Huang K et al. (JAN 2014)
Science China Life Sciences 57 2 162--70
Neural progenitor cells from human induced pluripotent stem cells generated less autogenous immune response
The breakthrough development of induced pluripotent stem cells (iPSCs) raises the prospect of patient-specific treatment for many diseases through the replacement of affected cells. However,whether iPSC-derived functional cell lineages generate a deleterious immune response upon auto-transplantation remains unclear. In this study,we differentiated five human iPSC lines from skin fibroblasts and urine cells into neural progenitor cells (NPCs) and analyzed their immunogenicity. Through co-culture with autogenous peripheral blood mononuclear cells (PBMCs),we showed that both somatic cells and iPSC-derived NPCs do not stimulate significant autogenous PBMC proliferation. However,a significant immune reaction was detected when these cells were co-cultured with allogenous PBMCs. Furthermore,no significant expression of perforin or granzyme B was detected following stimulation of autogenous immune effector cells (CD3+CD8− T cells,CD3+CD8+ T cells or CD3−CD56+ NK cells) by NPCs in both PBMC and T cell co-culture systems. These results suggest that human iPSC-derived NPCs may not initiate an immune response in autogenous transplants,and thus set a base for further preclinical evaluation of human iPSCs.
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Gasparetto M et al. (OCT 2012)
Experimental hematology 40 10 857--66.e5
Varying levels of aldehyde dehydrogenase activity in adult murine marrow hematopoietic stem cells are associated with engraftment and cell cycle status.
Aldehyde dehydrogenase (ALDH) activity is a widely used marker for human hematopoietic stem cells (HSCs),yet its relevance and role in murine HSCs remain unclear. We found that murine marrow cells with a high level of ALDH activity as measured by Aldefluor staining (ALDH(br) cells) do not contain known HSCs or progenitors. In contrast,highly enriched murine HSCs defined by the CD48(-)EPCR(+) and other phenotypes contain two subpopulations,one that stains dimly with Aldefluor (ALDH(dim)) and one that stains at intermediate levels (ALDH(int)). The CD48(-)EPCR(+)ALDH(dim) cells are virtually all in G(0) and yield high levels of engraftment via both intravenous and intrabone routes. In contrast the CD48(-)EPCR(+)ALDH(int) cells are virtually all in G(1),have little intravenous engraftment potential,and yet can engraft long-term after intrabone transplantation. These data demonstrate that Aldefluor staining of unfractionated murine marrow does not identify known HSCs or progenitors. However,varying levels of Aldefluor staining when combined with CD48 and EPCR detection can identify novel populations in murine marrow including a highly enriched population of resting HSCs and a previously unknown HSC population in G(1) with an intravenous engraftment defect.
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01700
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01702
产品名：
ALDEFLUOR™ 试剂盒
ALDEFLUOR™ DEAB试剂, 1.5 mM, 1 mL
ALDEFLUOR™检测缓冲液
Douvaras P et al. (MAY 2017)
Stem cell reports
Directed Differentiation of Human Pluripotent Stem Cells to Microglia.
Microglia,the immune cells of the brain,are crucial to proper development and maintenance of the CNS,and their involvement in numerous neurological disorders is increasingly being recognized. To improve our understanding of human microglial biology,we devised a chemically defined protocol to generate human microglia from pluripotent stem cells. Myeloid progenitors expressing CD14/CX3CR1 were generated within 30 days of differentiation from both embryonic and induced pluripotent stem cells (iPSCs). Further differentiation of the progenitors resulted in ramified microglia with highly motile processes,expressing typical microglial markers. Analyses of gene expression and cytokine release showed close similarities between iPSC-derived (iPSC-MG) and human primary microglia as well as clear distinctions from macrophages. iPSC-MG were able to phagocytose and responded to ADP by producing intracellular Ca(2+) transients,whereas macrophages lacked such response. The differentiation protocol was highly reproducible across several pluripotent stem cell lines.
