple cell lines requires efficient and streamlined gene editing technologies...
May 15, 2017: Methods: a Companion to Methods in Enzymology
https://www.readbyqxmd.com/read/28509371/quantitative-assessment-of-sialo-glycoproteins-and-n-glycans-during-cardiomyogenic-differentiation-of-human-induced-pluripotent-stem-cells
#10
Sarah Anna Konze, Samanta Cajic, Astrid Oberbeck, René Hennig, Andreas Pich, Erdmann Rapp, Falk Buettner
Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) may be used in regenerative medicine for individualized tissue transplants in future. For application in patients, the generated CMs have to be highly pure and well characterized. To overcome the prevalent scarcity of CM-specific markers, we quantitatively assessed cell surface exposed sialo-glycoproteins and N-glycans of hiPSCs, CM progenitors and CMs derived thereof. Applying a combination of metabolic labeling and specific sialo-glycoprotein capture, we could highly enrich and quantify membrane proteins during cardiomyogenic differentiation...
May 16, 2017: Chembiochem: a European Journal of Chemical Biology
https://www.readbyqxmd.com/read/28507211/autologous-pluripotent-stem-cell-derived-%C3%AE-like-cells-for-diabetes-cellular-therapy
#11
Jeffrey R Millman, Felicia W Pagliuca
Development of stem cell technologies for cell replacement therapy has progressed rapidly in recent years. Diabetes has long been seen as one of the first applications for stem cell-derived cells because of the loss of only a single cell type-the insulin-producing β-cell. Recent reports have detailed strategies that overcome prior hurdles to generate functional β-like cells from human pluripotent stem cells in vitro, including from human induced pluripotent stem cells (hiPSCs). Even with this accomplishment, addressing immunological barriers to transplantation remains a major challenge for the field...
May 2017: Diabetes
https://www.readbyqxmd.com/read/28506844/proarrhythmic-risk-assessment-using-conventional-and-new-in%C3%A2-vitro-assays
#12
Sonia Goineau, Vincent Castagné
Drug-induced QT prolongation is a major safety issue in the drug discovery process. This study was conducted to assess the electrophysiological responses of four substances using established preclinical assays usually used in regulatory studies (hERG channel or Purkinje fiber action potential) and a new assay (human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs)-field potential). After acute exposure, moxifloxacin and dofetilide concentration-dependently decreased IKr amplitude (IC50 values: 102 μM and 40 nM, respectively) and lengthened action potential (100 μM moxifloxacin: +23% and 10 nM dofetilide: +18%) and field potential (300 μM moxifloxacin: +76% and 10 nM dofetilide: +38%) durations...
May 12, 2017: Regulatory Toxicology and Pharmacology: RTP
https://www.readbyqxmd.com/read/28500272/probing-the-lithium-response-pathway-in-hipscs-implicates-the-phosphoregulatory-set-point-for-a-cytoskeletal-modulator-in-bipolar-pathogenesis
#13
Brian T D Tobe, Andrew M Crain, Alicia M Winquist, Barbara Calabrese, Hiroko Makihara, Wen-Ning Zhao, Jasmin Lalonde, Haruko Nakamura, Glenn Konopaske, Michelle Sidor, Cameron D Pernia, Naoya Yamashita, Moyuka Wada, Yuuka Inoue, Fumio Nakamura, Steven D Sheridan, Ryan W Logan, Michael Brandel, Dongmei Wu, Joshua Hunsberger, Laurel Dorsett, Cordulla Duerr, Ranor C B Basa, Michael J McCarthy, Namrata D Udeshi, Philipp Mertins, Steven A Carr, Guy A Rouleau, Lina Mastrangelo, Jianxue Li, Gustavo J Gutierrez, Laurence M Brill, Nikolaos Venizelos, Guang Chen, Jeffrey S Nye, Husseini Manji, Jeffrey H Price, Colleen A McClung, Hagop S Akiskal, Martin Alda, De-Maw M Chuang, Joseph T Coyle, Yang Liu, Yang D Teng, Toshio Ohshima, Katsuhiko Mikoshiba, Richard L Sidman, Shelley Halpain, Stephen J Haggarty, Yoshio Goshima, Evan Y Snyder
The molecular pathogenesis of bipolar disorder (BPD) is poorly understood. Using human-induced pluripotent stem cells (hiPSCs) to unravel such mechanisms in polygenic diseases is generally challenging. However, hiPSCs from BPD patients responsive to lithium offered unique opportunities to discern lithium's target and hence gain molecular insight into BPD. By profiling the proteomics of BDP-hiPSC-derived neurons, we found that lithium alters the phosphorylation state of collapsin response mediator protein-2 (CRMP2)...
