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Fig. 1. Histochemical and fluorescence in situ hybridizations identify the
molecular boundaries of the AHF/SpM in the craniofacial mesoderm at cardiac
crescent stages. The molecular identities of distinct cell populations in
the head were determined, using histochemical in situ hybridization (ISH) and
a double-fluorescence ISH method in St. 8+ chick embryos. AHF/SpM cells are
delineated by dashed ellipses. (A-D) Whole-mount ISH for the indicated
genes. Dashed lines indicate the plane of the stained transverse sections
(A'-D'). Nkx2.5 and Isl1 are
expressed in a broader portion of the SpM (defined as the AHF/SpM), whereas
C-actin and Gata4 are expressed in the more-differentiated
SpM. (E-E'') Fluorescent ISH for Isl1 (E) and
Nkx2.5 (E'), and their overlay (E''), showing that both
genes are expressed in both the differentiated SpM and AHF/SpM.
(F-F'') Fgf10 (F), Nkx2.5 (F') and overlay
(F''). (G-G'') The expression of CPM marker Cyp26c1
(G) and the AHF/SpM marker Nkx2.5 (G') defining the boundary
between the CPM and the AHF/SpM (G''). (H-H'') Expression of
the differentiated SpM marker Gata4 (H) and Nkx2.5
(H'). The merged image (H'') demonstrates that the AHF can be
molecularly identified as comprising Nkx2.5+ and
Gata4- cells. (I-I'') Tbx20
expression (I) marking both differentiated SpM and AHF/SpM. Tbx5
expression (I') labeled only the differentiated SpM. The merged image
(I'') shows that the AHF can be molecularly identified as comprising
Tbx20+/Tbx5- cells. nt, neural tube;
no, notochord; CPM, cranial paraxial mesoderm; AHF/SpM, anterior heart
field/splanchnic mesoderm.
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