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Fig. 6. Generation of mesoderm and the anterior PS from genetically
unmanipulated hES cell lines, KhES-3 and HES-3 cells. (A)
Phase-contrast microscopy of KhES-3 cells, treated with BIO at different
concentrations for 3 days. Scale bar: 100 µm. (B,C) The qPCR
analysis on RNA isolated after 3 days from untreated KhES-3 cells (vehicle, v)
or BIO-treated cells (BIO, 5 µM) with or without Noggin (Nog; 250 ng/ml)
for 3 days was performed using specific primers for the genes indicated.
(D) Expression of N-cadherin and FOXA2 in BIO-treated KhES-3 cells. The
hES cells were cultured with BIO (5 µM) in the presence or absence Noggin
for 3 days, and stained with specific antibodies as indicated. Scale bar: 100
µm. (E) Phase-contrast microscopy and localization of β-catenin
in BIO-Acetoxime-treated HES-3 cells. Cells were cultured for 3 days with
BIO-Acetoxime that exhibits greater selectivity for GSK3 than for BIO (left
panels, phase-contrast images), and subjected to immunostaining with
anti-β-catenin antibody (right panels). Scale bars: 100 µm. (F)
qPCR analysis of RNA isolated from HES-3 cells, with or without BIO-Acetoxime
(0-10 µM) for 3 days, was performed as described in B. (G)
Expression of N-cadherin and FOXA2 in BIO-Acetoxime-treated HES-3 cells. The
HES-3 cells were cultured with BIO-Acetoxime (10 µM) in the presence or
absence Noggin for 3 days, and subjected to immunostaining with
anti-N-cadherin and anti-FOXA2 antibodies. Scale bar: 100 µm.
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