May 12, 2017: Proceedings of the National Academy of Sciences of the United States of America
https://www.readbyqxmd.com/read/28494940/differentiation-defective-human-induced-pluripotent-stem-cells-reveal%C3%A2-strengths-and-limitations-of-the-teratoma-assay-and-in%C3%A2-vitro-pluripotency-assays
#14
Marga J Bouma, Maarten van Iterson, Bart Janssen, Christine L Mummery, Daniela C F Salvatori, Christian Freund
The ability to form teratomas in vivo containing multiple somatic cell types is regarded as functional evidence of pluripotency for human pluripotent stem cells (hPSCs). Since the Teratoma assay is animal dependent, laborious, and only qualitative, the PluriTest and the hPSC ScoreCard assay have been developed as in vitro alternatives. Here we compared normal hPSCs, induced hPSCs (hiPSCs) with reactivated reprogramming transgenes, and human embryonal carcinoma cells (hECs) in these assays. While normal hPSCs gave rise to typical teratomas, the xenografts of the hECs and the hiPSCs with reactivated reprogramming transgenes were largely undifferentiated and malignant...
May 9, 2017: Stem Cell Reports
https://www.readbyqxmd.com/read/28492526/differentiation-of-cardiomyocytes-and-generation-of-human-engineered-heart-tissue
#15
Kaja Breckwoldt, David Letuffe-Brenière, Ingra Mannhardt, Thomas Schulze, Bärbel Ulmer, Tessa Werner, Anika Benzin, Birgit Klampe, Marina C Reinsch, Sandra Laufer, Aya Shibamiya, Maksymilian Prondzynski, Giulia Mearini, Dennis Schade, Sigrid Fuchs, Christiane Neuber, Elisabeth Krämer, Umber Saleem, Mirja L Schulze, Marita L Rodriguez, Thomas Eschenhagen, Arne Hansen
Since the advent of the generation of human induced pluripotent stem cells (hiPSCs), numerous protocols have been developed to differentiate hiPSCs into cardiomyocytes and then subsequently assess their ability to recapitulate the properties of adult human cardiomyocytes. However, hiPSC-derived cardiomyocytes (hiPSC-CMs) are often assessed in single-cell assays. A shortcoming of these assays is the limited ability to characterize the physiological parameters of cardiomyocytes, such as contractile force, due to random orientations...
June 2017: Nature Protocols
https://www.readbyqxmd.com/read/28491099/induction-of-pluripotent-stem-cells-from-a-manifesting-carrier-of-duchenne-muscular-dystrophy-and-characterization-of-their-x-inactivation-status
#16
Yuko Miyagoe-Suzuki, Takashi Nishiyama, Miho Nakamura, Asako Narita, Fusako Takemura, Satoru Masuda, Narihiro Minami, Kumiko Murayama, Hirofumi Komaki, Yu-Ichi Goto, Shin'ichi Takeda
Three to eight percent of female carriers of Duchenne muscular dystrophy (DMD) develop dystrophic symptoms ranging from mild muscle weakness to a rapidly progressive DMD-like muscular dystrophy due to skewed inactivation of X chromosomes during early development. Here, we generated human induced pluripotent stem cells (hiPSCs) from a manifesting female carrier using retroviral or Sendai viral (SeV) vectors and determined their X-inactivation status. Although manifesting carrier-derived iPS cells showed normal expression of human embryonic stem cell markers and formed well-differentiated teratomas in vivo, many hiPS clones showed bi-allelic expression of the androgen receptor (AR) gene and loss of X-inactivation-specific transcript and trimethyl-histone H3 (Lys27) signals on X chromosomes, suggesting that both X chromosomes of the hiPS cells are in an active state...
