Establishment and characterization of induced pluripotent stem cell line (IGIBi002-A) from a β-thalassemia patient with IVS1-5 mutation by non-integrating reprogramming approach

Thakur, Priya; Bhargava, Nupur; Jaitly, Shashank; Gupta, Pragya; Bhattacharya, Sumitra; Padma, G.; Kondaveeti, Saroja; Jain, Suman; Ramalingam, Sivaprakash

doi:10.1016/j.scr.2020.102124

Cited by 5 publications

(2 citation statements)

References 5 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The development of immortalized erythroid progenitors using CRISPR technology to create isogenic cell lines for SCD and BT has been a highly sought-after goal in the field of erythroid biology. Despite several studies generating hiPSC lines from SCD and BT patients 20 – 23 no in vitro cellular system could produce reticulocytes equivalent to primary adult reticulocytes for these two blood disorders 24 . Recently, an immortalized erythroid line siBBE, was generated from peripheral blood stem cells of a HBE/β-thalassemia patient 25 , but the pathophysiological characteristics of this line were not directly compared with primary patient-derived cells.…”

Section: Discussionmentioning

confidence: 99%

Development of pathophysiologically relevant models of sickle cell disease and β-thalassemia for therapeutic studies

Gupta,

Goswami,

Kumari

et al. 2024

Nat Commun

Self Cite

View full text Add to dashboard Cite

Ex vivo cellular system that accurately replicates sickle cell disease and β-thalassemia characteristics is a highly sought-after goal in the field of erythroid biology. In this study, we present the generation of erythroid progenitor lines with sickle cell disease and β-thalassemia mutation using CRISPR/Cas9. The disease cellular models exhibit similar differentiation profiles, globin expression and proteome dynamics as patient-derived hematopoietic stem/progenitor cells. Additionally, these cellular models recapitulate pathological conditions associated with both the diseases. Hydroxyurea and pomalidomide treatment enhanced fetal hemoglobin levels. Notably, we introduce a therapeutic strategy for the above diseases by recapitulating the HPFH3 genotype, which reactivates fetal hemoglobin levels and rescues the disease phenotypes, thus making these lines a valuable platform for studying and developing new therapeutic strategies. Altogether, we demonstrate our disease cellular systems are physiologically relevant and could prove to be indispensable tools for disease modeling, drug screenings and cell and gene therapy-based applications.

show abstract

Section: Discussionmentioning

confidence: 99%

Development of pathophysiologically relevant models of sickle cell disease and β-thalassemia for therapeutic studies

Gupta,

Goswami,

Kumari

et al. 2024

Nat Commun

Self Cite

View full text Add to dashboard Cite

show abstract

“…The hiPSC colonies from human control DYR0100 (ATCC; ACS-1011) and two patientderived hiPSCs lines, IGIBi001-A (Bhargava et al, 2019) and IGIBi002-A (Thakur et al, 2021) were dissociated into single cells and plated at a density of 2500 cells per cm 2 in 24 well dishes using different dissociation reagents, TrypLE, 0.5mM EDTA, ReleSR, Accutase and Gentle Cell Dissociation reagent (GCDR), in StemFlex medium supplemented with 1X RevitaCell. Media was changed every day and harvested for counting after 60 hours.…”

Section: Evaluation Of Different Cell Dissociation Reagentsmentioning

confidence: 99%

Development of an efficient single-cell cloning and expansion strategy for genome edited induced pluripotent stem cells

et al. 2022

Self Cite

View full text Add to dashboard Cite

Disease-specific human induced pluripotent stem cells (hiPSCs) can be generated directly from individuals with known disease characteristics or alternatively be modified using genome editing approaches to introduce disease causing genetic mutations to study the biological response of those mutations. The genome editing procedure in hiPSCs is still inefficient, particularly when it comes to homology directed repair (HDR) of genetic mutations or targeted transgene insertion in the genome and single cell cloning of edited cells. In addition, genome editing processes also involve additional cellular stresses such as trouble with cell viability and genetic stability of hiPSCs. Therefore, efficient workflows are desired to increase genome editing application to hiPSC disease models and therapeutic applications. Apart from genome editing efficiency, hiPSC survival following single-cell cloning has proved to be challenging and has thus restricted the capability to easily isolate homogeneous clones from edited hiPSCs. To this end, we demonstrate an efficient workflow for feeder-free single cell clone generation and expansion in both CRISPR-mediated knock-out (KO) and knock-in (KI) hiPSC lines. Using StemFlex medium and CloneR supplement in conjunction with Matrigel cell culture matrix, we show that cell viability and expansion during single-cell cloning in edited and unedited cells is significantly enhanced. Our reliable single-cell cloning and expansion workflow did not affect the biology of the hiPSCs as the cells retained their growth and morphology, expression of various pluripotency markers and normal karyotype. This simplified and efficient workflow will allow for a new level of sophistication in generating hiPSC-based disease models to promote rapid advancement in basic research and also the development of novel cellular therapeutics.

show abstract

An Overview on Promising Somatic Cell Sources Utilized for the Efficient Generation of Induced Pluripotent Stem Cells

Ray

Joshi

Sundaravadivelu

et al. 2021

Stem Cell Rev and Rep

View full text Add to dashboard Cite

Establishment and characterization of induced pluripotent stem cell line (IGIBi002-A) from a β-thalassemia patient with IVS1-5 mutation by non-integrating reprogramming approach

Cited by 5 publications

References 5 publications

Development of pathophysiologically relevant models of sickle cell disease and β-thalassemia for therapeutic studies

Development of pathophysiologically relevant models of sickle cell disease and β-thalassemia for therapeutic studies

Development of an efficient single-cell cloning and expansion strategy for genome edited induced pluripotent stem cells

An Overview on Promising Somatic Cell Sources Utilized for the Efficient Generation of Induced Pluripotent Stem Cells

Contact Info

Product

Resources

About