The Questions of Developmental Biology

Chick cells are readily distinguishable from quail cells because the quail cell nucleus
stains much more darkly (owing to its dense nucleoli), thus providing a permanent marker for
distinguishing the two cell types (see Figure 1.10). Using these "yolk sac chimeras," Dieterlen-
Lièvre and Martin (1981) showed that the yolk sac stem cells do not contribute cells to the adult
animal. Instead, the definitive stem cells are formed within nodes of mesoderm that line the
mesentery and the major blood vessels. In the 4-day chick embryo, the aortic wall appears to be
the most important source of new blood cells, and it has been found to contain numerous
hematopoietic stem cells (Cormier and Dieterlen-Lièvre 1988).
Similarly, studies in fishes, mammals, and frogs indicate that the definitive hematopoietic
cells are formed near the aorta in a domain called the aorta-gonad-mesonephros (AGM) region.
The first blood islands in the mouse embryo appear in the mesoderm around the yolk sac, but by
day 11, pluripotential hematopoietic stem cells and CFU-S cells can be found in the AGM (Kubai
and Auerbach 1983; Godlin et al. 1993; Medvinsky et al. 1993). These are the hematopoietic stem
cells that will colonize the liver and constitute the fetal and adult circulatory system (Medvinsky
and Dzierzak 1996). Müller and colleagues (1994) have proposed that two waves of cells
colonize the fetal liver. According to this hypothesis, the first population of these cells comes
from the yolk sac and comprises predominantly pluripotential stem cells. The majority
population, however, comes from the AGM, and comprises both CFU-S and CFU-M,L cells
(Figure 15.24). This hypothesis was supported by the finding that mice deficient in the
transcription factor AML1 undergo normal yolk sac hematopoiesis, but no definitive (AGM)
hematopoiesis (Okuda et al. 1996). These mutant mice die at embryonic day 12.5. Their livers
contain a small number of primitive nucleated red blood cells, whereas control livers are full of
blood cells derived from the AGM. The AML protein is critical for activating the genes involved
in definitive hematopoiesis. At around the time of birth, stem cells from the liver populate the
bone marrow, which then becomes the major site of blood formation throughout adult life.
The pathways of hematopoiesis are difficult to unravel, but recently a new approach has
gained momentum. Since zebrafish can be easily screened for mutations of developmentally
important genes, over 26 different mutations of hematopoiesis have been found in this species
(Liao and Zon 1999). Some of these, such as the swirl mutation, inhibit the production of ventral
mesoderm. Mutations of the moonshine and cloche genes prevent hemangioblast development;
mutations of frascati and thunderbird act at the level of the pluripotential hematopoietic stem
cells, and several other mutations affect the pathways leading to the various differentiated blood
cell types. By studying gene expression patterns in these mutants, the stage at which the mutant
gene operates can be discerned (Figure 15.25; Thompson et al. 1998). For instance, the cloche
and spadetail mutants have defects in both primitive and definitive hematopoiesis. The cloche
mutant, however, also has defects in vascularization, indicating that cloche works at the level of
the hemangioblast, while the spadetail gene works later, probably at the level of the pluripotential
hematopoietic stem cell.