ate ce entaletadeli ct,ia
122
the tissues had sequestered radioproline during the
numbers of osteoblasts increase in rachitie rat
this point, it should be noted first that Tom
periosteum, endosteum, and the endosteal surface
in the area of the proximal tibial metaphysis was
averaged for 30-50 cells to obtain a quantitative
estimate of the utility of the tracer for bone matrix
synthesis (vide infra). With the staming method em-
genic cells of mice within 15-30 min after a injection. Secondly, in concert with results from
les using radioglycine as a tracer for collagen
mation, 1% 12) it is known that osteogenic cell
port a large porportion of their tracer content
cell with an eccentric nucleus, a prominent nucleolus
and a juxtanuclear vacuole (Golgi apparatus) located on or close to a bone surface. The relative
number of osteoblasts and their precursor cells, variously called reticular, mesenchymal, or osteoprogenitor cells, was not estimated in the primary spongiosa,
thesized collagen molecules by 4 hr. However, p:
is also an active metabolite and can be transfo
via glutamic acid into other amino acids whic!
initial 4 hr of the experiment. The number of silver
grains over the cytoplasm of labeled osteoblasts in the
ployed, an osteoblast was defined as a basophilic
although Rohr‘) has indicated that the absolute
found peak uptake of radioproline in the ¢
the surfaces of bone as an integral part of newly
utilized for a number of other compounds, pro!
and mucopolysaccharides. Thus the early grain c«
per se cannot be depended upon to measure onl:
rate of collagen synthesis by labeled osteoblasts.
The reliability of the initial gram counts wa:
sayed independently by measuring the position o
labeled matrical (collagen and mucopolysacchar!
band of silver grains buried within the corte
days after radioproline administration (Figure
The thickness of lamellar bone deposited by
osteoblasts on the growing surfaces of the (endosteal, .periosteal) and transverse epiphy
bone during this interval was calculated by mex
ing the total distance from the leading edge of
continuous band of labeled matrix to the anaton
surfaces. This was done in the center of approxim:
60 adjacent high powerfields (400 x) with an oc
micrometer. This value was divided by 3 so that
rate of appositional bone growth could be expre:
in microns per day, An estimation of the thickne-
the band of labeled matrix was also attempted, fo
lieu of definitive grain count data early after tr:
administration, this measure should reflect accura'
the rate at which the osteogenic cells on these faces were producing newstruetural collagen. T!
two sets of data were also expressed as an Osi
blast Activity Index (QAI), which is defined as
observed thickness of the ?3H-proline label divided
Fic. 95Autoradiographs of the articular surface of the
transverse epiphyseal bone from rats sacrificed at two intervals
of time after a single injection of *H-proline. A. a band of silver
grains at a-b representing collagen newly formed byosteoblasts
(ob!) 4 hr after injection. The evtoplasm of the osteoblasts is
lightly labeled. B, the position of the silver grains over labeled
collagen lamellae 3 days postinjection. Interval a-c, thickness
of lamellar bone formed in 3 davs. Interval b~c, thickness of
the bandof silver grains. Note the trail of silver grains (interval
a-h) due to reutilization of radioproline. Hematoxylin and
eosin. Original magnification 250 X.
the observed apposition rate (microns/microns/da
It is unlikely that these data would be complica
at the light microscope level by any change in
catabolic rate of newly synthesized collagen m«
cules or differential packing of collagen fibers b
into the skeleton.{?- 15) A ratio of 1 would suge
that the osteoblasts formed the labeled bone mat
in exactly 24 hours’ time. Other values would
inversely proportional to the pace at which |
osteogenic cells were performing. It was difficult
apply this type of analysis to trabecular bone, fir
beeause it is less well-oriented than compact bo
and second, because our animals were not sacrific
at narrowtime intervals.