568
ETIENNE WOLFF
3. Protein Factors for Differentiation in vitro
Fraser (1957) used the criterion of the number of differentiated
somites to test and compare the nutritive value of various natural
and artificial media. Blastoderms were cultivated from the primitivestreak stage for 22-23 h. The nutritive substances were added to the
basal medium, which consisted of agar gel in Ringer's solution.
This work tended to demonstrate that proteins or simple peptides,
as well as glucose, are indispensable for the differentiation of somites.
O f all the substances tested, whole egg-white gave rise to the best
differentiation (an average of 5*6 somites). Table I shows the results obtained by Fraser with different substances or extracts. The dialysate
of egg-white has an effect only slightly less intense than that of whole
egg-white, whereas neither the remainder after dialysis nor crystallized
ovalbumin had any effect at all. This tends to prove that small molecules are more active in this process than big ones. When treated by
different processes (boiling, aeration, treatment by alkalis) the dialysate
loses its properties, either partially or totally.
The free amino acids of egg-white, are moderately favourable to the
differentiation of somites (average of 1-9). O f them, glutamic acid
is inactive and only alanine associated with glucose has a real effect
which is reinforced by the addition of hydrogen sulphide.
The author has concluded that several factors are effective in the
activity of whole egg-white: glucose; alanine which is perhaps transformed into cysteine by contact with the hydrogen sulphide; another
compound of low molecular weight which is inactivated by boiling,
aeration, and treatment with ether or acetone; and finally a fraction
belonging to a complex of large molecules from the egg-white and
perhaps incorporating a sulphydryl group.
In further work, Fraser (1960b) has shown that cysteine does in fact
have a noteworthy effect on the development of somites (5-6 and 6-4
with 5 mg°/o and 10 mg%, respectively), and on the development of
the whole embryo. Other sulphydryl compounds have a feeble activity
(thiamine, homo-cysteine, and coenzyme A ) ; still others have no
activity at all (glutathione and methionine). For this reason, it is not
likely that the effects of cysteine can be linked to its reducing power.
In fact, vitamin B 12 which is considered to be capable of maintaining
SH groups in an active state, has no influence on the differentiation
of somites.
Chloride of mercury blocks SH groups by forming mercaptides with
them. It also has an inhibitory effect on the formation of somites. But
if cysteine is added to the medium after this treatment, providing that
the dose and duration of treatment have not been too great, the
ETIENNE WOLFF
3. Protein Factors for Differentiation in vitro
Fraser (1957) used the criterion of the number of differentiated
somites to test and compare the nutritive value of various natural
and artificial media. Blastoderms were cultivated from the primitivestreak stage for 22-23 h. The nutritive substances were added to the
basal medium, which consisted of agar gel in Ringer's solution.
This work tended to demonstrate that proteins or simple peptides,
as well as glucose, are indispensable for the differentiation of somites.
O f all the substances tested, whole egg-white gave rise to the best
differentiation (an average of 5*6 somites). Table I shows the results obtained by Fraser with different substances or extracts. The dialysate
of egg-white has an effect only slightly less intense than that of whole
egg-white, whereas neither the remainder after dialysis nor crystallized
ovalbumin had any effect at all. This tends to prove that small molecules are more active in this process than big ones. When treated by
different processes (boiling, aeration, treatment by alkalis) the dialysate
loses its properties, either partially or totally.
The free amino acids of egg-white, are moderately favourable to the
differentiation of somites (average of 1-9). O f them, glutamic acid
is inactive and only alanine associated with glucose has a real effect
which is reinforced by the addition of hydrogen sulphide.
The author has concluded that several factors are effective in the
activity of whole egg-white: glucose; alanine which is perhaps transformed into cysteine by contact with the hydrogen sulphide; another
compound of low molecular weight which is inactivated by boiling,
aeration, and treatment with ether or acetone; and finally a fraction
belonging to a complex of large molecules from the egg-white and
perhaps incorporating a sulphydryl group.
In further work, Fraser (1960b) has shown that cysteine does in fact
have a noteworthy effect on the development of somites (5-6 and 6-4
with 5 mg°/o and 10 mg%, respectively), and on the development of
the whole embryo. Other sulphydryl compounds have a feeble activity
(thiamine, homo-cysteine, and coenzyme A ) ; still others have no
activity at all (glutathione and methionine). For this reason, it is not
likely that the effects of cysteine can be linked to its reducing power.
In fact, vitamin B 12 which is considered to be capable of maintaining
SH groups in an active state, has no influence on the differentiation
of somites.
Chloride of mercury blocks SH groups by forming mercaptides with
them. It also has an inhibitory effect on the formation of somites. But
if cysteine is added to the medium after this treatment, providing that
the dose and duration of treatment have not been too great, the