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Fig. 2. FGFR-ERK1/2 signaling is activated in migrating PGCs. (A) Whole-mount
dp-ERK1/2 (red) staining in transverse slices from E9.5 OCT4
PE:GFP
embryos; germ cells are shown in green. To show the specificity of the
dp-ERK1/2 antibody, trunk regions of embryos were cultured with (right) or
without (center) MEK inhibitor (50 µM of U0120) for 4 hours, fixed, sliced
and stained for dp-ERK1/2. Slices in which the primary antibody was left out
of the protocol (left) were used as negative controls. (B-D) Higher
magnification views of hind gut regions of the embryos shown in A. Confocal
sections reveal that cytoplasmic staining observed in PGCs of the control
slice (arrowheads in C) was diminished in the U0126-treated slice (D) and the
negative control slice (B). (E-H) The percentage of dp-ERK1/2-positive PGCs
was dramatically affected by 4-hour treatments of FGFs (FGF7 and FGF2) or FGFR
inhibitor (SU5402). (E) Dp-ERK1/2 staining of control embryo cultured with
DMSO. The numbers of dp-ERK1/2-positive PGCs (red; arrows) were increased in
embryos incubated with (F) 10 ng/ml of FGF7 or (G) 10 ng/ml of FGF2, and
decreased in embryos treated with (H) 5 µM of SU5402. (B-H) dp-ERK1/2
images are on the right; GFP overlaid image, left. (I) The percentage of
dp-ERK1/2-positive PGCs obtained from each treatment is summarized. Error bars
show the s.e.m. Asterisks indicate the degree of statistical significance
(P<0.01), of increase (*) and decrease (**)
compared with the controls. Scale bars: in A, 200 µm; in B, 20 µm for
B-D; in E, 40 µm for E-H.
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