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514 results for “Human induced pluripotent stem cells”
Efficient embryoid-based method to improve generation of optic vesicles from human induced pluripotent stem cells data
<p>Animal models have provided many insights into ocular development and disease, but they remain suboptimal for understanding human oculogenesis. Eye development requires spatiotemporal gene expression patterns and disease phenotypes can differ significantly between humans and animal models, with patient-associated mutations causing embryonic lethality reported in some animal models. The emergence of human induced pluripotent stem cell (hiPSC) technology has provided a new resource for dissecting the complex nature of early eye morphogenesis through the generation of three-dimensional (3D) cellular models. By using patient-specific hiPSCs to generate <em>in vitro </em>optic vesicle-like models, we can enhance the understanding of early developmental eye disorders and provide a pre-clinical platform for disease modelling and therapeutics testing. A major challenge of <em>in vitro </em>optic vesicle generation is the low efficiency of differentiation in 3D cultures. To address this, we adapted a previously published protocol of retinal organoid differentiation to improve embryoid body formation using a microwell plate. Established morphology, upregulated transcript levels of known early eye-field transcription factors and protein expression of standard retinal progenitor markers confirmed the optic vesicle/presumptive optic cup identity of <em>in vitro </em>models between day 20 and 50 of culture. This adapted protocol is relevant to researchers seeking a physiologically relevant model of early human ocular development and disease with a view to replacing animal models.</p>
Generation of beta-like cell subtypes from differentiated human induced pluripotent stem cells in 3D spheroids
<p>This repository contains single-cell RNA-sequencing data files (raw FASTQ files generated from Illumina HiSeq sequencing) related to the article entitled "Generation of beta-like cell subtypes from differentiated human induced pluripotent stem cells in 3D spheroids" by Lisa Morisseau et al. (2023) published in the Molecular Omics journal (DOI: 10.1039/d3mo00050h).</p> <p> </p>
GC-MS raw data_Figure 6E_Lysophosphatidic Acid Shifts Metabolic and Transcriptional Landscapes to Induce a Distinct Cellular State in Human Pluripotent Stem Cells
<p><strong>Sample name</strong></p> <p>hESCs (H1 cells) were given treatments for two days and then collected for GC-MS analysis.</p> <p>E8: E8 medium</p> <p>AX: E8 + 1.6% AlbuMAX;</p> <p>BSA: E8 + 1% albumin;</p> <p>BSA+hCDL: E8 + 1% albumin + 0.1% hCDL;</p> <p>LPA+BSA: E8 + 1 μM LPA + 1% albumin;</p> <p>LPA+BSA+hCDL: E8 + 1 μM LPA + 1% albumin + 0.1% hCDL</p> <p>STD: standard lipids mixture used as reference</p> <p><strong>Extraction and Methylation</strong></p> <p>Sample preparation was conducted according to the previously reported method (Araujo et al., 2008) with the modification. Briefly, spent medium was removed, and cells were rinsed with 1 mL/well 0.9% (w/v) saline twice. Then 0.5 mL/well -80°C 80% methanol was added to quench the metabolism. Five wells of cells (from 6-well plate) were scrapped off into a glass screw-cap tube. Then 4 mL heptadecanoate containing chloroform (4 μg/mL, internal standard for fatty acids) was added into the tube. Vortex, and then centrifuge at 2000 rpm for 5 min. Cellular debris was carefully removed, and nitrogen blow the solution till dry. Add 1.5 mL hexane and 1.5 mL 14% boron trifluoride (BF<sub>3</sub>)/methanol solution. Seal the tube with nitrogen gas, heat it at 100°C for 1 h using MK200-2 dry bath incubator (Aosheng), and then cool down to room temperature. Add 1 mL water into the tube, vortex and then centrifuge at 3000 rpm for 10 min. The upper layer was transferred into a new 1.5-mL eppendorf tube and evaporated by nitrogen gas. The residue was re-dissolved in 100 μL hexane for GC-MS analysis.