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Fig. 8. Overexpression of cNanog induces growth factor-independent
proliferation in both mESC and cESC. mESC or cESC were transfected with
either mNanog, cNanog or empty expression vector. For mESC, LIF was
removed, clones scored (A), stained for alkaline phosphatase activity
and observed (B-G). Similar results were obtained in two independent
experiments. Scale bars: 400 µm. (H) Gene expression analysis
revealed expression of mOct4 as well as of Sox2, Utf1, Rex1
and Tert, but with the notable exception of Fgf4. A value of
1 was given to the level of expression in the clones obtained without LIF in
the presence of mNanog. (I) For cESC, growth factors and
cytokines (bFgf, Scf, Igf1, Il6, Il6R) (-F) were removed, serum concentration
reduced as indicated (10% to 1%) and clones scored. (J,K) Gene
expression analysis revealed maintenance of cPouV expression in the
presence of 10% serum without factors (J), but a complete loss of expression
when serum was reduced to 1% without factors (K). Under these conditions, the
expression of cNanog is upregulated and the cells continue to
proliferate.
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