112
FRED H. WILT
formation and erythropoiesis ensued in the endoderm-mesoderm combinations. If endoderm was removed prior to condensation of mesoderm
into blood islands, the ectoderm-mesoderm isolates showed a peculiar
behavior. Very small numbers of red blood cells did form, and they
contained Hb, but no endothelium developed. It was concluded that the
endoderm played: (1) an important role in organization of mesoderm
into blood islands which leads to endothelium formation; and (2) was
a potent stimulus for the number of red blood cells formed and amount
of Hb synthesized. The possibility that material may pass from endoderm
to blood island cells and play a role in the initiation of Hb synthesis
was proposed, and the observations of Mato et al. (1964) are consistent
with this view.
It should be noted here that the results of experiments on Hb formation
may be influenced by the conditions of culture. In general, erythrocyte
formation and Hb synthesis are not as sensitive to poor culture conditions as many other concurrent morphogenetic events, e.g., somite and
heart formation. Almost any available organ culture method in which
the tissue is at a medium-gas interface is adequate, especially blastoderms (or pieces from them) having attained the 1-somite stage. However, the frequency of erythrocyte formation and general health of the
culture is much better in rich media, and for this purpose the whole-egg
agar medium of Spratt is very good (Spratt and Haas, 1960a). A more
marked sensitivity to the type of medium is apparent if pre-streak and
early-streak blastoderms are used, and organ cultures of unincubated
blastoderms are quite fastidious, really only doing well in our experience
on Spratt's whole egg medium.
The time when Hb first appears in the development of the chick
embryo is fairly well settled; a variety of different techniques show it
is first detectable at the 6- or 7-somite stage. The practiced eye can
visualize the faint red color of Hb in the living blastoderm by the 10- to
12-somite stages. While techniques sensitive enough to reveal the presence of the very first molecules of Hb are not available, there is little
question that an accelerated rate of synthesis has commenced by the
7-somite stage. The benzidine stain shows that Hb is present in blood
island cells of the 7-somite blastoderm (Slonimski, 1927; O'Brien, 1960,
1961). Hb is not detected elsewhere at this stage by these techniques.
O'Brien (1960) states that the stainable Hb is first seen in the cell
nucleus, and subsequently a halo of Hb surrounds the nucleus; soon
thereafter the entire cell stains for Hb. It is of real interest to determine
if Hb synthesis is first initiated in the nucleus. Hammel and Bessman
(1964) have provided evidence that the isolated nucleus of duck reticulocytes may synthesize some Hb (cf. Mirsky and Allfrey, 1958). Efforts
to detect a concentration of protein-bound heme by autoradiographic
FRED H. WILT
formation and erythropoiesis ensued in the endoderm-mesoderm combinations. If endoderm was removed prior to condensation of mesoderm
into blood islands, the ectoderm-mesoderm isolates showed a peculiar
behavior. Very small numbers of red blood cells did form, and they
contained Hb, but no endothelium developed. It was concluded that the
endoderm played: (1) an important role in organization of mesoderm
into blood islands which leads to endothelium formation; and (2) was
a potent stimulus for the number of red blood cells formed and amount
of Hb synthesized. The possibility that material may pass from endoderm
to blood island cells and play a role in the initiation of Hb synthesis
was proposed, and the observations of Mato et al. (1964) are consistent
with this view.
It should be noted here that the results of experiments on Hb formation
may be influenced by the conditions of culture. In general, erythrocyte
formation and Hb synthesis are not as sensitive to poor culture conditions as many other concurrent morphogenetic events, e.g., somite and
heart formation. Almost any available organ culture method in which
the tissue is at a medium-gas interface is adequate, especially blastoderms (or pieces from them) having attained the 1-somite stage. However, the frequency of erythrocyte formation and general health of the
culture is much better in rich media, and for this purpose the whole-egg
agar medium of Spratt is very good (Spratt and Haas, 1960a). A more
marked sensitivity to the type of medium is apparent if pre-streak and
early-streak blastoderms are used, and organ cultures of unincubated
blastoderms are quite fastidious, really only doing well in our experience
on Spratt's whole egg medium.
The time when Hb first appears in the development of the chick
embryo is fairly well settled; a variety of different techniques show it
is first detectable at the 6- or 7-somite stage. The practiced eye can
visualize the faint red color of Hb in the living blastoderm by the 10- to
12-somite stages. While techniques sensitive enough to reveal the presence of the very first molecules of Hb are not available, there is little
question that an accelerated rate of synthesis has commenced by the
7-somite stage. The benzidine stain shows that Hb is present in blood
island cells of the 7-somite blastoderm (Slonimski, 1927; O'Brien, 1960,
1961). Hb is not detected elsewhere at this stage by these techniques.
O'Brien (1960) states that the stainable Hb is first seen in the cell
nucleus, and subsequently a halo of Hb surrounds the nucleus; soon
thereafter the entire cell stains for Hb. It is of real interest to determine
if Hb synthesis is first initiated in the nucleus. Hammel and Bessman
(1964) have provided evidence that the isolated nucleus of duck reticulocytes may synthesize some Hb (cf. Mirsky and Allfrey, 1958). Efforts
to detect a concentration of protein-bound heme by autoradiographic