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Fig. 8. ß-catenin overexpression results in a less restricted population of
neurons compared with RA-derived neurons. The phenotype of neurons generated
by ß-catenin 
N and C overexpression in high-density EB
cultures was compared with neurons generated by RA treatment of low-density EB
cultures. (A-D) Hoxc4, red; ß-tubulin 3, green; (E-H) NeuN, green; GABA,
red; (I-L) tyrosine hydroxylase, red; ß-tubulin 3, green; Hoechst stain
blue (white arrows indicate TH-positive neurons); (M-P) Map2, green;
ß-tubulin 3, red; (Q-T) synaptophysin, green; ß-tubulin 3, red.
(A-D) All neurons generated by either ß-catenin overexpression or RA
treatment were positive for Hoxc4, a homeobox gene specific for caudal
neurons. (E-H) Many neurons were positive for the neurotransmitter GABA and
all neurons were positive for NeuN. (I-L) Some neurons induced by
overexpression of ß-catenin were immunoreactive for tyrosine hydroxylase
(TH), the rate-limiting enzyme in catecholamine biosynthesis. By contrast, TH
immunoreactivity was not observed in RA-treated conditions. (M-T) All
ß-tubulin 3-immunoreactive cells also expressed Map2 and synaptophysin.
(U) Quantification of the percentage of gabaergic neurons found in the
cultures. There was no statistically significant difference between the
RA-treated and untreated cultures (by ANOVA). (V) Neurons generated from
untransfected ES cells differentiated at low density also express Hoxc4,
suggesting that these are caudal neurons (green, Hoxc4; red, ß-tubulin
3).
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