13. D I F F E R E N T I A T I O N
477
resuspended, pelleted and organ-cultured in the same fashion as the
freshly liberated chondrocytes. Cells after 10 days in dispersed culture
failed on organ culture to form extracellular metachromatic material,
or to show other histological signs of chondrogenesis. Incorporation of
3 5
S was reduced after 5 days in culture to approximately half that of
freshly liberated chondrocytes and reached minimal levels by 11 days.
Cells beyond the 10-day point when cultured in the absence of embryo
juice or in the presence of embryo juice from successively older embryos
—procedures reported in earlier literature to favor differentiation—
were equally unable to initiate chondrogenesis.
Hilfer (1962) examined the specialized behavior of thyroid cells from
16-day chick embryos. After dispersal with trypsin and versene such
cells would reaggregate and, under organ-culture conditions, reform
follicles containing colloid. Chromatographic analysis showed a normal
pattern of iodinated precursors and hormones. If the cells were dispersed on glass—and maintained with little or no proliferation—they
formed sheets, showed limited morphological organization, and rapidly
lost demonstrable thyroxin. Intracellular glycoprotein and mono- and
diiodotyrosine both persisted over a number of days. Thyroxin and
follicle formation did not occur when the cells were reaggregated and
organ-cultured, with or without thyrotropic hormone. Recombination
of the cells with thyroid fibroblasts resulted in appearance of some
follicles with extracellular colloid, but did not lead to demonstrable
thyroxin. Hilfer believes that under spreading conditions certain
differentiated cellular properties may persist while others are lost.
These results of careful studies using sensitive biochemical criteria are
in accord with a number of earlier reports indicating that differentiated
behavior declines with culturing (Grobstein, 1959a; see also, Billen,
1959; Billen and Debrunner, 1960). The apparent difference between
these reports and those of Waltz, Tullner, Evans, Hertz and Earle (1954),
Schindler, Day and Fischer (1959),, Thompson, Vincent, Jensen, Price
and Shapiro (1959),, and Konigsberg (1961) may rest on differences in
culture procedure (e.g. presence and absence of plasma) or may indicate
differences in the stability of various specialized behaviors. In any
event, it is clear that further critical studies of particular differentiations, employing multiple criteria and examining relations to propagation and culture environment, are prerequisite to any generalization
about this high-priority problem.
V I . H E T E R O T Y P I C F A C T O R S
The possibility that differentiation of one tissue in culture might be
477
resuspended, pelleted and organ-cultured in the same fashion as the
freshly liberated chondrocytes. Cells after 10 days in dispersed culture
failed on organ culture to form extracellular metachromatic material,
or to show other histological signs of chondrogenesis. Incorporation of
3 5
S was reduced after 5 days in culture to approximately half that of
freshly liberated chondrocytes and reached minimal levels by 11 days.
Cells beyond the 10-day point when cultured in the absence of embryo
juice or in the presence of embryo juice from successively older embryos
—procedures reported in earlier literature to favor differentiation—
were equally unable to initiate chondrogenesis.
Hilfer (1962) examined the specialized behavior of thyroid cells from
16-day chick embryos. After dispersal with trypsin and versene such
cells would reaggregate and, under organ-culture conditions, reform
follicles containing colloid. Chromatographic analysis showed a normal
pattern of iodinated precursors and hormones. If the cells were dispersed on glass—and maintained with little or no proliferation—they
formed sheets, showed limited morphological organization, and rapidly
lost demonstrable thyroxin. Intracellular glycoprotein and mono- and
diiodotyrosine both persisted over a number of days. Thyroxin and
follicle formation did not occur when the cells were reaggregated and
organ-cultured, with or without thyrotropic hormone. Recombination
of the cells with thyroid fibroblasts resulted in appearance of some
follicles with extracellular colloid, but did not lead to demonstrable
thyroxin. Hilfer believes that under spreading conditions certain
differentiated cellular properties may persist while others are lost.
These results of careful studies using sensitive biochemical criteria are
in accord with a number of earlier reports indicating that differentiated
behavior declines with culturing (Grobstein, 1959a; see also, Billen,
1959; Billen and Debrunner, 1960). The apparent difference between
these reports and those of Waltz, Tullner, Evans, Hertz and Earle (1954),
Schindler, Day and Fischer (1959),, Thompson, Vincent, Jensen, Price
and Shapiro (1959),, and Konigsberg (1961) may rest on differences in
culture procedure (e.g. presence and absence of plasma) or may indicate
differences in the stability of various specialized behaviors. In any
event, it is clear that further critical studies of particular differentiations, employing multiple criteria and examining relations to propagation and culture environment, are prerequisite to any generalization
about this high-priority problem.
V I . H E T E R O T Y P I C F A C T O R S
The possibility that differentiation of one tissue in culture might be