Current Protocols in Stem Cell Biology最新文献_第8页

Development of a Three-Dimensional Bioengineering Technology to Generate Lung Tissue for Personalized Disease Modeling 三维生物工程技术生成肺组织用于个性化疾病建模的发展

Current Protocols in Stem Cell Biology Pub Date : 2018-06-07 DOI: 10.1002/cpsc.56

Dan C. Wilkinson, Michael Mellody, Luisa K. Meneses, Ashley C. Hope, Bruce Dunn, Brigitte N. Gomperts

引用次数: 108

Cancer Stem Cell Migration in Three-Dimensional Aligned Collagen Matrices 肿瘤干细胞在三维排列的胶原基质中的迁移

Current Protocols in Stem Cell Biology Pub Date : 2018-05-24 DOI: 10.1002/cpsc.57

Arja Ray, Rachel K. Morford, Paolo P. Provenzano

引用次数: 5

Fluorescence Endomicroscopy Imaging of Mesenchymal Stem Cells in the Rat Lung 大鼠肺间充质干细胞的荧光内镜成像

Current Protocols in Stem Cell Biology Pub Date : 2018-05-04 DOI: 10.1002/cpsc.52

Jessica R. Perez, Norma Ybarra, Frederic Chagnon, Olivier Lesur, Jan Seuntjens, Issam El Naqa

引用次数: 1

Generation of Human Induced Pluripotent Stem Cells Using a Defined, Feeder-Free Reprogramming System 使用一种明确的、无馈源的重编程系统生成人类诱导多能干细胞

Current Protocols in Stem Cell Biology Pub Date : 2018-05-04 DOI: 10.1002/cpsc.48

Seonmi Park, Gustavo Mostoslavsky

引用次数: 13

Derivation of Epithelial-Only Airway Organoids from Human Pluripotent Stem Cells 从人多能干细胞中衍生上皮性气道类器官

Current Protocols in Stem Cell Biology Pub Date : 2018-05-04 DOI: 10.1002/cpsc.51

Katherine B. McCauley, Finn Hawkins, Darrell N. Kotton

{"title":"Derivation of Epithelial-Only Airway Organoids from Human Pluripotent Stem Cells","authors":"Katherine B. McCauley, Finn Hawkins, Darrell N. Kotton","doi":"10.1002/cpsc.51","DOIUrl":"10.1002/cpsc.51","url":null,"abstract":"New protocols to efficiently generate functional airway epithelial organoids from human pluripotent stem cells (PSCs) would represent a major advance towards effective disease modeling, drug screening and cell based therapies for lung disorders. This unit describes an approach using stage-specific signaling pathway manipulation to differentiate cells to proximal airway epithelium via key developmental intermediates. Cells are directed via definitive endoderm (DE) to anterior foregut, and then specified to NKX2-1+ lung epithelial progenitors. These lung progenitors are purified using cell surface marker sorting and replated in defined culture conditions to form three-dimensional, epithelial-only airway organoids. This directed differentiation approach using serum-free, defined media also includes protocols for evaluation of DE induction, intracellular FACS analysis of NKX2-1 specification efficiency and enrichment, and approaches for characterization and expansion of airway organoids. Taken together, this represents an efficient and reproducible approach to generate expandable airway organoids from human PSCs for use in numerous downstream applications. © 2018 by John Wiley & Sons, Inc.","PeriodicalId":53703,"journal":{"name":"Current Protocols in Stem Cell Biology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2018-05-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/cpsc.51","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"36337446","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 40

Methods for Generating Vascularized Islet-Like Organoids Via Self-Condensation 血管化胰岛样器官的自凝生成方法

Current Protocols in Stem Cell Biology Pub Date : 2018-05-04 DOI: 10.1002/cpsc.49

Yoshinobu Takahashi, Takanori Takebe, Hideki Taniguchi

引用次数: 21

CRISPR/Cas9-based Targeted Genome Editing for the Development of Monogenic Diseases Models with Human Pluripotent Stem Cells 基于CRISPR/ cas9的靶向基因组编辑用于人类多能干细胞单基因疾病模型的开发

Current Protocols in Stem Cell Biology Pub Date : 2018-04-26 DOI: 10.1002/cpsc.50

Navin Gupta, Koichiro Susa, Yoko Yoda, Joseph V. Bonventre, M. Todd Valerius, Ryuji Morizane

{"title":"CRISPR/Cas9-based Targeted Genome Editing for the Development of Monogenic Diseases Models with Human Pluripotent Stem Cells","authors":"Navin Gupta, Koichiro Susa, Yoko Yoda, Joseph V. Bonventre, M. Todd Valerius, Ryuji Morizane","doi":"10.1002/cpsc.50","DOIUrl":"10.1002/cpsc.50","url":null,"abstract":"Human pluripotent stem cells (hPSCs) represent a formidable tool for disease modeling, drug discovery, and regenerative medicine using human cells and tissues in vitro. Evolving techniques of targeted genome editing, specifically the CRISPR/Cas9 system, allow for the generation of cell lines bearing gene-specific knock-outs, knock-in reporters, and precise mutations. However, there are increasing concerns related to the transfection efficiency, cell viability, and maintenance of pluripotency provided by genome-editing techniques. The procedure presented here employs transient antibiotic selection that overcomes reduced transfection efficiency, avoids cytotoxic flow sorting for increased viability, and generates multiple genome-edited pluripotent hPSC lines expanded from a single parent cell. Avoidance of xenogeneic contamination from feeder cells and reduced operator workload, owing to single-cell passaging rather than clump passaging, are additional benefits. The outlined methods may enable researchers with limited means and technical experience to create human stem cell lines containing desired gene-specific mutations. © 2018 by John Wiley & Sons, Inc.","PeriodicalId":53703,"journal":{"name":"Current Protocols in Stem Cell Biology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2018-04-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/cpsc.50","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"36337445","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 9

