Generation of human induced pluripotent stem cells from peripheral blood using the STEMCCA lentiviral vector

Gianotti-Sommer, Andreia

doi:10.3824/stembook.1.93.1

Cited by 6 publications

(6 citation statements)

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“…hiPSCs (BU5, BU6, and BS31) were generated by hSTEMCCA lentiviral transduction of human peripheral blood mononuclear cells as described previously . iPSCs were maintained in mTESR (StemCell Technologies, Vancouver, Canada) on matrigel optimized for hPSC growth (Corning, Corning, NY Matrigel hESC‐qualified Matrix, #354277) and passaged approximately every 7–8 days through ReLeSR (StemCell Tech., Vancouver, Canada) dissociation, following manufacturer instructions.…”

Section: Methodsmentioning

confidence: 99%

Notch and Aryl Hydrocarbon Receptor Signaling Impact Definitive Hematopoiesis from Human Pluripotent Stem Cells

et al. 2018

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Induced pluripotent stem cells (iPSCs) stand to revolutionize the way we study human development, model disease, and eventually, treat patients. However, these cell sources produce progeny that retain embryonic and/or fetal characteristics. The failure to mature to definitive, adult‐type cells is a major barrier for iPSC‐based disease modeling and drug discovery. To directly address these concerns, we have developed a chemically defined, serum and feeder‐free–directed differentiation platform to generate hematopoietic stem‐progenitor cells (HSPCs) and resultant adult‐type progeny from iPSCs. This system allows for strict control of signaling pathways over time through growth factor and/or small molecule modulation. Through direct comparison with our previously described protocol for the production of primitive wave hematopoietic cells, we demonstrate that induced HSPCs are enhanced for erythroid and myeloid colony forming potential, and strikingly, resultant erythroid‐lineage cells display enhanced expression of adult β globin indicating definitive pathway patterning. Using this system, we demonstrate the stage‐specific roles of two key signaling pathways, Notch and the aryl hydrocarbon receptor (AHR), in the derivation of definitive hematopoietic cells. We illustrate the stage‐specific necessity of Notch signaling in the emergence of hematopoietic progenitors and downstream definitive, adult‐type erythroblasts. We also show that genetic or small molecule inhibition of the AHR results in the increased production of CD34+CD45+ HSPCs while conversely, activation of the same receptor results in a block of hematopoietic cell emergence. Results presented here should have broad implications for hematopoietic stem cell transplantation and future clinical translation of iPSC‐derived blood cells. Stem Cells 2018;36:1004–1019

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Section: Methodsmentioning

confidence: 99%

Notch and Aryl Hydrocarbon Receptor Signaling Impact Definitive Hematopoiesis from Human Pluripotent Stem Cells

et al. 2018

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“…iPSC lines (BU6 and BS31 [BR-SP-31-1]) were generated through hSTEMCCA lentiviral transduction of human peripheral blood mononuclear cells as previously described and met stringent quality control parameters for pluripotency and functionality. [30][31][32] iPSCs were maintained in mTESR (StemCell Technologies) on matrigel (Corning Matrigel hESC-qualified Matrix; #354277) using ReLeSR (StemCell Technologies) for passaging.…”

Section: Methods Ipsc Generation and Maintenancementioning

confidence: 99%

Induced pluripotent stem cell–based mapping of β-globin expression throughout human erythropoietic development

et al. 2018

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Robust β-globin expression in erythroid cells derived from induced pluripotent stem cells (iPSCs) increases the resolution with which red blood cell disorders such as sickle cell disease and β thalassemia can be modeled in vitro. To better quantify efforts in augmenting β-globin expression, we report the creation of a β-globin reporter iPSC line that allows for the mapping of β-globin expression throughout human erythropoietic development in real time at single-cell resolution. Coupling this tool with single-cell RNA sequencing (scRNAseq) identified features that distinguish β-globin-expressing cells and allowed for the dissection of the developmental and maturational statuses of iPSC-derived erythroid lineage cells. Coexpression of embryonic, fetal, and adult globins in individual cells indicated that these cells correspond to a yolk sac erythromyeloid progenitor program of hematopoietic development, representing the onset of definitive erythropoiesis. Within this developmental program, scRNAseq analysis identified a gradient of erythroid maturation, with β-globin-expressing cells showing increased maturation. Compared with other cells, β-globin-expressing cells showed a reduction in transcripts coding for ribosomal proteins, increased expression of members of the ubiquitin-proteasome system recently identified to be involved in remodeling of the erythroid proteome, and upregulation of genes involved in the dynamic translational control of red blood cell maturation. These findings emphasize that definitively patterned iPSC-derived erythroblasts resemble their postnatal counterparts in terms of gene expression and essential biological processes, confirming their potential for disease modeling and regenerative medicine applications.

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“…Reprogramming of iPSCs from somatic cells was performed as previously described (Supplementary Fig. 1) (Gianotti-Sommer et al, 2008;Somers et al, 2010;Sommer et al, 2009;Takahashi et al, 2006;Takahashi et al, 2007). We preferentially focused on identifying individuals with homozygous mutations to exclude the potential confounding effect of compound heterozygous mutations from different classes that might obscure the interpretation of CFTR modulator responses.…”

Section: Cf Ipscs Efficiently Differentiate Into Airway Epithelial Spheroidsmentioning

confidence: 99%

A multimodal iPSC platform for cystic fibrosis drug testing

Berical

et al. 2021

Preprint

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Cystic fibrosis (CF) is a monogenic lung disease caused by dysfunction of the cystic fibrosis transmembrane regulator (CFTR) anion channel, resulting in significant morbidity and mortality. The progress in elucidating the role of the CFTR channel using established animal and cell-based models led to the recent discovery of effective CFTR modulators for most individuals with CF. However, a subset of individuals with CF do not respond to these modulators and there is an urgent need to develop novel therapeutic strategies. In this study, we assembled a panel of iPSCs derived from individuals with common or rare variants representative of three distinct classes of CFTR dysfunction. To measure CFTR function in patient-specific iPSCs we adapted two established in vitro assays of CFTR function to iPSC-derived airway cells. In both a 3-D spheroid assay using forskolin-induced swelling as well as planar cultures composed of polarized mucociliary airway epithelial cells, we quantified CFTR baseline function and response to CFTR modulators and detected genotype-specific differences. Our results demonstrate the potential of the human iPSC platform as a research tool to study cystic fibrosis and in particular accelerate therapeutic development for CF caused by rare mutations.

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Generation of human induced pluripotent stem cells from peripheral blood using the STEMCCA lentiviral vector

Cited by 6 publications

References 0 publications

Notch and Aryl Hydrocarbon Receptor Signaling Impact Definitive Hematopoiesis from Human Pluripotent Stem Cells

Notch and Aryl Hydrocarbon Receptor Signaling Impact Definitive Hematopoiesis from Human Pluripotent Stem Cells

Induced pluripotent stem cell–based mapping of β-globin expression throughout human erythropoietic development

A multimodal iPSC platform for cystic fibrosis drug testing

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