</p> <p><strong>GC-MS method</strong></p> <p>Samples were analyzed using an Agilent GC-MS system (Agilent) consisting of a 6890 gas chromatography and a 5973 mass spectrometer. Fatty acid methyl esters were separated by an Omegawax™ 250 fused silica capillary column (30 m × 0.25 mm i.d., 0.25 μm film thickness, Supelco, Bellefonte, PA). The optimized oven temperature program was: initial temperature set at 180°C and held for 3 min; ramped to 206°C at 2°C/min and held at 206°C for 25 min, then, ramped to 240°C at 10°C/min and held for 5 min. Overall, the total run time was 50 min. Carrier gas was high-purity helium at a flow rate of 1.5 mL/min. Injector temperature was set at 250°C. Injection volume was 2 μL with a split ratio of 1:15. The mass spectrometer was operated in electron-impact (EI) mode at 70 eV ionization energy. The temperatures of quadrupole and ionization source were set at 150°C and 280°C, respectively. The spectra from 3 to 50 min were acquired with the <em>m/z</em> range of 35–550 at a scan rate of 0.34 s per scan.</p>
LC-MS raw data_Lysophosphatidic Acid Shifts Metabolic and Transcriptional Landscapes to Induce a Distinct Cellular State in Human Pluripotent Stem Cells
<p><strong>LC-MS/MS analysis</strong></p> <p><strong>Metabolite extraction</strong></p> <p>For LC-MS/MS quantification, cell sample preparation was conducted as described in the previous literatures (Ying, Kimmelman et al. 2012, Zhang, Badur et al. 2016). Briefly, the spent medium was removed, and cells were rinsed with 1 mL/well 0.9% (w/v) saline twice. Then 0.5 mL/well -80°C 0.2 μg/mL norvaline containing 80% methanol was added to quench the metabolism. Cells were scraped off into 1.5-mL eppendorf tube and stored in -80℃ overnight. The mixtures were vortexed and then centrifuged 12500 × <em>g </em>for 15 min at 4℃. The supernatant was used for LC-MS analysis.</p> <p><strong>LC-MS/MS method</strong></p> <p>Waters Xevo TQD coupled with Waters Acquity UPLC system was used for quantification. Acquity UPLC BEH HILIC column (2.1 × 100 mm, 1.7 μm), Acquity UPLC BEH C18 column (2.1 × 100 mm, 1.7 μm), and Acquity UPLC BEH amide column (2.1 × 100 mm, 1.7 μm) were used for the separation of metabolites. Column temperature was set at 40 °C.</p> <p>For the quantification of norvaline, amino acids, GSH, GSSG, SAH, SAM, ascorbic acid and myo-inositol, amide column was used for the separation. Acetonitrile with 0.1% formic acid (A) and water with 0.1% formic acid (B) were used as mobile phases. The gradient setting is: 0-4 min, 99% A to 90% A; 4-10 min, 90% A to 67% A; 10-13 min, 67% A to 1% A; 13-15 min, 1% A; 15-16.5 min, 1% A to 99% A; 16.5-20 min, 99% A. Flowrate was set as 0.4 mL/min.</p> <p>For the quantification of metabolites involved in TCA cycle, energy related and ribonucleotides, an amide column was used for the separation. Acetonitrile with 0.1% formic acid (A) and water with 0.1% formic acid (B) were used as mobile phases. The gradient setting is: 0-2 min, 80% A; 2-3 min, 80% A to 20% A; 3-5 min, 20% A; 5-6 min, 20% A to 80% A; 6-10 min, 80% A. Flowrate was set as 0.4 mL/min.</p> <p>For the quantification of acetate, acetyl-CoA and metabolites involved in glycolysis and pentose phosphate pathway, HILIC column was used for the separation. Acetonitrile (A) and 10 mM ammonium bicarbonate were used as mobile phases. The gradient setting is: 0-2 min, 10% A; 2-5 min, 10% A to 5% A; 5-6 min, 5% A to 10% A; 6-10 min, 10% A. Flowrate was set as 0.2 mL/min.