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文献

Robinson M et al. (AUG 2016) Stem Cell Reviews and Reports 12 4 476--483

Functionalizing Ascl1 with Novel Intracellular Protein Delivery Technology for Promoting Neuronal Differentiation of Human Induced Pluripotent Stem Cells

Pluripotent stem cells can become any cell type found in the body. Accordingly,one of the major challenges when working with pluripotent stem cells is producing a highly homogenous population of differentiated cells,which can then be used for downstream applications such as cell therapies or drug screening. The transcription factor Ascl1 plays a key role in neural development and previous work has shown that Ascl1 overexpression using viral vectors can reprogram fibroblasts directly into neurons. Here we report on how a recombinant version of the Ascl1 protein functionalized with intracellular protein delivery technology (Ascl1-IPTD) can be used to rapidly differentiate human induced pluripotent stem cells (hiPSCs) into neurons. We first evaluated a range of Ascl1-IPTD concentrations to determine the most effective amount for generating neurons from hiPSCs cultured in serum free media. Next,we looked at the frequency of Ascl1-IPTD supplementation in the media on differentiation and found that one time supplementation is sufficient enough to trigger the neural differentiation process. Ascl1-IPTD was efficiently taken up by the hiPSCs and enabled rapid differentiation into TUJ1-positive and NeuN-positive populations with neuronal morphology after 8 days. After 12 days of culture,hiPSC-derived neurons produced by Ascl1-IPTD treatment exhibited greater neurite length and higher numbers of branch points compared to neurons derived using a standard neural progenitor differentiation protocol. This work validates Ascl1-IPTD as a powerful tool for engineering neural tissue from pluripotent stem cells. View Publication

产品号#：

05832

05833

05838

05872

05873

05940

07174

07180

07183

07190

27147

07191

36254

07930

07931

07940

07955

07956

07959

07954

27845

27945

27840

27865

27940

27965

05835

05839

08581

08582

产品名：

STEMdiff™ 神经花环选择试剂

STEMdiff™神经前体细胞培养基

STEMdiff™神经祖细胞冻存液

玻连蛋白 XF™

CellAdhere™ 稀释缓冲液

DMEM/F-12 with 15 mM HEPES

CryoStor® CS10

STEMdiff™ 神经诱导培养基

STEMdiff™SMADi神经诱导试剂盒

STEMdiff™SMADi神经诱导试剂盒，2套

文献

Zhou J et al. (AUG 2016) Neurochemical Research 41 8 2065--2074

Generation of Human Embryonic Stem Cell Line Expressing zsGreen in Cholinergic Neurons Using CRISPR/Cas9 System

Lineage specific human embryonic stem cell (hESC) reporter cell line is a versatile tool for biological studies on real time monitoring of differentiation,physiological and biochemical features of special cell types and pathological mechanism of disease. Here we report the generation of ChAT-zsGreen reporter hESC line that express zsGreen under the control of the choline acetyltransferase (ChAT) promoter using CRISPR (Clustered Regularly Interspersed Short Palindromic Repeats)/Cas9 system. We show that the ChAT-zsGreen hESC reporter cell lines retain the features of undifferentiated hESC. After cholinergic neuronal differentiation,cholinergic neurons were clearly labeled with green fluorescence protein (zsGreen). The ChAT-zsGreen reporter hESC lines are invaluable not only for the monitoring cholinergic neuronal differentiation but also for study physiological and biochemical hallmarks of cholinergic neurons. View Publication

文献

Dafinca R et al. (APR 2016) Stem cells (Dayton,Ohio) 34 8 2016

C9orf72 Hexanucleotide Expansions are Associated with Altered ER Calcium Homeostasis and Stress Granule Formation in iPSC-Derived Neurons from Patients with Amyotrophic Lateral Sclerosis and Frontotemporal Dementia.

