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Fig. 8. Steel is required for proper germ cell localization and for
proliferation at E10.5. (A-I) Bax/Steel embryos were
immunostained for phospho-histone H3, cleaved Parp, and the PGC marker SSEA1.
(A-C) Bax-/- Steel+/+,
(D-F) Bax+/- Steel+/-, and
(G-I) Bax-/- Steel-/-.
Apoptotic PGCs (arrowheads) were enriched in Bax+/-
embryos that lacked at least one allele of Steel (D). Proliferating
germ cells (arrows) were enriched in embryos with an intact allele of
Steel (C,F). Germ cells in Bax/Steel DKO embryos were
mislocalized (G-I), and were found in the ventral aspect of the hindgut and
further ventral structures (arrows), with few near the genital ridges
(arrowhead). SSEA1 staining (B,E,H) confirmed the presence of PGCs,
independent of Oct4. No differences in PGC numbers or locations were observed
between SSEA1 stained and unstained slices. (J)
Cleaved-PARP+ PGCs from Bax+/-
Steel+/- (D) and Bax-/-
Steel-/- (G) embryos were compared and found to be
significantly reduced in the absence of Bax. (K) Phospho-histone
H3-positive PGCs were significantly reduced in Steel-/-
embryos (I) compared with Steel heterozygous (F) and wild-type (C)
littermates. (L) PGCs were reduced in Steel heterozygous
compared with wild-type embryos, and nearly absent in
Steel-/- embryos. The additional loss of Bax rescues PGC
numbers to E9.0 levels. 
P<0.01 for
Bax+/- Steel-/- vs
Bax-/- Steel-/- embryos. (M)
The percentage of ectopic germ cells in Bax/Steel embryos is markedly
higher in Steel-null embryos. Horizontal brackets indicate samples that were
compared for statistical significance measurements.
P<0.05,
P<0.01,
P<0.001. Error bars represent
means ± s.e.m.
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