Fig. 3. Perturbed homeostasis in the mesoderm and ectoderm component of the amnion in Smad5^m1/m1WT^lacZ and Smad5^m1/m1 embryos. (A,B) Dorsal view of a low-percentage E9.5 Smad5^m1/m1WT^lacZ chimeric embryo, unstained (A) and stained for ß-galactosidase activity (B). The head is patterned normally and the headfolds and neural tube are closed, but embryonic turning is mildly affected (a twisted tail). The amnion remains locally thickened (white arrow), and red blood cells can be observed within this aggregate of cells (A; open arrowhead). (C-E) Section through low- percentage control Smad5^+/m1WT^lacZ (chWT) (C) and Smad5^m1/m1WT^lacZ chimeras (chKO) (D-E). Smad5^+/m1 and Smad5^m1/m1 ES cell derivatives colonize both the mesoderm and ectoderm of the amnion extensively (E). Aggregates of cells on the amnion are of mixed wild type (blue) and Smad5^m1/m1 (purple) origin in a Smad5^m1/m1WT^lacZ chimera (D). The amnion ectoderm is cuboidal in one part of the aggregate of cells (open arrow). Blood vessels develop always in the mesodermal side of the aggregate. In another Smad5^m1/m1WT^lacZ embryo the amnion ectoderm is completely of wild-type origin (E). (F) Twist is expressed in amnion mesoderm, including in the clump of cells in Smad5^m1/m1 mutant amnion, but Twist^neg areas can also be distinguished (dashed circle). (G) Thickened, AP-2⁺ amnion ectoderm is observed locally in a clump of cells (arrow) in a Smad5^m1/m1 mutant amnion. Al, allantois; Am, amnion; AmEc, amnion ectoderm; AmMe, amnion mesoderm; Bv, blood vessel; Ex, extraembryonic region; Fg, foregut; He, heart; Hf, headfold; Nt, neural tissue; Op, optic anlage; So, somite; Ys, yolk sac. Scale bars: 100 µm in A, E-G; 50 µm in B-D.