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Transcriptomically-guided mesendoderm induction of human pluripotent stem cells using a systematically defined culture scheme

ABSTRACT: Human pluripotent stem cells (hPSCs) are an essential cell source in tissue engineering, studies of development, and disease modeling. Efficient and broadly amenable protocols for rapid lineage induction of hPSCs are of great interest in the stem cell biology field. We describe a simple yet robust method for differentiation of hPSCs into mesendoderm in defined conditions utilizing single-cell seeding and BMP4 and Activin A (BA) treatment. Gene sets and gene ontology terms related to mesoderm and endoderm differentiation were enriched after 48 hours of BA treatment. After 14 days of differentiation, BA-treated cells expressed genes indicative of mesoderm and endoderm-derived tissue types, including lung, kidney and adipose tissue. BA treatment was readily incorporated into existing protocols for chondrogenic and endothelial progenitor cell differentiation. Teratomas formed from BA-treated cells contained higher ratios of mesoderm and endoderm to ectoderm tissue compared to PSC-derived teratomas. Collectively, these data demonstrate that single-cell seeding with BA treatment is a powerful method for induction of mesendoderm that can be integrated into protocols for mesoderm and endoderm differentiation.

ORGANISM(S): Homo sapiens

PROVIDER: GSE129570 | GEO | 2019/04/11

REPOSITORIES: GEO

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Project description:Human pluripotent stem cells (hPSCs) tend to acquire genomic aberrations in culture, the most common of which is the trisomy of chromosome 12. Interestingly, trisomy 12 is also prevalent in germ cell tumors (GCTs). Here, we aimed to dissect the cellular and molecular implications of trisomy 12 in hPSCs. A genome-wide gene expression analysis revealed that trisomy 12 profoundly affects the global gene expression profile of hPSCs, inducing a transcriptional program very similar to that of CGTs. Direct comparison of the proliferation, replication, differentiation and apoptosis between diploid and aneuploid hPSCs revealed that trisomy 12 significantly increases the proliferation rate of hPSCs. Increased replication largely accounts for the increased proliferation observed, and may explain the selection advantage that trisomy 12 confers to hPSCs. A comparison of the tumors induced by diploid and aneuploid hPSCs further demonstrated that trisomy 12 increases the tumorigenicity of hPSCs, inducing transcriptionally-distinct teratomas, from which pluripotent cells can be recovered. Lastly, a chemical screen of 89 anticancer drugs against diploid and aneuploid hPSCs discovered that trisomy 12 raises the sensitivity of hPSCs to several replication inhibitors, suggesting that the increased proliferation and tumorigenicity of these aberrant cells also makes them more vulnerable, and might potentially be used for their selective elimination from culture. Together, our findings demonstrate the extensive effect of trisomy 12 on the gene expression signature and on the cellular behavior of hPSCs, and highlight the danger posed by this trisomy for the successful use of hPSCs in basic research and in regenerative medicine. Expression data from diploid and aneuoploid human pluripotent stem cells, teratomas derived from them, and pluripotent-like cells recovered from these teratomas total RNA was isolated from undifferentiated human pluripotent stem cells grown under standard human ES conditions, or from teratomas derived from them, or from ES-like cells recovered from these teratomas.

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Transcriptomically-guided mesendoderm induction of human pluripotent stem cells using a systematically defined culture scheme

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