Deterministic HOX patterning in human pluripotent stem cell-derived neuroectoderm.
Bottom Line: Despite the precision of HOX patterning in vivo, in vitro approaches for differentiating human pluripotent stem cells (hPSCs) to posterior neural fates coarsely pattern HOX expression thereby generating cultures broadly specified to hindbrain or spinal cord regions.Here, we demonstrate that successive activation of fibroblast growth factor, Wnt/β-catenin, and growth differentiation factor signaling during hPSC differentiation generates stable, homogenous SOX2(+)/Brachyury(+) neuromesoderm that exhibits progressive, full colinear HOX activation over 7 days.This fully defined approach significantly expands capabilities to derive regional neural phenotypes from diverse hindbrain and spinal cord domains.
Affiliation: Department of Biomedical Engineering, University of Wisconsin, Madison, WI 53706, USA; Wisconsin Institute for Discovery, University of Wisconsin, Madison, WI 53706, USA.Show MeSH
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Mentions: Gouti et al. recently demonstrated that simultaneous induction of Wnt/β-catenin and FGF signaling can differentiate hPSCs to SOX2+/Brachyury(T)+ NMPs, but NMPs persisted for only 3 days before shifting to a mesodermal fate under their treatment regimen (Gouti et al., 2014). We observed a similar trend with simultaneous CHIR and FGF8b treatment inducing uniform Brachyury but also causing a sharp decrease in SOX2 expression (23% ± 0% SOX2+), indicating a mesodermal fate shift (Figure 2A, Route 1). Conversely, pre-treatment with FGF8b prior to CHIR (pre-FGF8b/CHIR) yielded uniform expression of both Brachyury and SOX2 that could be maintained (75%–100% SOX2+/Brachyury+) for 7 days (Figure 2A, Route 2, and Figure 2B). Thus, FGF signaling upstream of Wnt/β-catenin signaling effectively induces a stable NMP identity during hPSC differentiation.
Affiliation: Department of Biomedical Engineering, University of Wisconsin, Madison, WI 53706, USA; Wisconsin Institute for Discovery, University of Wisconsin, Madison, WI 53706, USA.