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mTeSR™1
mTeSR™1
Park Y et al. (MAR 2014)
Journal of Biotechnology 174 1 39--48
Hepatic differentiation of human embryonic stem cells on microcarriers
Translation of stem cell research to industrial and clinical settings mostly requires large quantities of cells,especially those involving large organs such as the liver. A scalable reactor system is desirable to ensure a reliable supply of sufficient quantities of differentiated cells. To increase the culture efficiency in bioreactor system,high surface to volume ratio needs to be achieved. We employed a microcarrier culture system for the expansion of undifferentiated human embryonic stem cells (hESCs) as well as for directed differentiation of these cells to hepatocyte-like cells. Cells in single cell suspension were attached to the bead surface in even distribution and were expanded to 1??106cells/ml within 2 days of hESC culture with maintenance of the level of pluripotency markers. Directed differentiation into hepatocyte-like cells on microcarriers,both in static culture and stirred bioreactors,induced similar levels of hepatocyte-like cell differentiation as observed with cells cultured in conventional tissue culture plates. The cells expressed both immature and mature hepatocyte-lineage genes and proteins such as asialoglycoprotein receptor-1 (ASGPR-1) and albumin. Differentiated cells exhibited functional characteristics such as secretion of albumin and urea,and CYP3A4 activity could be detected. Microcarriers thus offer the potential for large-scale expansion and differentiation of hESCs induced hepatocyte-like cells in a more controllable bioreactor environment. ?? 2014.
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mTeSR™1
mTeSR™1
Genga RM et al. (MAY 2016)
Methods 101 36--42
Controlling transcription in human pluripotent stem cells using CRISPR-effectors
The ability to manipulate transcription in human pluripotent stem cells (hPSCs) is fundamental for the discovery of key genes and mechanisms governing cellular state and differentiation. Recently developed CRISPR-effector systems provide a systematic approach to rapidly test gene function in mammalian cells,including hPSCs. In this review,we discuss recent advances in CRISPR-effector technologies that have been employed to control transcription through gene activation,gene repression,and epigenome engineering. We describe an application of CRISPR-effector mediated transcriptional regulation in hPSCs by targeting a synthetic promoter driving a GFP transgene,demonstrating the ease and effectiveness of CRISPR-effector mediated transcriptional regulation in hPSCs.
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mTeSR™1
mTeSR™1
Castañ et al. (FEB 2016)
PLoS ONE 11 2 e0149502
SETD7 regulates the differentiation of human embryonic stem cells
The successful use of specialized cells in regenerative medicine requires an optimization in the differentiation protocols that are currently used. Understanding the molecular events that take place during the differentiation of human pluripotent cells is essential for the improvement of these protocols and the generation of high quality differentiated cells. In an effort to understand the molecular mechanisms that govern differentiation we identify the methyltransferase SETD7 as highly induced during the differentiation of human embryonic stem cells and differentially expressed between induced pluripotent cells and somatic cells. Knock-down of SETD7 causes differentiation defects in human embryonic stem cell including delay in both the silencing of pluripotency-related genes and the induction of differentiation genes. We show that SETD7 methylates linker histone H1 in vitro causing conformational changes in H1. These effects correlate with a decrease in the recruitment of H1 to the pluripotency genes OCT4 and NANOG during differentiation in the SETD7 knock down that might affect the proper silencing of these genes during differentiation.
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mTeSR™1
mTeSR™1
Zhang J et al. (NOV 2011)
Stem Cell Reviews and Reports 7 4 987--996
Electrically Guiding Migration of Human Induced Pluripotent Stem Cells
A major road-block in stem cell therapy is the poor homing and integration of transplanted stem cells with the targeted host tissue. Human induced pluripotent stem (hiPS) cells are considered an excellent alternative to embryonic stem (ES) cells and we tested the feasibility of using small,physiological electric fields (EFs) to guide hiPS cells to their target. Applied EFs stimulated and guided migration of cultured hiPS cells toward the anode,with a stimulation threshold of textless30 mV/mm; in three-dimensional (3D) culture hiPS cells remained stationary,whereas in an applied EF they migrated directionally. This is of significance as the therapeutic use of hiPS cells occurs in a 3D environment. EF exposure did not alter expression of the pluripotency markers SSEA-4 and Oct-4 in hiPS cells. We compared EF-directed migration (galvanotaxis) of hiPS cells and hES cells and found that hiPS cells showed greater sensitivity and directedness than those of hES cells in an EF,while hES cells migrated toward cathode. Rho-kinase (ROCK) inhibition,a method to aid expansion and survival of stem cells,significantly increased the motility,but reduced directionality of iPS cells in an EF by 70-80%. Thus,our study has revealed that physiological EF is an effective guidance cue for the migration of hiPS cells in either 2D or 3D environments and that will occur in a ROCK-dependent manner. Our current finding may lead to techniques for applying EFs in vivo to guide migration of transplanted stem cells.
