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Fig. 2. TGFß induces Smad2 phosphorylation in the yolk sacs of wild-type and
Eng mutant embryos. PSmad1/5/8 (PS1) was detected by
immunohistochemistry in both endothelial and mesothelial cells of wild-type
yolk sacs (A). In Eng mutant yolk sacs, Smad1 was phosphorylated in
mesothelial cells (green arrow) but not in endothelial cells (B); after a 1
hour treatment with BMP2 (C) or TGFß1 (D) PSmad1 was detected in
endothelial cells. PSmad2 (PS2) was detected by immunohistochemistry in the
mesothelial layer of both untreated (E) and TGFß1-treated (F) wild-type
yolk sacs. In the Eng null yolk sac, PSmad2 was not detectable in the
mesothelial layer (G) but phosphorylation of Smad2 was restored after
TGFß1 treatment (green arrow, H) for 1 hour. To confirm the specificity
of PSmad2 antibody, Tgfbr2 null yolk sac were analysed. PSmad2 was
not present in either endothelial or mesothelial cell layers (I) and its
phosphorylation was not restored after TGFß1 treatment (J). (K)
Percentage of endothelial cells expressing PSmad1/5/8 and PSmad2 in sections
from wild-type and Eng mutant treated with BMP2 or treated with
TGFß1. Bars represent the mean±s.e.m. of five independent
experiments. (L) Percentage of mesothelial cells expressing PSmad2 in sections
from wild-type, Eng mutant and
tie-1-Cre/TßRIIfl/fl yolk sacs with
or without 1 hour TGFß1 treatment. Bars represent the mean±s.e.m.
of cells from four independent experiments (*P<0.05;
**P<0.01). Scale bar: 0.5 mm. Abbreviations: EC,
endothelial cell layer; end, endoderm; mes, mesothelial cell layer.
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