</p> <p>For the quantification of LPA, LPC and PC, HILIC column was used for the separation. Acetonitrile (A) and 10 mM ammonium bicarbonate aqueous solution (B) were used as mobile phases. The gradient setting is: 0-2 min, 95% A; 2-4 min, 95% A to 10% A; 4-7 min, 10% A; 7-9 min, 10% A to 95% A; 9-15 min, 95% A. Flowrate was set as 0.2 mL/min.</p> <p>For the quantification of CDL lipids, C18 column was used for the separation. 98% Acetonitrile aqueous solution (A) and 10 mM ammonium acetate 90% acetonitrile aqueous solution (B) were used as mobile phases. The gradient setting is: 0-5 min, 0.1% A; 5-6 min, 0.1% A to 99.9% A; 6-11 min, 99.9% A; 11-12 min, 99.9% A to 0.1% A; 12-15 min, 0.1% A. Flowrate was set as 0.4 mL/min.</p> <p>Argon was used as source gas, capillary voltage was 3500 V, and desolvation temperature was 500 °C. Multiple reaction monitoring (MRM) was conducted, and the ion transitions are listed in the supplemental Table S2. Selected ion recording (SIR) was conducted for the detection of CDL-related lipids, and the setting is listed in the supplemental Table S3.</p> <p>Standard solutions of TCA metabolites (100 μg/mL) and intermediates of glycolysis and pentose phosphate pathway (10 μg/mL) were prepared to confirm the retention time. Peak intensity of product ion was used for the quantification. Data analysis was performed by TargetLynx software (Waters) with statistical analysis in Graphpad Prism (version 8.4.0) and R.</p>
Transcriptomic characterization of 2D and 3D human induced pluripotent stem cell-based in vitro models as New Approach Methodologies for developmental neurotoxicity testing
<p><strong>Abstract:</strong> The safety and developmental neurotoxicity (DNT) potential of chemicals remain critically understudied due to limitations of current in vivo testing guidelines, which are low throughput, resource-intensive, and hindered by species differences that limit their relevance to human health. To address these issues, robust new approach methodologies (NAMs) using deeply characterized cell models are essential. This study presents the comprehensive transcriptomic characterization of two advanced human-induced pluripotent stem cell (hiPSC)-derived models: a 2D adherent and a 3D neurosphere model of human neural progenitor cells (hiNPCs) differentiated up to 21 days. Using high-throughput RNA sequencing, we compared gene expression profiles of 2D and 3D models at three developmental stages (3, 14, and 21 days of differentiation). Both models exhibit maturation towards post-mitotic neurons, with the 3D model maturing faster and showing a higher prevalence of GABAergic neurons, while the 2D model is enriched with glutamatergic neurons. Both models demonstrate broad applicability domains, including excitatory and inhibitory neurons, astrocytes, and key endocrine and especially the understudied cholinergic receptors. Comparison with human fetal brain samples confirms their physiological relevance. This study provides novel in-depth applicability insights into the temporal and dimensional aspects of hiPSC-derived neural models for DNT testing. The complementary use of these two models is highlighted: the 2D model excels in synaptogenesis assessment, while the 3D model is particularly suited for neural network formation as observed as well in previous functional studies with these models. This research marks a significant advancement in developing human-relevant, high-throughput DNT assays for regulatory purposes.</p> <p><strong>This data sets contains:</strong></p> <p><strong>Tab. S1</strong> - Significant genes results</p> <p><strong>Tab. S2</strong> - Enriched pathways_GO_Biological Processes</p> <p><strong>Tab. S3</strong> - Enriched pathways_GO_Cellular Components</p> <p><strong>Tab. S4</strong> - Enriched pathways_GO_Molecular Function</p> <p><strong>Tab. S5</strong> - Enriched pathways_KEGG</p> <p><strong>Tab. S6</strong> - EnrichEnriched pathways_Panther</p> <p><strong>Tab. S7</strong> - Enriched pathways_Reactome</p> <p><strong>Tab. S8</strong> - Gene counts</p> <p><strong>Tab. S9</strong> - Gene selection for targeted analysis</p>