Stem Cell–Derived Retinal Pigment Epithelial Layer Model from Adult Human Globes Donated for Corneal Transplants 角膜移植捐献成人眼球干细胞源性视网膜色素上皮层模型

Current Protocols in Stem Cell Biology Pub Date : 2018-04-26 DOI: 10.1002/cpsc.53

Marie Fernandes, Brian McArdle, Lauren Schiff, Timothy A. Blenkinsop

引用次数: 12

Derivation of Multipotent Mesenchymal Stromal Cells from Ovine Bone Marrow 绵羊骨髓多能间充质间质细胞的制备

Current Protocols in Stem Cell Biology Pub Date : 2018-02-28 DOI: 10.1002/cpsc.43

Daniel Vivas, Marta Caminal, Irene Oliver-Vila, Joaquim Vives

{"title":"Derivation of Multipotent Mesenchymal Stromal Cells from Ovine Bone Marrow","authors":"Daniel Vivas, Marta Caminal, Irene Oliver-Vila, Joaquim Vives","doi":"10.1002/cpsc.43","DOIUrl":"10.1002/cpsc.43","url":null,"abstract":"In the field of orthopedics, translational research of novel therapeutic approaches involves the use of large animal models (such as sheep, goat, pig, dog, and horse) due to the similarities with humans in weight, size, joint structure, and bone/cartilage healing mechanisms. Particularly in the development of cell-based therapies, the lack of manageable immunocompromised preclinical large animal models prevents the use of human cells, which makes it necessary to produce equivalent homologous cell types for the study of their pharmacodynamics, pharmacokinetics, and toxicology. The methods described herein allow for the isolation, expansion, manipulation, and characterization of fibroblastic-like ovine bone marrow–derived multipotent mesenchymal stromal cells (BM-MSC) that, similar to human BM-MSC, adhere to standard plastic surfaces; express specific surface markers such as CD44, CD90, CD140a, CD105, and CD166; and display trilineage differentiation potential in vitro. Homogeneous cell cultures result from a 3-week bioprocess yielding cell densities in the range of 2–4 × 104 MSC/cm2 at passage 2, which corresponds to ∼8 cumulative population doublings. Large quantities of BM-MSC resulting from following this methodology can be readily used in proof of efficacy and safety studies in the preclinical development stage. © 2018 by John Wiley & Sons, Inc.","PeriodicalId":53703,"journal":{"name":"Current Protocols in Stem Cell Biology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2018-02-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/cpsc.43","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"35889246","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 21

Conditional Manipulation of Gene Function in Human Cells with Optimized Inducible shRNA 利用优化的可诱导shRNA条件操纵人细胞的基因功能

Current Protocols in Stem Cell Biology Pub Date : 2018-02-28 DOI: 10.1002/cpsc.45

Alessandro Bertero, Loukia Yiangou, Stephanie Brown, Daniel Ortmann, Matthias Pawlowski, Ludovic Vallier

{"title":"Conditional Manipulation of Gene Function in Human Cells with Optimized Inducible shRNA","authors":"Alessandro Bertero, Loukia Yiangou, Stephanie Brown, Daniel Ortmann, Matthias Pawlowski, Ludovic Vallier","doi":"10.1002/cpsc.45","DOIUrl":"10.1002/cpsc.45","url":null,"abstract":"The difficulties involved in conditionally perturbing complex gene expression networks represent major challenges toward defining the mechanisms controlling human development, physiology, and disease. We developed an OPTimized inducible KnockDown (OPTiKD) platform that addresses the limitations of previous approaches by allowing streamlined, tightly-controlled, and potent loss-of-function experiments for both single and multiple genes. The method relies on single-step genetic engineering of the AAVS1 genomic safe harbor with an optimized tetracycline-responsive cassette driving one or more inducible short hairpin RNAs (shRNAs). OPTiKD provides homogeneous, dose-responsive, and reversible gene knockdown. When implemented in human pluripotent stem cells (hPSCs), the approach can be then applied to a broad range of hPSC-derived mature cell lineages that include neurons, cardiomyocytes, and hepatocytes. Generation of OPTiKD hPSCs in commonly used culture conditions is simple (plasmid based), rapid (two weeks), and highly efficient (>95%). Overall, this method facilitates the functional annotation of the human genome in health and disease. © 2018 by John Wiley & Sons, Inc.","PeriodicalId":53703,"journal":{"name":"Current Protocols in Stem Cell Biology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2018-02-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/cpsc.45","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"35890119","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 9