[CANCER RESEARCH 40, 4467-4472,
0008-5472/80/0040-OOOOS02.00
December 1980]
Duration of Cell Cycle and Its Phases Measured in Synchronized Cells
of Squamous Cell Carcinoma of Rat Trachea1
Ronald E. Gordon and Bernard P. Lane
Department of Pathology. State University of New York at Stony Brook. Stony Brook. New York 11794
ABSTRACT
Synchronization of tumor cells can facilitate measurement of
cycle time and permit identification of subpopulations which
deviate from the average in either total cycle time or duration
of specific phases. We utilized exposure to high concentrations
of thymidine to arrest cycling cells in tumor fragments in
culture. Pieces of six squamous cell carcinomas, induced in
heterotopically
transplanted rat tracheas by exposure to
benzo(a)pyrene, were placed in culture and subjected to two
sequential thymidine blocks. The labeling indices in cultures
studied at 2 and 8 hr after release from the second block were
equal to the growth fractions. By 16 hr after release from the
block, no DMA synthesis was observed in any culture. In three
tumors for which cycle and cycle phase duration was meas
ured, mitosis occurred synchronously 12 hr after release from
the thymidine block, and a second period of DMA synthesis
began 16 hr later. 7"cwas 28 hr, Ts averaged 9 hr, TG,averaged
15 hr, and TG2averaged 3 hr, allowing 1 hr for TM. These
numbers correspond to the values for normal cells participating
in wound healing in this epithelium. There were also no deviant
populations present. The cycle and its phases therefore cannot
serve as markers for cancer in this tissue, but the degree of
synchrony achieved will permit correlative studies of phasespecific cell properties which have been suggested as candi
dates for markers of cancer.
INTRODUCTION
Many morphological and biochemical characteristics of nor
mal cells vary with position in the cell cycle. Differences in
types and rates of synthesis of structural proteins, RNA, DMA,
and enzymes as well as variations in the transport of nutrients
during the generative cycle have been reviewed by Baserga
(1-3). It has also been shown that changes in cell surface
structure and in the quality and quantity of membrane mole
cules occur throughout the cell cycle in normal cells (16).
Differences in some of these characteristics have been pro
posed as markers for distinguishing neoplastic cells from the
normal population (1, 3, 7, 9, 11, 18, 21). Since the markers
may themselves vary with the cell cycle, to evaluate these
possibilities one must know in which phase of the generative
cycle the cells that are being studied reside. This correlation
may be achieved by simultaneously determining the phase of
the cycle while examining other features. Alternatively, mitotic
activity can be synchronized so that cell cycle phases can be
predicted. This approach is preferable because concurrent
assessment of cell cycle phase which might impede measures
of cell function or structure is then no longer required.
We describe a method for synchronizing mitotic activity in
tumor cells and present data concerning duration of the cell
cycle and its constituent phases.
MATERIALS
AND METHODS
Experimental Design
Fragments of tumor were placed in culture and treated with
dThd2 to arrest cells moving through the cell cycle (8, 24). The
success of the synchronization method was tested by autoradiographic assessment of DNA synthesis over a period of hours
after removal of the dThd block to determine whether the
labeling index equaled the growth fraction and whether all cells
which synthesize DNA start at the same time shortly after
release from the block. The independent effects of the organ
culture manipulations were monitored by measuring growth
fractions, and those of circadian rhythms were measured by
comparing mitotic activity and labeling after release at different
hours. After the degree of synchrony was determined, the cell
cycle and individual phase durations were measured for pos
sible subpopulation deviations and comparisons with previ
ously determined phase durations in normal regenerative tra
chéalepithelium (10).
Source of Tumor Tissues
The study used 6 squamous cell carcinomas induced in
heterotopic trachéalgrafts treated with benzo(a)pyrene (14).
