A Comprehensive, Ethnically Diverse Library of Sickle Cell Disease-Specific Induced Pluripotent Stem Cells

Park, Seonmi; Gianotti-Sommer, Andreia; Molina-Estevez, F. Javier; Vanuytsel, Kim; Skvir, Nicholas; Leung, Amy; Rozelle, Sarah S.; Shaikho, Elmutaz M.; Weir, Isabelle R; Jiang, Zhihua; Luo, Hong-Yuan; Chui, David H.K.; Figueiredo, Maria Stella; Alsultan, A.; Al-Ali, Amein K.; Sebastiani, Paola; Steinberg, Martin H.; Mostoslavsky, Gustavo; Murphy, George J.

doi:10.1016/j.stemcr.2016.12.017

Cited by 50 publications

(50 citation statements)

References 17 publications

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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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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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“…iPSCs were reprogrammed from lymphocytes from the blood of two normal donors and CD34 + cells were isolated from the blood of these same individual for studies using RNA sequencing. iPSCs cells from blood of patients with sickle cell anemia were also reprogramed from peripheral blood lymphocytes and driven toward erythroid differentiation. Both iPSC and CD34 + cells were differentiated into erythroid lineage cells as previously described , (Supporting Information Methods).…”

Section: Methodsmentioning

confidence: 99%

A candidate transacting modulator of fetal hemoglobin gene expression in the Arab—Indian haplotype of sickle cell anemia

et al. 2016

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Fetal hemoglobin (HbF) levels are higher in the Arab-Indian (AI) β-globin gene haplotype of sickle cell anemia compared with African-origin haplotypes. To study genetic elements that effect HbF expression in the AI haplotype we completed whole genome sequencing in 14 Saudi AI haplotype sickle hemoglobin homozygotes—seven selected for low HbF (8.2±1.3%) and seven selected for high HbF (23.5±.2.6%). An intronic single nucleotide polymorphism (SNP) in ANTXR1, an anthrax toxin receptor (chromosome 2p13), was associated with HbF. These results were replicated in two independent Saudi AI haplotype cohorts of 120 and 139 patients, but not in 76 Saudi Benin haplotype, 894 African origin haplotype and 44 Arab Indian haplotype patients of Indian descent, suggesting that this association is effective only in the Saudi AI haplotype background. ANTXR1 variants explained 10% of the HbF variability compared with 8% for BCL11A. These two genes had independent, additive effects on HbF and together explained about 15% of HbF variability in Saudi AI sickle cell anemia patients. ANTXR1 was expressed at mRNA and protein levels in erythroid progenitors derived from induced pluripotent stem cells (iPSCs) and CD34+ cells. As CD34+ cells matured and their HbF decreased ANTXR1 expression increased; as iPSCs differentiated and their HbF increased, ANTXR1 expression decreased. Along with elements in cis to the HbF genes, ANTXR1 contributes to the variation in HbF in Saudi AI haplotype sickle cell anemia and is the first gene in trans to HBB that is associated with HbF only in carriers of the Saudi AI haplotype.

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“…Two independent human iPSC lines, bBU1c2 27 and BU3-NKX2-1 GFP -SFTPC tdTomato 11,13 (BU3NGST) were differentiated into CXCR4/c-Kit +/+ definitive endoderm, then treated with dual-smad inhibition as described above. Endodermal cells were then further incubated in conditions to promote lineage specification through Wnt activation with the GSK3β inhibitor CHIR99021 (CHIR), BMP4, and retinoic acid (RA) (ref.…”

Section: Resultsmentioning

confidence: 99%

Generation of mesenchyme free intestinal organoids from human induced pluripotent stem cells

et al. 2020

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Efficient generation of human induced pluripotent stem cell (hiPSC)-derived human intestinal organoids (HIOs) would facilitate the development of in vitro models for a variety of diseases that affect the gastrointestinal tract, such as inflammatory bowel disease or Cystic Fibrosis. Here, we report a directed differentiation protocol for the generation of mesenchyme-free HIOs that can be primed towards more colonic or proximal intestinal lineages in serum-free defined conditions. Using a CDX2 eGFP iPSC knock-in reporter line to track the emergence of hindgut progenitors, we follow the kinetics of CDX2 expression throughout directed differentiation, enabling the purification of intestinal progenitors and robust generation of mesenchyme-free organoids expressing characteristic markers of small intestinal or colonic epithelium. We employ HIOs generated in this way to measure CFTR function using cystic fibrosis patient-derived iPSC lines before and after correction of the CFTR mutation, demonstrating their future potential for disease modeling and therapeutic screening applications.

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A Comprehensive, Ethnically Diverse Library of Sickle Cell Disease-Specific Induced Pluripotent Stem Cells

Cited by 50 publications

References 17 publications

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

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

A candidate transacting modulator of fetal hemoglobin gene expression in the Arab—Indian haplotype of sickle cell anemia

Generation of mesenchyme free intestinal organoids from human induced pluripotent stem cells

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