Differentiation of human pluripotent stem cells to multipotent neural crest cells
OwnerStephen Dalton, Laura M. Menendezinventors
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Abstract
The present invention relates to the differentiation of human pluripotent cells, including human pluripotent stems cells to produce a self-renewing multipotent neural crest cell population in a single step method without the requirement of isolation of intermediate cells and without appreciable contamination (in certain preferred instances, virtually none) with Pax6+ neural progenitor cells in the population of p75+ Hnk1+ Ap2+ multipotent neural crest-like cells. The multipotent neural crest cell population obtained can be clonally amplified and maintained for >25 passages (>100 days) while retaining the capacity to differentiate into peripheral neurons, smooth muscle cells and mesenchymal precursor cells.
Claims50
A method of producing p75+ Hnk1+ Ap2+ multipotent neural crest-like cells from pluripotent stem cells comprising differentiating said pluripotent stem cells in a differentiation medium consisting essentially of an effective amount of a Wnt signaling promoter in combination with an effective amount of an Activin A/Smad pathway inhibitor, and optionally, an effective amount of a bone morphogenic protein inhibitor, the resulting neural crest-like stem cells being appreciably free of PAX6+ neural progenitor cells.
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A method of producing p75+ Hnk1+ Ap2+ multipotent neural crest-like cells from human pluripotent stem cells, in particular human embryonic stem cells, comprising differentiating said pluripotent stem cells in a differentiation medium consisting essentially of an effective amount of a Wnt signaling promoter in combination with an effective amount of an Activin A/Smad pathway inhibitor, optionally in combination with an effective amount of a BMP inhibitor for a period of ranging from about 7 to about 20 days, the resulting neural crest-like cells being appreciably free of Pax6+ neural progenitor cells.
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A composition comprising a population of p75+ Hnk1+ Ap2+ multipotent neural crest-like cells in a differentiation medium consisting essentially of at least one Wnt signaling promoter in combination with an Activin A/Smad pathway inhibitor and optionally a BMP inhibitor, said composition being appreciably free of PAX6+ neural progenitor cells.
A composition consisting essentially of a population of p75+ Hnk1+ Ap2+ multipotent neural crest-like stem cells in a differentiation medium produced by exposing human pluripotent stem cells to a combination of effective amounts of BIO, Wnt3a and SB431542, and optionally noggin in a differentiation medium for a period ranging from about 10 to about 14 days, said composition being appreciably free of PAX6+ neural progenitor cells.
Dependencies are inferred from the claim text.