A targeted metabolomics-based assay using human induced pluripotent stem cell-derived cardiomyocytes identifies structural and functional cardiotoxicity potential
<p>Implementing screening assays that identify functional and structural cardiotoxicity earlier in the drug development pipeline has the potential to improve safety and the cost and time required to bring new drugs to market. In this study, a metabolic biomarker-based assay was developed that predicts the cardiotoxicity potential of a drug based on changes in the metabolism and viability of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CM). Assay development and testing was conducted in two phases: (1) biomarker identification and (2) targeted assay development. In the first phase, metabolomic data from hiPSC-CM spent media following exposure to 66 drugs was used to identify biomarkers that identified both functional and structural cardiotoxicants. Four metabolites that represent different metabolic pathways (arachidonic acid, lactic acid, 2'-deoxycytidine, and thymidine) were identified as indicators of cardiotoxicity. In phase two, a targeted, exposure-based biomarker assay was developed that measured these metabolites and hiPSC-CM viability across an eight-point concentration curve. Metabolite-specific predictive thresholds for identifying the cardiotoxicity potential of a drug were established and optimized for balanced accuracy or sensitivity. When predictive thresholds were optimized for balanced accuracy, the assay predicted the cardiotoxicity potential of 81 drugs with 86% balanced accuracy, 83% sensitivity, and 90% specificity. Alternatively, optimizing the thresholds for sensitivity yields a balanced accuracy of 85%, 90% sensitivity, and 79% specificity. This new hiPSC-CM-based assay provides a paradigm that can identify structural and functional cardiotoxic drugs that could be used in conjunction with other endpoints to provide a more comprehensive evaluation of a drug's cardiotoxicity potential.</p>
Generation of human induced pluripotent stem cells-derived cortical neurons for high throughput imaging of neurite morphology and neuron maturation
<p>Figure 5: Whole cell patch clamping showed the differentiated neurons are functional.</p>
Generation of human induced pluripotent stem cells-derived cortical neurons for high throughput imaging of neurite morphology and neuron maturation
<p>Figure 3 - Neurite outgrowth dataset. Comparing neurite outgrowth at Day1 and 15 post-seeding cortical neural progenitors. </p>
Safety and Efficacy of Induced Pluripotent Stem Cell-derived Engineered Human Myocardium as Biological Ventricular Assist Tissue in Terminal Heart Failure
ClinicalTrials.gov study NCT04396899. IPD Sharing: Not stated. Countries: 1. Publications: 0.
Generation of Cancer Antigen-Specific T-cells From Human Induced Pluripotent Stem Cells (iPSC) for Research and Potential FutureTherapy
ClinicalTrials.gov study NCT03407040. IPD Sharing: Not stated. Countries: 1. Publications: 3.
Derivation of Induced Pluripotent Stem Cells From an Existing Collection of Human Somatic Cells
ClinicalTrials.gov study NCT00801333. IPD Sharing: Not stated. Countries: 1. Publications: 1.
