Dataset Information

NextGen Consortium: Globin Gene Expression in Sickle Cell Genotype-Specific iPS Cells

ABSTRACT:

Implement an efficient, highly reproducible and 'scalable' system for the production of large numbers of sickle cell anemia-specific iPS cells (iPSC).
Derive and characterize a novel, in vitro system for the production of an unlimited supply of erythroid lineage cells from the directed differentiation of 'clinical grade' transgene-free iPS cells; use this system to recapitulate erythroid-lineage ontogeny in vitro with the sequential development of primitive and definitive erythropoiesis, accompanied by the appropriate expression of stage-specific globin genes.
Identify developmental gene expression profile differences between erythroid precursors that produce primarily HbF and those that produce primarily HbA or HbS.
Determine the effects of the three known HbF major quantitative trait loci (QTL) on globin gene expression in disease-specific iPS cells during in vitro erythropoiesis.
Search for novel HbF genetic modifiers associated with markedly elevated HbF levels found in sickle cell anemia patients naturally, or in response to hydroxyurea treatment, by examining gene expression profiles and mRNA sequence of their iPSC-derived erythroid cells.
Develop and use a CRISPR-based gene editing platform to study the effect of novel HbF genetic modifiers, explore globin switching, and correct the HbS mutation in sickle iPSC lines.

PROVIDER: phs001212 | dbGaP |

SECONDARY ACCESSION(S): PRJNA353151PRJNA353152

REPOSITORIES: dbGaP

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Dataset's files

Source:

			Action	DRS
	GapExchange_phs001212.v1.p1.xml	Xml
	dbGaPEx2.1.5.xsd	Other
	Study_Report.phs001212.Globin_SickleCelliPSC.v1.p1.MULTI.pdf	Pdf
	manifest_phs001212.Globin_SickleCelliPSC.v1.p1.c1.GRU.pdf	Pdf
	datadict_v2.xsl	Other

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Project description:Background: Hepatocyte-like cells (HLCs) differentiated from induced pluripotent stem cells (iPSCs) have emerged as a promising cell culture model to study metabolism, biotransformation, viral infections and genetic liver diseases. However the derivation of iPSCs is restricted by ethical and procedural constraints. Furthermore, incomplete HLC differentiation remains a major challenge. iPSC may carry-on a tissue of origin dependent expression memory influencing iPSC differentiation into different cell types. If liver derived iPSCs (Li-iPSCs) maintain a liver specific gene expression profile is not known. It is also not known, if Li-iPSCs would allow the generation of more fully differentiated HLCs.Methods: Primary liver cells (PLCs) were expanded from liver needle biopsies and reprogrammed into Li-iPSCs using a non-integrative Sendai virus-based system. Li-iPSCs were differentiated into HLCs using established differentiation protocols. The HLC phenotype was characterized at the protein, functional and transcriptional level. RNA sequencing data were generated from the originating liver biopsies, the Li-iPSCs, fibroblast derived iPSCs, and differentiated HLCs, and used to characterize and compare their transcriptome profiles.Results: PLCs were obtained from small pieces of liver biopsies and could be reprogrammed into Li-iPSCs. Li-iPSCs indeed retain a liver specific transcriptional footprint. Li-iPSCs can be propagated to provide an unlimited supply of cells for differentiation into Li-HLCs. Similar to HLCs derived from fibroblasts, Li-HLCs could not be fully differentiated into hepatocytes. Relative to the originating liver, Li-HLCs showed lower expression of liver specific transcription factors and increased expression of genes involved in the differentiation of other tissues.Conclusions: PLCs and Li-iPSCs obtained from small pieces of human needle liver biopsies constitute a novel unlimited source for the production of HLCs. Despite the preservation of a liver specific gene expression footprint in Li-iPSCs, the generation of fully differentiated hepatocytes cannot be achieved with the current differentiation protocols.

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NextGen Consortium: Globin Gene Expression in Sickle Cell Genotype-Specific iPS Cells

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