2017: Stem Cells International
https://www.readbyqxmd.com/read/28487523/a-preliminary-analysis-of-volatile-metabolites-of-human-induced-pluripotent-stem-cells-along-the-in-vitro-differentiation
#17
Rosamaria Capuano, Paola Spitalieri, Rosa Valentina Talarico, Ana Carolina Domakoski, Alexandro Catini, Roberto Paolesse, Eugenio Martinelli, Giuseppe Novelli, Federica Sangiuolo, Corrado Di Natale
Cellular metabolism of stem cell biology is still an unexplored field. However, considering the amount of information carried by metabolomes, this is a promising field for a fast identification of stem cells itself and during the differentiation process. One of the goals of such application is the identification of residual pluripotent cells before cell transplantation to avoid the occurrence of teratomas. In this paper, we investigated in vitro the volatile compounds (VOCs) released during human induced pluripotent stem cells (hiPSCs) reprogramming...
May 9, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28476540/epigenetic-and-transcriptional-modulation-of-wdr5-a-chromatin-remodeling-protein-in-huntington-s-disease-human-induced-pluripotent-stem-cell-hipsc-model
#18
Simona Baronchelli, Alberto La Spada, Aikaterini Ntai, Andrea Barbieri, Paola Conforti, Gloria Saccani Jotti, Serena Redaelli, Angela Bentivegna, Pasquale De Blasio, Ida Biunno
DNA methylation (DNAm) changes are of increasing relevance to neurodegenerative disorders, including Huntington's disease (HD). We performed genome-wide screening of possible DNAm changes occurring during striatal differentiation in human induced pluripotent stem cells derived from a HD patient (HD-hiPSCs) as cellular model. We identified 240 differentially methylated regions (DMRs) at promoters in fully differentiated HD-hiPSCs. Subsequently, we focused on the methylation differences in a subcluster of genes related to Jumonji Domain Containing 3 (JMJD3), a demethylase that epigenetically regulates neuronal differentiation and activates neuronal progenitor associated genes, which are indispensable for neuronal fate acquisition...
May 2, 2017: Molecular and Cellular Neurosciences
https://www.readbyqxmd.com/read/28474496/an-arduous-journey-from-human-pluripotent-stem-cells-to-functional-pancreatic-%C3%AE-cells
#19
REVIEW
Larry Sai Weng Loo, Hwee Hui Lau, Joanita Binte Jasmen, Chang Siang Lim, Adrian Kee Keong Teo
Type 1 and type 2 diabetes are caused by either a destruction or decrease in the number of functional insulin-producing β-cells, respectively. Therefore, the generation of functional β-cells from human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs), collectively known as human pluripotent stem cells (hPSCs), for potential cell replacement therapy and disease modeling is an intensely investigated area. Recent scientific breakthroughs enabled derivation of large quantities of human pancreatic β-like cells in vitro although with varied glucose-stimulated insulin secretion kinetics...
May 4, 2017: Diabetes, Obesity & Metabolism
https://www.readbyqxmd.com/read/28465161/assessment-of-elisa-as-endpoint-in-neuronal-cell-based-assay-for-bont-detection-using-hipsc-derived-neurons
#20
Sabine Pellett, William H Tepp, Eric A Johnson, Dorothea Sesardic
INTRODUCTION: Botulinum neurotoxins (BoNTs), the causative agents of botulism, are widely used as powerful bio-pharmaceuticals to treat neuro-muscular disorders. Due to the high potency and potential lethality of BoNTs, careful monitoring of the biologic activity of BoNT-based pharmaceuticals is required to ensure safe usage. For decades, the only approved method for potency determination of pharmaceutical BoNTs was the mouse bioassay (MBA), but in recent years improvements in cell-assay technologies have enabled MBA replacement by cell-based assays for specific product evaluations...
April 29, 2017: Journal of Pharmacological and Toxicological Methods
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