Female Fischer rats (Charles River Breeding Laboratories, Inc.,
Wilmington, Mass.) weighing 100 to 150 g were used as tissue
donors and recipients for the graft procedure. Donor tracheas
were placed in s.c. pouches on the backs of hosts. Two weeks
later, agar suspensions containing 2.5 mg of benzo(a)pyrene
were placed in the lumens of the grafts. Neoplasms which grew
to measure 15 to 20 mm in greatest dimension arose in the
grafts from 3 to 6 months after onset of exposure to
benzo(a)pyrene. The tumors exhibited central necrosis with a
viable 3- to 5-mm rim composed of micronodules with interven
ing vascular connective tissue.
Culture and Synchronization of Tumor Fragments
With the rats lightly anesthetized with ether, fragments of
viable tumor tissue were removed from the well-vascularized
periphery of the cancers and minced. These pieces were
immediately immersed in serum-free McCoy's modified Me
dium 5A with streptomycin and penicillin where they remained
for 1 hr. The cells composing the tumor pieces were then
synchronized by the use of a double dThd block which arrests
cells at the Gi-S interface (8, 24). The McCoy's modified
' Supported by Contract NO 1 CP 33361. National Cancer Institute.
Received January 2, 1979; accepted August 26, 1980.
DECEMBER
2 The abbreviation
used is: dThd. thymidine.
1980
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4467
R. E. Gordon and B. P. Lane
Medium 5A was replaced with the same medium containing 2
HIMdThd. After 30 hr, fresh McCoy's modified Medium 5A was
substituted for the dThd-containing
which had collected at the interface
the medium containing 2 mw dThd
the McCoy's medium without dThd,
medium to release cells
of Gìand S. Six hr later,
was again substituted for
and the tissue was incu
bated for 30 hr before return to medium without dThd. The
second block serves to increase the degree of synchrony.
background which was never more than 2 grains per cell.
However, in the area used for scoring, there were no cells with
grain counts in the range of 2 to 10 grains. For several sections
in which absolute grain counts were done, the counts were
unimodal and fell within 1 S.D. from the mean.
All slides were read by a single observer in a double-blind
protocol, and 5% of the slides were selected at random for
repeat scoring by that observer and by another observer as a
measure of reproducibility and absence of bias.
Effects of Culture and Circadian Rhythms on Labeling
Labeling indices were measured without dThd block at 6
a.m., 12 noon, 6 p.m., and 12 midnight on the first day to test
for circadian rhythm. Labeling indices were repeated on the
second and third days to determine whether the size of the
normal in situ proliferative population changed due to culture
conditions. The labeling was performed by placing the tissue
for 30 min in fresh McCoy's medium to which [3H]dThd (specific
activity, 20 |uCi/mmol) was added to make a final concentration
of 1 juCi/ml of medium. The tissue was then cold chased with
two 10-min washes in McCoy's medium containing excess
dThd (1.2 jug/ml) in order to displace any unincorporated
[3H]dThd. Labeling indices of arrested organ cultures were also
measured at 2, 8, and 16 hr after release from the second
dThd block to determine if synchrony was achieved. Unsynchronized tumor tissue was pulse labeled at each medium
change in the above protocol to determine if the medium
changes in themselves had an effect on the labeling indices.
Growth fractions were measured on the first, second, and
third days of culture by replacing the medium with fresh
McCoy's medium containing [3H]dThd (0.5 juCi/ml; specific
activity, 20 Ci/mmol). The tissue was incubated in this medium
for 24 hr. After this interval, the tissue was cold chased with
two 10-min washes of medium containing excess dThd (1.2
,ug/ml).
One additional tumor-bearing animal was pulse labeled in
vivo with an i.p. injection of [3H]dThd at a concentration of 1
fiCi/g body weight to compare in vivo versus in vitro labeling
indices. The animal was sacrificed 1 hr after pulse labeling,
and tumor fragments were prepared for autoradiography.