A targeted metabolomics-based assay using human induced pluripotent stem cell-derived cardiomyocytes identifies structural and functional cardiotoxicity potential
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Dataset related to article "Neonicotinoid pesticides affect developing neurons in experi-mental mouse models and in human induced pluripotent stem cell (iPSC)-derived neural cultures and organoids"
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Neonicotinoid pesticides affect developing neurons in experi-mental mouse models and in human induced pluripotent stem cell (iPSC)-derived neural cultures and organoids
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Data from: Comparison of osteoclast differentiation protocols from human induced pluripotent stem cells of different tissue origins
<p class="MsoNormal"><span class="apple-converted-space"><span><strong>Background</strong>: </span></span><span class="apple-converted-space"><span>Ever since their discovery, induced pluripotent stem cells (iPSCs) have been extensively differentiated into a large variety of cell types. However, a limited amount of work has been dedicated to differentiating iPSCs into osteoclasts. </span></span><span>While several differentiation protocols have been published, it remains unclear which protocols or differentiation methods are preferable regarding the differentiation of osteoclasts.</span></p> <p class="MsoNormal"><span><strong>Methods</strong>: </span><span>In this study, we compare the osteoclastogenesis capacity of a peripheral blood mononuclear cell (PBMC)-derived iPSC line to a fibroblast-derived iPSC line in conjunction with either embryoid body-based or monolayer-based differentiation strategies. Both cell lines and differentiation protocols were investigated regarding their ability to generate osteoclasts and their inherent robustness and ease of use. The ability of both cell lines to remain undifferentiated while propagating using a feeder-free system was assessed using alkaline phosphatase staining. This was followed by evaluating mesodermal differentiation and the characterization of hematopoietic progenitor cells using flow cytometry. Finally, osteoclast yield and functionality based on resorptive activity, Cathepsin K, and tartrate-resistant acid phosphatase (TRAP) expression were assessed. Results were validated using qRT-PCR throughout the differentiation stages.</span></p> <p class="MsoNormal"><span><strong>Results</strong>: </span><span class="apple-converted-space"><span>Embryoid-body-based differentiation yielded CD45<sup>+</sup>, CD14<sup>+</sup>, and CD11b<sup>+</sup> subpopulations, which in turn differentiated into osteoclasts which demonstrated TRAP positivity, Cathepsin K expression, and mineral resorptive capabilities. This was regardless of which iPSC line was used. Monolayer-based differentiation yielded lower quantities of hematopoietic cells that were mostly CD34<sup>+</sup> and did not subsequently differentiate into osteoclasts.</span></span></p> <p class="MsoNormal"><span class="apple-converted-space"><span><strong>Conclusions</strong>: </span></span><span>The outcome of this study demonstrates the successful differentiation of osteoclasts from iPSCs in conjunction with the embryoid-based differentiation method, while the monolayer-based method did not yield osteoclasts. No differences were observed regarding osteoclast differentiation between the PBMC and fibroblast-derived iPSC lines.</span></p>
Modeling Bronchial Epithelium in Severe Asthma With Human Induced Pluripotent Stem Cells (iPSC)
ClinicalTrials.gov study NCT05616338. IPD Sharing: YES. Countries: 1. Publications: 0.
Data from: Comparison of osteoclast differentiation protocols from human induced pluripotent stem cells of different tissue origins
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Effects of Spaceflight on Human Induced Pluripotent Stem Cell-Derived Cardiomyocyte Structure and Function
With extended stays aboard the International Space Station (ISS) becoming commonplace there is a need to better understand the effects of microgravity on cardiac function. We utilized human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) to study the effects of microgravity on cell-level cardiac function and gene expression. The hiPSC-CMs were cultured aboard the ISS for 5.5 weeks and their gene expression structure and functions were compared to ground control hiPSC-CMs. Exposure to microgravity on the ISS caused alterations in hiPSC-CM calcium handling. RNA-sequencing analysis demonstrated 2,635 genes were differentially expressed among flight post-flight and ground control samples including genes involved in mitochondrial metabolism. This study represents the first use of hiPSCs to model the effects of spaceflight on human cardiomyocyte structure and function.
Reprogramming-associated aberrant DNA methylation determines hematopoietic differentiation capacity of human induced pluripotent stem cells [PSCderived_HPCs_methylation]
GEO Series GSE60811. Homo sapiens. 10 samples. Type: Methylation profiling by genome tiling array.
Bulk RNA-seq of neuromuscular system models generated from human induced pluripotent stem cells (hiPSCs)
GEO Series GSE226477. Homo sapiens. 15 samples. Type: Expression profiling by high throughput sequencing.
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Allen Brain Atlas
Allen Brain Atlas is an Allen Institute collection of brain map atlases, datasets, APIs, and analysis tools covering mouse, human, and non-human primate brain resources.
Annotated Behaviour and Observability Dataset (ABODe)
ABODe is a University of Edinburgh DataShare dataset for behavior classification in group-housed mice using home-cage video, identities, bounding boxes, ground-plate positions, and annotator labels.
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