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mTeSR™1
mTeSR™1
Lu Y et al. (FEB 2012)
Stem cells and development 21 3 394--403
Avian-Induced Pluripotent Stem Cells Derived Using Human Reprogramming Factors
Avian species are important model animals for developmental biology and disease research. However,unlike in mice,where clonal lines of pluripotent stem cells have enabled researchers to study mammalian gene function,clonal and highly proliferative pluripotent avian cell lines have been an elusive goal. Here we demonstrate the generation of avian induced pluripotent stem cells (iPSCs),the first nonmammalian iPSCs,which were clonally isolated and propagated,important attributes not attained in embryo-sourced avian cells. This was accomplished using human pluripotency genes rather than avian genes,indicating that the process in which mammalian and nonmammalian cells are reprogrammed is a conserved process. Quail iPSCs (qiPSCs) were capable of forming all 3 germ layers in vitro and were directly differentiated in culture into astrocytes,oligodendrocytes,and neurons. Ultimately,qiPSCs were capable of generating live chimeric birds and incorporated into tissues from all 3 germ layers,extraembryonic tissues,and potentially the germline. These chimera competent qiPSCs and in vitro differentiated cells offer insight into the conserved nature of reprogramming and genetic tools that were only previously available in mammals.
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mTeSR™1
mTeSR™1
Krawetz R and Rancourt DE (JAN 2012)
Methods in molecular biology (Clifton,N.J.) 873 227--235
Suspension bioreactor expansion of undifferentiated human embryonic stem cells
Embryonic stem cells (ESCs) are unique cells,which have the ability to differentiate into all cell types that comprise the adult organism. Furthermore,ESCs can infinitely self-renew under optimized conditions. These features place human ESCs (hESCs) in a position where these cells can be exploited for tissue engineering and regenerative medicine approaches in treating human degenerative disorders. However,cell therapy approaches will require large amounts of clinically useable cells,not typically achievable using standard static cell culture methods. Here,we describe a method wherein clinically relevant numbers of hESCs can be generated in a cost and time effective manner.
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07920
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ACCUTASE™
mTeSR™1
mTeSR™1
ACCUTASE™
R. J. Ihry et al. ( 2018)
Nature Medicine
P53 inhibits CRISPR-Cas9 engineering in human pluripotent stem cells
Economic development has become a prominent issue for state governments. Nevertheless,states vary in the economic policies they choose. Two general approaches to the issue are discussed: the maintenance/attraction strategy and the creation strategy. Factor analysis allows us to gauge state effort on these two criteria. Regression analysis shows that political culture is an important factor in predicting which approach a state chooses,with traditionalistic states favoring the maintenance/attraction strategy,and moralistic states favoring the creation alternative. Other predictors of state policy choices include the condition of the economy and the diffusion of innovations. Also discussed is the interaction of political culture with other relevant variables in shaping state policies.
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(Feb 2025)
Nature Communications 16
Single cell suppression profiling of human regulatory T cells
Regulatory T cells (Treg) play an important role in regulating immune homeostasis in health and disease. Traditionally their suppressive function has been assayed by mixing purified cell populations,which does not provide an accurate picture of a physiologically relevant response. To overcome this limitation,we here develop ‘single cell suppression profiling of human Tregs’ (scSPOT). scSPOT uses a 52-marker CyTOF panel,a cell division detection algorithm,and a whole PBMC system to assess the effect of Tregs on all other cell types simultaneously. In this head-to-head comparison,we find Tregs having the clearest suppressive effects on effector memory CD8 T cells through partial division arrest,cell cycle inhibition,and effector molecule downregulation. Additionally,scSPOT identifies a Treg phenotypic split previously observed in viral infection and propose modes of action by the FDA-approved drugs Ipilimumab and Tazemetostat. scSPOT is thus scalable,robust,widely applicable,and may be used to better understand Treg immunobiology and screen for therapeutic compounds. Traditional regulatory T cell (Tregs) assays utilize mixture of purified cell population. Here the authors develop a ‘single cell suppression profiling of human Tregs’ (scSPOT) with 52-marker CyTOF panel,a cell division detection algorithm,and a whole PBMC system to assess Treg suppressive function on all cell types simultaneously.
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EasySep™小鼠TIL（CD45）正选试剂盒
科学海报A Simple, Reproducible Method to Generate Red Blood Cells From Human Pluripotent Stem Cells
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