Autoradiographic Procedure
After labeling, the tissue was immediately immersed in fixa
tive containing 3% glutaraldehyde with 0.2 M sodium cacodylate buffered at pH 7.3. After overnight fixation, the tissue was
transferred to 1% unbuffered osmium tetroxide. One hr later,
the tissue was dehydrated in graded steps of ethanol and
embedded in Epon 812. One-ftm-thick plastic sections were
heat fixed to acid-cleaned glass slides and dipped in 1:1
dilution of NTB2 photographic emulsion. The slides were kept
under cool dry conditions in light-tight boxes for 4 weeks,
developed in full-strength Kodak developer D19, and fixed with
full-strength Kodak Rapid Fix. After drying, the slides were
stained with méthylèneblue and azure II.
An 80-/im-wide peripheral rim of each tumor fragment section
was scored for percentage of cells labeled. A minimum of 2000
cells was counted for each tumor at each time studied. Only
nucleated cells were counted, and cells showing pyknosis or
karyolysis were excluded. Cells were scored as positive for
[3H]dThd incorporation if 3 or more silver grains were present
directly over the nucleus. This threshold value was greater than
4468
Cell Cycle and Phase Durations
Cell cycle and phase durations were measured by the tech
niques described by Gordon and Lane (10).
Determination of Duration of DMA Synthesis ( Ts). Two hr
after release from the synchronization procedure, the tissue
was exposed to [3H]dThd at a concentration of 5 /nCi/ml
McCoy's medium (specific activity, 60 fiCi/mmol) for 30 min
followed by three 10-min cold chases with McCoy's medium
containing excess dThd (1.2 jug/ml). The tissue was transferred
to McCoy's medium, and, at hourly intervals between 6 and 12
hr after the [3H]dThd pulse, groups of fragments were labeled
for 30 min with medium containing [14C]dThd at a concentration
of 1 /iCi/ml medium (specific activity, 42 mCi/mmol). The
fragments were cold chased in excess dThd, immediately fixed
in 3% glutaraldehyde with 0.2 M sodium cacodylate buffered
at pH 7.3, and prepared for double-label autoradiography. Data
obtained from slides of 3 tumors raised on separate animals
were statistically compared to one another for each time stud
ied by standard error of the mean. The initial label was given 2
hr after release because preliminary studies on a similar tumor
in which replicate fragments were pulse labeled at 0.5-hr
intervals from 20 min to 4 hr after removal from dThd excess
showed maximal labeling at 2 hr thereafter, and there was less
than 0.5% labeling at 1.5 hr. As an independent test of the
dThd effect on cycle progression, unsynchronized fragments
from one tumor were also pulse labeled with [3H]dThd and then
pulse labeled with [14C]dThd from 6 to 12 hr later. The disap
pearance of double-labeled cells was at 9 hr. Thus, flux through
S of unsynchronized cells equaled progress through S by
synchronized cells. Another set of unsynchronized specimens
was pulse labeled with [3H]dThd and then subjected to a pulse
of [14C]dThd at hourly intervals from 12 to 32 hr later. The
earliest appearance of double label is a measure of G, + G2
+ M, and the last appearance of double-labeled cells is a
measure of G, + G2 + M + 2S. These values were used as
controls for the effects of dThd on cycling.
Technique for Determining Duration of the Cell Cycle ( Tc).
Two hr after release from synchronization, the fragments taken
from 3 separate tumor-bearing animals were pulse labeled with
[3H]dThd at a concentration of 5 /¿Ci/mlin McCoy's medium
for 30 min, cold chased with excessive dThd, and then placed
in nonradioactive McCoy's medium. Beginning 14 hr after the
[3H]dThd exposure at 1-hr intervals through 40 hr, groups of
fragments were pulse labeled with ['"CJdThd for 30 min, cold
chased with excess dThd, and immediately fixed with glutar
aldehyde. One-^m sections of Epon-embedded tissue from
alternate hours were prepared for double-label autoradiogra
phy to determine the shape of the curve. Specimens from the
remaining hours which fell within the periods of DMA synthesis
were studied.
