Heightened potency of human pluripotent stem cell lines created by transient BMP4 exposure

Abstract

Human pluripotent stem cells (PSCs) show epiblast-type pluripotency that is maintained with ACTIVIN/FGF2 signaling. Here, we report the acquisition of a unique stem cell phenotype by both human ES cells (hESCs) and induced pluripotent stem cells (iPSCs) in response to transient (24-36 h) exposure to bone morphogenetic protein 4 (BMP4) plus inhibitors of ACTIVIN signaling (A83-01) and FGF2 (PD173074), followed by trypsin dissociation and recovery of colonies capable of growing on a gelatin substratum in standard medium for human PSCs at low but not high FGF2 concentrations. The self-renewing cell lines stain weakly for CDX2 and strongly for NANOG, can be propagated clonally on either Matrigel or gelatin, and are morphologically distinct from human PSC progenitors on either substratum but still meet standard in vitro criteria for pluripotency. They form well-differentiated teratomas in immune-compromised mice that secrete human chorionic gonadotropin (hCG) into the host mouse and include small areas of trophoblast-like cells. The cells have a distinct transcriptome profile from the human PSCs from which they were derived (including higher expression of NANOG, LEFTY1, and LEFTY2). In nonconditioned medium lacking FGF2, the colonies spontaneously differentiated along multiple lineages, including trophoblast. They responded to PD173074 in the absence of both FGF2 and BMP4 by conversion to trophoblast, and especially syncytiotrophoblast, whereas an A83-01/PD173074 combination favored increased expression of HLA-G, a marker of extravillous trophoblast. Together, these data suggest that the cell lines exhibit totipotent potential and that BMP4 can prime human PSCs to a self-renewing alternative state permissive for trophoblast development. The results may have implications for regulation of lineage decisions in the early embryo.biological sciences | developmental biology | pluripotent stem cells | totipotent | trophoblast

Comparison of syncytiotrophoblast generated from human embryonic stem cells and from term placentas

“…1C). Discrete zones of CG-alpha (CGA)-positive cells became visible within the colonies at day 4, and these increased in number and size over subsequent days until, at day 8, they occupied between 5% and 10% of the surface areas of the colonies (17). On this day, many of these areas were greater than 100 μm in diameter and contained many nuclei (Fig.…”

“…used in combination with two small compounds, the activin A signaling inhibitor A83-01, and the FGF2 signaling inhibitor PD173074 (BMP4/A83-01/PD173074; BAP treatment), the process becomes more efficient and synchronous, such that by 48 h, almost all of the cells in the colonies have become transiently positive for the transcription factor CDX2 and completely positive for the trophoblast marker KRT7 (14,17). By day 6 of treatment, areas of syncytium emerge, and the culture medium begins to accumulate significant quantities of the placental hormones, chorionic gonadotropin (CG), placental growth factor (PGF), and progesterone (14).…”

Vulnerability of primitive human placental trophoblast to Zika virus

“…It is now well established that human embryonic stem cells (ESC) and induced pluripotent stem cells can be driven along the trophoblast lineage by exposing them to BMP4 and inhibiting the signaling pathways that maintain the pluripotent phenotype (29)(30)(31). Within 5-6 d of initiating differentiation, areas of syncytium expressing typical markers of STB begin to appear within the colonies.…”

“…They display a progression of differentiation over time, expressing markers of trophoblast stem cells soon after initiating differentiation, but, a few days later, after the stem cell signature genes have more or less been down-regulated, STB formation begins (29,31,44,65,66). By day 8, the transcriptome signature of the ESCd > 70 STB fraction is so different from that of PHTd, the placenta-derived STB fraction, that it is highly improbable that the two could be functional homologs (32).…”

The secretome of MUSE cells contains factors that may play a role in regulation of stemness, apoptosis and immunomodulation

“…They are involved in heightening stem differentiation potential and control cell commitment. [54][55][56] A huge number of transcription factors were identified with IPA and Venn analysis as potential common upstream regulators of protein products evidenced in MSC and Muse cell secretomes. Their activities cover all the aspect of cell biology and it is difficult to relate a single factor with a specific biological activity that could be ascribed to proteins present in secretomes.…”