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Dataset Information


Analysis of multi-lineage gene expression dynamics during primordial germ cell induction from human induced pluripotent stem cells.

ABSTRACT: BACKGROUND:In mammals, specification of primordial germ cells (PGCs) is established in the early post-implantation embryo. The bone morphogenetic protein (BMP)-SMAD and WNT3-?-catenin signaling initiate the gene regulatory network for PGC specification. The activation of SOX17-BLIMP1 axis is critical for human PGC program. Moreover, EpCAM and INTEGRIN?6 were identified as surface markers of human PGC-like cells (PGCLCs) recently. However, the signaling mechanism for PGC specification in non-rodent mammals remains to be clarified. METHODS:We differentiated human induced pluripotent stem cells (hiPSCs) into PGCLCs in vitro in response to Activin A and BMP4. The percentage of EpCAM/INTEGRIN?6 double-positive cells (PGCLCs) was analyzed by flow cytometry. The expression of PGC genes was evaluated by qRT-PCR and immunofluorescence. The expression dynamic of multi-lineage genes during the differentiation process was evaluated by qRT-PCR. RESULTS:Under the stimulation for PGCLC induction, the embryoids derived from hiPSCs initiated significant upregulation of the early PGC genes (BLIMP1, TFAP2C, and NANOS3), but maintained low or no levels of DPPA3 and late PGC genes (DAZL and DDX4). The percentage of EpCAM/INTEGRIN?6 double-positive PGCLCs reached the highest at day 6 of induction. After pre-induction, the incipient mesoderm-like cells (iMeLCs) upregulated most of the mesoderm genes (EOMES, T, MSXI, RUNX2, and MIXL1). The differentiating embryoids showed high levels of key pluripotency genes, OCT4 and NANOG, but became negative for SOX2. In contrast to iMeLCs, the differentiating embryoids downregulated mesoderm genes RUNX2 and EOMES, and ectoderm gene PAX6, but increased the expression of endoderm gene SOX17. CONCLUSIONS:During PGCLC induction process in vitro, the differentiating embryoids not only activated the PGC-related genes, but also displayed complex regulation of pluripotency genes and multi-lineage genes. These results would be meaningful for future research investigating the regulation of human early germ line development.


PROVIDER: S-EPMC7055065 | BioStudies | 2020-01-01

REPOSITORIES: biostudies

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