CANCER
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Cytokinetics
Double-label
autoradiograms
were prepared
of Culture Synchronized
Table 1
by the tech
nique described by Gordon and Lane (10). A first layer of
Kodak NTB2 photographic emulsion was applied by dip tech
nique to the 1-|um sections at 1:3 dilution. This layer was
exposed for 9 days, developed, and counted for the number of
labeled cells by phase microscopy. A second layer of Kodak
NTB2 at 1:1 dilution was applied by dip technique over the first
but separated from the first by a thin gelatin layer (0.8%). The
second layer was exposed for 17 days, developed, and stained
with méthylèneblue and azure II. The technique for reading
the double-labeled autoradiographs was fully described by
Gordon and Lane (10).
Technique for Determining Duration of G¡( TG,).Two hr after
release from synchronization,
fragments were exposed to
[3H]dThd at a concentration of 5 juCi/ml McCoy's medium for
30 min. The fragments were cold chased with excess dThd and
then placed in normal McCoy's medium. Groups of fragments
were fixed with glutaraldehyde at hourly intervals between 10
and 14 hr after the [3H]dThd pulse label. They were embedded
in Epon 812, sectioned, and prepared for single-label autoradiography. A minimum of 100 mitotic figures in at least 10
fragments per time point per tumor were scored for the per
centage of mitotic cells exhibiting silver grains over their chro
mosomes.
Calculation of the Duration of G2 (T,,.) and Approximation
of (TM). TG, was calculated by subtracting the Ts + TQ?+ TM
from the 7"c.TMwas approximated and supported by measure
ments made from other rat tissues (4).
RESULTS
Rat Trachea/ Tumor
Daily labeling indices and growth fractions ot culture tumor fragments
Tumor fragments (rom 3 carcinomas were pulse labeled in vitro with [ 'H|dThd
(5 jiCi/ml medium) for 30 mm at 3 different times of day to determine if tumor cell
proliferation exhibits a circadian rhythm. Pulse labels were done on 2 successive
days to determine whether circadian rhythms appeared after a period in culture.
Additional tissue samples were pool labeled with [ 'H]dThd (0.5 //Ci/ml medium)
for 24 hr on Days 1.2. and 3 to determine if the size of the proliferative population
changes over the duration of culture. The specimens, pulse labeled, fixed,
embedded, and sectioned, were prepared for autoradiography and represent the
labeling index Pool-labeled specimens prepared similarly represent growth
fractions
Tumor222222333333Time1
index0.21702100.2030.1900.2060.2110.2020.1960
fraction0.5900.5910.5980.6080.
2oon6
n
p.m.12
midnight6
a.m.12
noon12
noon1
n2oon6
p.m.12
midnight6
a.m.12
noon1
noon1
2
n2oon6
p.m.12
midnight6
a.m.1
noon1
2
2 noonLabeling
2140.2160.2010.2100.2050.2130.1930.1980.2060.203Day1111
5860.595
Table 2
Labeling indices for determining effectiveness of double dThd block on
synchronization of tumor cells
Tumor fragments were cultured in medium containing 2 mM dThd for 30-hr
periods, returned to normal medium for 6-hr intervals, and then exposed for 30
hr more to 2 mM dThd. To measure the extent of cell cycle synchronizing to
tumor, samples were pulse labeled with [ 'HJdThd (5 fiCi/ml medium) at 2, 8, and
16 hr after release from the second dThd block.
Cultured fragments of squamous cell carcinoma retained
their histological organization and cellular differentiation when
cultured. There were no avascular or necrotic fragments. La
beling indices of unsynchronized
tissues were the same
throughout the 3-day study and did not demonstrate any ap
preciable circadian rhythm (Table 1). Comparison of in vivo
and in vitro labeling indices for one of the tumors revealed no
differences. There was also very little difference in labeling
index among the 6 tumors studied. Growth fractions were
measured to determine whether there was any change in the
size of the proliferative population during the study. These 24hr pool labels showed no significant differences during the 3day period (Table 1). There was no variation of labeling indices
associated with changes of culture medium. Grain counts in
the areas studied in specimens which had been labeled in vitro
were 10 grains or more per nucleus with background less than
2 grains.
