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11.25 14 LE
MICROCOPY RESOLUTION TEST CHART
NATIONAL BUREAU OF STANDARDS - 1963
- ORNLP-2812
ADO 10.566
MASTER
1.
. .00; 21:50
Cowls 660634-1
AN IN VITRO BONE MARROW SYSTEM FOR STUDYING
RECOVERY FROM RADIATION INJURY
L. H. SMITH
(Biology Division, Oak Riäge National Laboratory, Oak Ridge, Tennessee)

LEGAL NOTICE
RELEASED FOR ANNOUNCEMENT
IN NUCLEAR SCIENCE ABSTRACTS
This report no prepared us us uccount of Guverament sponsored work. Neither the United
Sulos, dor the Commission, oor way person acting oa beball of the Commission:
A. Makes any warranty or representati ja, expressed or implied, with respect to the accu-
racy, completeness, or usefulness of be informazon contained in the report, or that the use
of away iaformation, apparitus, metbod, or process disclosed la is report may not latriage
prinatoly owned rights; or
B. Asrumes any llabillues with respect to the use of, or for damages resulting frota the
use of any information, appundus, melbod, or process disclosed in a report.
As used in the above, "person acting on betw of the Commission" includes any em-
ployee or coatractor of the Commission, or employee of such contractor, to the extent that
soch coaployee or contructor o! the Commission, or employee of such contractor prepares,
disseminates, or provides access to any information pursuaat to his era ploymeal or contract
with the Commission, or his employment with such contractor.
Research sponsored by the U. 9. Atomic Energy Commission under
contract with the Union Carbide Corporation
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Recovery froni radiation injury at the cell level has been demonstrated
for bacteria, higher plants, protozcans and, more recently with altered
mammalian cell lines in culture. We have attempted to apply some of these
recovery measures, which have been successful for the aforementioned forms,
to normal mouse bone marrow cells in vitro.
These cells were chosen for
study because they constitute a critically radiation sensitive tissue in
so far as survival of the animal is concerned and because their reproductive
capacity can be readily assessed and quantified by transplantation into
syngeneic irradiated recipients. For this assessment we have used the
spleen "Fe method, a diagram of which is shown in figure 1. With this
method it was found that a linear relation exists between spleen "Fe up-
take and marrow cell dose over the range 0.05 to 0.8 x 10° cells. By
comparing this relation for unirradiated cells with that for irradiated
cells, we were able to construct a survival curve for in vitro radio-
sensitivity of the erythropoietic compartment of bone marrow. The
exponential survival curve indicated at Day of about 75 R and an exiira-
poration to zero X-ray dose at about 1.6.
To study effects of environment on irradiated marrow cells in vitro
with the purpose of effecting recovery, requires definition and character-
ization of normal cell maintenance over a short period of time. Many
culture variables were tested, and the following conditions were found
to be optimal for normal unirradiated cells: TC 199 plus 25% mammalian
serum; 5.0 to 50.0 x 106 cells/ml; pH 6.8 - 7.6; and stationary cultures.
Under these conditions, the major variable was temperature. At 37, 25 or
2°c, suspensions lost about 25, 10 or 5% erythropoietic activity after
1-
w
6 hours incubation. Because work of other investigators has shown that
for some organisms survival is related to temperature during or after
irradiation, we tested these parameters in the mouse marrow system. At
irradiation temperatures of 2, 25, or 37", respective Dazı, and extra-
polation numbers were essentially the same suggesting that radiosensitivity
of marrow cells is not a function of temperature during exposure. On the
other hand, activity of irradiated cells during 6 hour incubation at 37°
declined at a much greater rate than those incubated at 25 or 2°; and
there was no indication that recovery occurred at incubation temperatures
ranging from 2 to 37° (tested at 2° to 3º intervals). These data not only
indicate that recovery cannot be enhanced by manipulating the temperature
but that expression of X-ray injury may be a temperature dependent event.
The efficacy of tissue extracts including heterologous DNA and RNA
have been tested in the mouse marrow system. Although these series of
experiments are incomplete, we have not found a substance which, when present
in the postradiation environment for up to 24 hours, effects recovery froin
X-ray injury. Current studies comparing single vs divided exposures indicate,
however, that recovery of mouse marrow cells from the first increment of
the divided exposure can be demonstrated in vitro.
The advantage of the in vitro mouse bone marrow system is two-fold.
First projiferative activity or capacity of normal cells is measured in
vivo; and secondly effects can be measured in an isolated in vitro system.
The major disadvantage is the difficulty of maintaining functional capacity
of cells for more than a few hours, especially at 37°c.
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LEGEND FOR FIGURES
FIG. 1. - Spleen "Fe method for measuring erythropoietic activity
nf mouse bone marrow cells.
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END
DATE FILMED
9 / 14/ 66


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