An expanded hexanucleotide repeat in a noncoding region of the C9orf72 gene is a major cause of amyotrophic lateral sclerosis (ALS),accounting for up to 40% of familial cases and 7% of sporadic ALS in European populations. We have generated induced pluripotent stem cells (iPSCs) from fibroblasts of patients carrying C9orf72 hexanucleotide expansions,differentiated these to functional motor and cortical neurons and performed an extensive phenotypic characterization. In C9orf72 iPSC-derived motor neurons,decreased cell survival is correlated with dysfunction in Ca(2+) homeostasis,reduced levels of the anti-apoptotic protein Bcl-2,increased endoplasmic reticulum (ER) stress and reduced mitochondrial membrane potential. Furthermore,C9orf72 motor neurons,and also cortical neurons,show evidence of abnormal protein aggregation and stress granule formation. This study is an extensive characterization of iPSC-derived motor neurons as cellular models of ALS carrying C9orf72 hexanucleotide repeats,which describes a novel pathogenic link between C9orf72 mutations,dysregulation of calcium signalling and altered proteostasis and provides a potential pharmacological target for the treatment of ALS and the related neurodegenerative disease frontotemporal dementia (FTD). This article is protected by copyright. All rights reserved. View Publication

产品号#：

85850

85857

产品名：

mTeSR™1

文献

Nekrasov ED et al. (DEC 2016) Molecular Neurodegeneration 11 1 1--15

Manifestation of Huntington's disease pathology in human induced pluripotent stem cell-derived neurons.

Background: Huntington's disease (HD) is an incurable hereditary neurodegenerative disorder,which manifests itself as a loss of GABAergic medium spiny (GABA MS) neurons in the striatum and caused by an expansion of the CAG repeat in exon 1 of the huntingtin gene. There is no cure for HD,existing pharmaceutical can only relieve its symptoms. Results: Here,induced pluripotent stem cells were established from patients with low CAG repeat expansion in the huntingtin gene,and were then efficiently differentiated into GABA MS-like neurons (GMSLNs) under defined culture conditions. The generated HD GMSLNs recapitulated disease pathology in vitro,as evidenced by mutant huntingtin protein aggregation,increased number of lysosomes/autophagosomes,nuclear indentations,and enhanced neuronal death during cell aging. Moreover,store-operated channel (SOC) currents were detected in the differentiated neurons,and enhanced calcium entry was reproducibly demonstrated in all HD GMSLNs genotypes. Additionally,the quinazoline derivative,EVP4593,reduced the number of lysosomes/autophagosomes and SOC currents in HD GMSLNs and exerted neuroprotective effects during cell aging. Conclusions: Our data is the first to demonstrate the direct link of nuclear morphology and SOC calcium deregulation to mutant huntingtin protein expression in iPSCs-derived neurons with disease-mimetic hallmarks,providing a valuable tool for identification of candidate anti-HD drugs. Our experiments demonstrated that EVP4593 may be a promising anti-HD drug. [ABSTRACT FROM AUTHOR] View Publication

产品号#：

05854

05855

85850

85857

产品名：

mFreSR™

mTeSR™1

文献

Francis KR et al. (APR 2016) Nature medicine 22 4 388--396

Modeling Smith-Lemli-Opitz syndrome with induced pluripotent stem cells reveals a causal role for Wnt/&dollar;&dollar;-catenin defects in neuronal cholesterol synthesis phenotypes.

Smith-Lemli-Opitz syndrome (SLOS) is a malformation disorder caused by mutations in DHCR7,which impair the reduction of 7-dehydrocholesterol (7DHC) to cholesterol. SLOS results in cognitive impairment,behavioral abnormalities and nervous system defects,though neither affected cell types nor impaired signaling pathways are fully understood. Whether 7DHC accumulation or cholesterol loss is primarily responsible for disease pathogenesis is also unclear. Using induced pluripotent stem cells (iPSCs) from subjects with SLOS,we identified cellular defects that lead to precocious neuronal specification within SLOS derived neural progenitors. We also demonstrated that 7DHC accumulation,not cholesterol deficiency,is critical for SLOS-associated defects. We further identified downregulation of Wnt/$$-catenin signaling as a key initiator of aberrant SLOS iPSC differentiation through the direct inhibitory effects of 7DHC on the formation of an active Wnt receptor complex. Activation of canonical Wnt signaling prevented the neural phenotypes observed in SLOS iPSCs,suggesting that Wnt signaling may be a promising therapeutic target for SLOS. View Publication