In cell populations not dividing synchronously, the labeling
index is usually a fraction of the proliferative population, and,
if the cells are completely asynchronous, the percentage of
cycling cells which are labeled during a short exposure to,
[3H]dThd is an accurate reflection of the relationship between
ments on the third day of culture. Eight hr after release, the
labeling indices of all the tumors were still equal to the growth
fractions of tumor fragments on their third day of culture (Table
2). By 16 hr after release, the labeling index declined to zero
(Table 2). These data show that the double dThd block was
suitable for synchronization of the proliferative population of
tumor cells.
Application of the double label method permits direct meas
ure of the duration of S and determination of the degree of
synchrony which was achieved by the double dThd arrest. The
specimens subjected to a pulse label of [3H]dThd 2 hr after
release from the dThd arrest and to a pulse label of [14C]dThd
the duration of S and the duration of the cell cycle (19). After
synchronization, pulse label of the cells in the S phase should
produce a labeling index equal to the growth fraction. As the
cells leave the S phase, the labeling index should decline
rapidly to zero.
Two hr after the release from the second dThd block (Table
2), the labeling index in synchronized tumor fragments equaled
the measured growth fraction of unsynchronized tumor frag-
at hourly intervals thereafter (Chart 1) show a sharp decline in
double-labeled cells between 8 and 9 hr after introduction to
the [3H]dThd. This indicates that the duration of S is 8 to 9 hr
in a well-synchronized population of tumor cells. The unsyn
chronized tumor cells in fragments not exposed to the dThd
block exhibited double-labeled cells until the second pulse was
given at 9 hr. This is evidence that the dThd block did not
increase the length of the S phase.
Labeling index
Tumor1234562
hr0.5840.5760.5910.5960.5890.6038hr0.5970.5630.58916hr0.0040
0010003
DECEMBER 1980
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4469
R. E. Gordon and B. P. Lane
Labeling the synchronized cells as they enter the S phase
and then observing them at a later time autoradiographically
as labeled mitosis occurs (Table 3) permit accurate measure
ment of the interval from the onset of S to mitosis. This period
which includes 7S, TG?,and Vz TMwas determined to be 12 hr.
Since TSand V?Tu are 8.5 to 9.5 hr, 7G?is, by subtraction, 2.5
to 3.5 hr.
The duration of the cell cycle (7~c)was measured by labeling
the synchronized population of cells with [3H]dThd as they
80-,
U)
70-
o
60-
50-<_j40-U.
UJ
entered S and then, at hourly intervals after the wave of mitosis,
pulse labeling the fragments with ["*C]dThd to determine the
onset of the second phase of S for cells which had participated
in the first S phase (Chart 2). The 28-hr interval between these
points was considered to be the Tc.
TGI was indirectly calculated to be 14.5 to 16.5 by subtract
ing TS, rGj, and the approximation of TMfrom 7"c.Measures of
S and overall cycle time of unsynchronized cells gave similar
values for the random sample of cells which were pulse labeled.
\\^Z
*H*
ONLY•
/4C
O30-O^
»or//[3h]dThd
20-LÜ
PULSE
U5
io-*o.0-•
IIi
16
80-
20
]
24
HOUR Op[l4c]
28
32
dThd
36
4O
PULSE
Chart 2. The percentage of cells in synchronized cultured tumor fragments
labeled with 3H only. MC only, and both on double-label autoradiographs. The
data demonstrate the synchronous departure of cells from S phase between 8.5
and 9.5 hr after its onset. Tumors pulse labeled with |3H)dThd 2 hr after release
from the synchronizing block were pulse labeled at intervals thereafter. The
tissues were immediately fixed, embedded, sectioned, and prepared for doublelabel autoradiography. Each time point is the mean from the comparison of 3
tumors raised in separate rats. The autoradiographic results were subjected to
statistical analysis, and no point exceeded ±1 S.E.
