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Fig. 8. Progenitors with compromised TCF/Lef signaling are depleted from the
epithelial lineage. (A) Metanephric mesenchymes are infected with
Ad-GFP or GFP-tagged dominant-negative TCF (Ad-DN-TCF) followed by culture in
the presence of inductive media containing Tgf
, Fgf2 and Lif and virus
withdrawal after 14 hours. Efficiency of infection of Pax2-positive epithelial
progenitors (red) with Ad-GFP (green, cytoplasmic staining) or Ad-DN-TCF
(green, nuclear staining) is similar as determined after 14 hours of culture.
Conversely, after 48 hours, cells expressing DN-TCF are observed at a
decreased frequency in the Pax2-positive cell population as compared with
cells expressing GFP only. Occasionally, staining for activated caspase 3
(blue) is observed in Pax2-positive cells infected by Ad-DN-TCF (arrowheads),
indicating ongoing apoptosis in this population (note occasional apoptosis
also in the Pax2-negative cell population with both viruses). After 6 days of
culture, DN-TCF-expressing cells (green) are rarely observed in Pax2-positive
Cdh1-negative (Pax2+ Cdh1-) epithelial progenitors or
arising Pax2+ Cdh1+ epithelial clusters, whereas cells
infected with Ad-GFP (green) readily contribute to either of these pools
(asterisks). (B) Quantitative evaluation of percentage of
green-fluorescent cells in either Pax2+ progenitors or
Pax2+ Cdh1+ epithelia demonstrates progressive depletion
from the differentiating epithelial lineage of cells expressing GFP-tagged
DN-TCF, but not GFP only.
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