产品号#：

07923

85850

85857

产品名：

Dispase (1 U/mL)

mTeSR™1

文献

Chamma I et al. (MAR 2016) Nature Communications 7 10773

Mapping the dynamics and nanoscale organization of synaptic adhesion proteins using monomeric streptavidin

The advent of super-resolution imaging (SRI) has created a need for optimized labelling strategies. We present a new method relying on fluorophore-conjugated monomeric streptavidin (mSA) to label membrane proteins carrying a short,enzymatically biotinylated tag,compatible with SRI techniques including uPAINT,STED and dSTORM. We demonstrate efficient and specific labelling of target proteins in confined intercellular and organotypic tissues,with reduced steric hindrance and no crosslinking compared with multivalent probes. We use mSA to decipher the dynamics and nanoscale organization of the synaptic adhesion molecules neurexin-1β,neuroligin-1 (Nlg1) and leucine-rich-repeat transmembrane protein 2 (LRRTM2) in a dual-colour configuration with GFP nanobody,and show that these proteins are diffusionally trapped at synapses where they form apposed trans-synaptic adhesive structures. Furthermore,Nlg1 is dynamic,disperse and sensitive to synaptic stimulation,whereas LRRTM2 is organized in compact and stable nanodomains. Thus,mSA is a versatile tool to image membrane proteins at high resolution in complex live environments,providing novel information about the nano-organization of biological structures. View Publication

产品号#：

05711

产品名：

NeuroCult™ SM1 神经添加物

文献

Takayama Y and Kida YS (FEB 2016) PloS one 11 2 e0148559

In Vitro Reconstruction of Neuronal Networks Derived from Human iPS Cells Using Microfabricated Devices.

Morphology and function of the nervous system is maintained via well-coordinated processes both in central and peripheral nervous tissues,which govern the homeostasis of organs/tissues. Impairments of the nervous system induce neuronal disorders such as peripheral neuropathy or cardiac arrhythmia. Although further investigation is warranted to reveal the molecular mechanisms of progression in such diseases,appropriate model systems mimicking the patient-specific communication between neurons and organs are not established yet. In this study,we reconstructed the neuronal network in vitro either between neurons of the human induced pluripotent stem (iPS) cell derived peripheral nervous system (PNS) and central nervous system (CNS),or between PNS neurons and cardiac cells in a morphologically and functionally compartmentalized manner. Networks were constructed in photolithographically microfabricated devices with two culture compartments connected by 20 microtunnels. We confirmed that PNS and CNS neurons connected via synapses and formed a network. Additionally,calcium-imaging experiments showed that the bundles originating from the PNS neurons were functionally active and responded reproducibly to external stimuli. Next,we confirmed that CNS neurons showed an increase in calcium activity during electrical stimulation of networked bundles from PNS neurons in order to demonstrate the formation of functional cell-cell interactions. We also confirmed the formation of synapses between PNS neurons and mature cardiac cells. These results indicate that compartmentalized culture devices are promising tools for reconstructing network-wide connections between PNS neurons and various organs,and might help to understand patient-specific molecular and functional mechanisms under normal and pathological conditions. View Publication

产品号#：

85850

85857

产品名：

mTeSR™1

文献

Xia N et al. (FEB 2016) Scientific Reports 6 20270

Transcriptional comparison of human induced and primary midbrain dopaminergic neurons

Generation of induced dopaminergic (iDA) neurons may provide a significant step forward towards cell replacement therapy for Parkinson's disease (PD). To study and compare transcriptional programs of induced cells versus primary DA neurons is a preliminary step towards characterizing human iDA neurons. We have optimized a protocol to efficiently generate iDA neurons from human pluripotent stem cells (hPSCs). We then sequenced the transcriptomes of iDA neurons derived from 6 different hPSC lines and compared them to that of primary midbrain (mDA) neurons. We identified a small subset of genes with altered expression in derived iDA neurons from patients with Parkinson's Disease (PD). We also observed that iDA neurons differ significantly from primary mDA neurons in global gene expression,especially in genes related to neuron maturation level. Results suggest iDA neurons from patient iPSCs could be useful for basic and translational studies,including in vitro modeling of PD. However,further refinement of methods of induction and maturation of neurons may better recapitulate full development of mDA neurons from hPSCs. View Publication

产品号#：

85850

85857

产品名：

mTeSR™1

文献

Fuller HR et al. (JAN 2015) Frontiers in cellular neuroscience 9 January 506

Spinal Muscular Atrophy Patient iPSC-Derived Motor Neurons Have Reduced Expression of Proteins Important in Neuronal Development.

Spinal muscular atrophy (SMA) is an inherited neuromuscular disease primarily characterized by degeneration of spinal motor neurons,and caused by reduced levels of the SMN protein. Previous studies to understand the proteomic consequences of reduced SMN have mostly utilized patient fibroblasts and animal models. We have derived human motor neurons from type I SMA and healthy controls by creating their induced pluripotent stem cells (iPSCs). Quantitative mass spectrometry of these cells revealed increased expression of 63 proteins in control motor neurons compared to respective fibroblasts,whereas 30 proteins were increased in SMA motor neurons vs. their fibroblasts. Notably,UBA1 was significantly decreased in SMA motor neurons,supporting evidence for ubiquitin pathway defects. Subcellular distribution of UBA1 was predominantly cytoplasmic in SMA motor neurons in contrast to nuclear in control motor neurons; suggestive of neurodevelopmental abnormalities. Many of the proteins that were decreased in SMA motor neurons,including beta III-tubulin and UCHL1,were associated with neurodevelopment and differentiation. These neuron-specific consequences of SMN depletion were not evident in fibroblasts,highlighting the importance of iPSC technology. The proteomic profiles identified here provide a useful resource to explore the molecular consequences of reduced SMN in motor neurons,and for the identification of novel biomarker and therapeutic targets for SMA. View Publication

产品号#：

05832

85850

85857

产品名：

STEMdiff™ 神经花环选择试剂

mTeSR™1

文献

Ito N et al. (APR 2016) Disease models & mechanisms 9 4 451--462

Decreased N-TAF1 expression in X-linked dystonia-parkinsonism patient-specific neural stem cells.

X-linked dystonia-parkinsonism (XDP) is a hereditary neurodegenerative disorder involving a progressive loss of striatal medium spiny neurons. The mechanisms underlying neurodegeneration are not known,in part because there have been few cellular models available for studying the disease. The XDP haplotype consists of multiple sequence variations in a region of the X chromosome containingTAF1,a large gene with at least 38 exons,and a multiple transcript system (MTS) composed of five unconventional exons. A previous study identified an XDP-specific insertion of a SINE-VNTR-Alu (SVA)-type retrotransposon in intron 32 ofTAF1,as well as a neural-specific TAF1 isoform,N-TAF1,which showed decreased expression in post-mortem XDP brain compared with control tissue. Here,we generated XDP patient and control fibroblasts and induced pluripotent stem cells (iPSCs) in order to further probe cellular defects associated with this disease. As initial validation of the model,we compared expression ofTAF1and MTS transcripts in XDP versus control fibroblasts and iPSC-derived neural stem cells (NSCs). Compared with control cells,XDP fibroblasts exhibited decreased expression ofTAF1transcript fragments derived from exons 32-36,a region spanning the SVA insertion site. N-TAF1,which incorporates an alternative exon (exon 34'),was not expressed in fibroblasts,but was detectable in iPSC-differentiated NSCs at levels that were ∼threefold lower in XDP cells than in controls. These results support the previous findings that N-TAF1 expression is impaired in XDP,but additionally indicate that this aberrant transcription might occur in neural cells at relatively early stages of development that precede neurodegeneration. View Publication

文献

Duan S et al. (DEC 2015) Nature communications 6 10068

PTEN deficiency reprogrammes human neural stem cells towards a glioblastoma stem cell-like phenotype.

PTEN is a tumour suppressor frequently mutated in many types of cancers. Here we show that targeted disruption of PTEN leads to neoplastic transformation of human neural stem cells (NSCs),but not mesenchymal stem cells. PTEN-deficient NSCs display neoplasm-associated metabolic and gene expression profiles and generate intracranial tumours in immunodeficient mice. PTEN is localized to the nucleus in NSCs,binds to the PAX7 promoter through association with cAMP responsive element binding protein 1 (CREB)/CREB binding protein (CBP) and inhibits PAX7 transcription. PTEN deficiency leads to the upregulation of PAX7,which in turn promotes oncogenic transformation of NSCs and instates 'aggressiveness' in human glioblastoma stem cells. In a large clinical database,we find increased PAX7 levels in PTEN-deficient glioblastoma. Furthermore,we identify that mitomycin C selectively triggers apoptosis in NSCs with PTEN deficiency. Together,we uncover a potential mechanism of how PTEN safeguards NSCs,and establish a cellular platform to identify factors involved in NSC transformation,potentially permitting personalized treatment of glioblastoma. View Publication

产品号#：

05700

05701

05702

05750

05850

05857

05870

05875

85850

85857

85870

85875

产品名：

NeuroCult™ 基础培养基(小鼠&大鼠)

NeuroCult™ 扩增添加物 (小鼠&大鼠)

NeuroCult™ 扩增试剂盒 (小鼠&大鼠)

NeuroCult™ NS-A 基础培养基（人）

mTeSR™1

文献

Baarine M et al. (NOV 2015) PLoS ONE 10 11 e0143238

Functional characterization of IPSC-derived brain cells as a model for X-linked adrenoleukodystrophy

X-ALD is an inherited neurodegenerative disorder where mutations in the ABCD1 gene result in clinically diverse phenotypes: the fatal disorder of cerebral childhood ALD (cALD) or a milder disorder of adrenomyeloneuropathy (AMN). The various models used to study the pathobiology of X-ALD disease lack the appropriate presentation for different phenotypes of cALD vs AMN. This study demonstrates that induced pluripotent stem cells (IPSC) derived brain cells astrocytes (Ast),neurons and oligodendrocytes (OLs) express morphological and functional activities of the respective brain cell types. The excessive accumulation of saturated VLCFA,a hallmark" of X-ALD� View Publication

产品号#：

85850

85857

05835

05839

08581

08582

产品名：

mTeSR™1

STEMdiff™ 神经诱导培养基

STEMdiff™SMADi神经诱导试剂盒

STEMdiff™SMADi神经诱导试剂盒，2套

Functionalizing Ascl1 with Novel Intracellular Protein Delivery Technology for Promoting Neuronal Differentiation of Human Induced Pluripotent Stem Cells

Generation of Human Embryonic Stem Cell Line Expressing zsGreen in Cholinergic Neurons Using CRISPR/Cas9 System

C9orf72 Hexanucleotide Expansions are Associated with Altered ER Calcium Homeostasis and Stress Granule Formation in iPSC-Derived Neurons from Patients with Amyotrophic Lateral Sclerosis and Frontotemporal Dementia.

Manifestation of Huntington's disease pathology in human induced pluripotent stem cell-derived neurons.

Modeling Smith-Lemli-Opitz syndrome with induced pluripotent stem cells reveals a causal role for Wnt/&dollar;&dollar;-catenin defects in neuronal cholesterol synthesis phenotypes.

Mapping the dynamics and nanoscale organization of synaptic adhesion proteins using monomeric streptavidin

In Vitro Reconstruction of Neuronal Networks Derived from Human iPS Cells Using Microfabricated Devices.

Transcriptional comparison of human induced and primary midbrain dopaminergic neurons

Spinal Muscular Atrophy Patient iPSC-Derived Motor Neurons Have Reduced Expression of Proteins Important in Neuronal Development.

Decreased N-TAF1 expression in X-linked dystonia-parkinsonism patient-specific neural stem cells.

PTEN deficiency reprogrammes human neural stem cells towards a glioblastoma stem cell-like phenotype.

Functional characterization of IPSC-derived brain cells as a model for X-linked adrenoleukodystrophy

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