DISCUSSION
02
8
10
HOUR OF [l4C]dThd
12
14
PULSE
Chart 1. The percentage of cells in synchronized cultured tumor fragments
labeled with 'H only, "*C only, and both on double-label autoradiographs after
being exposed to pHjdThd 2 hr after release from synchrony and pulse labeled
at later times with (""C]dThd The tissues were immediately fixed, embedded,
sectioned, and prepared for double-label autoradiography. Each rime point is the
mean from the comparison of 3 tumors raised on separate rats. The autoradiographic results were subjected to statistical analysis, and no poinf exceeded
±1S.E.
Table 3
Percentage ol labeled mitotic cells observed at intervals after DNA synthesis
Tumor fragments were synchronized by double dThd block and pulse labeled
with [ 'HjdThd 2 hr after release from the block. At intervals thereafter, specimens
were fixed, embedded, sectioned, and prepared for autoradiography. The meas
ured period from the time of ( 'H]dThd pulse label to the peak incidence of labeled
mitosis is equal to 7S + Ta, + Vz Tu.
Time after [3H|dThd
pulse label
(hr)1011121314%
4470
mitosisTumor
of labeled
4029530Tumor
5039830Tumor
6029340
The ability to culture tumor fragments the in vivo structure
and function of which are maintained (13, 20, 26) may prove
helpful in the study of some characteristics likey to be affected
by uncontrolled extraneous variables. One such variable is
duration of the individual phases of the cell cycle. In most
studies of biochemical function or structural features, it is not
possible to simultaneously recognize the cycle phase. If the
cells were pulse labeled with radioactive dThd, one can ascer
tain when some of the cells were in the S phase of the cycle.
However, the phase cannot be determined by many cells which
were not labeled during the pulse. A way in which one can be
certain of the phase of all cells is to synchronize their entry
into a particular phase of the cell cycle.
Many attempts have been made to induce synchrony of
various cells in culture and tissue in situ. Methods which use
physical agents, such as heat, cold, mechanical trauma, or
surgical excision, and chemical agents, such as isoproterenol,
actinomycin D, puromycin, bromodeoxyuridine,
1-/8-D-arabinofuranosylcytosine,
and the stathmokinetic drugs, can also
be used to induce synchrony (15). The problem associated
with physical agents is that the resultant synchrony is not as
complete as with chemical means. However, most of the chem
icals used are toxic and have their own independent effects.
dThd is an exception in that it is normally available to and used
by the cells, but, when given in excess, reversibly arrests some
mammalian cell types at the Gt-S interface without any toxic
effects (8, 15, 24). While dThd has been shown to be toxic in
some cell systems (4) and to be more toxic for tumor cells than
normal cells (25), in our experiments the histology and growth
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Cytokinetics
fraction of tumor fragments treated with dThd were no different
than those which had not been so treated.
Labeling of tumor fragments by incubation in medium con
taining [3H]dThd is known to result in variable amounts of
incorporation which is in part attributable to limited penetration
(12) and may also be affected by oxygen tension (6). It has
been shown that scoring only the outermost 100-/im perimeter
of the labeled fragment avoids counting as negative those
cycling cells which have not had access to label or with
incorporation that is depressed by low oxygen tension. By
using high concentrations of [3H]dThd and restricting scoring
to the peripheral zones of high labeling, we apparently ex
cluded from our sample cells with supranuclear grain counts
near background. The data also reveal that, while it has been
shown biochemically that in some systems there is a low level
of DMA synthesis throughout the cycle (22) and that even
within S phase there are periods of varying amounts of DNA
synthesis, such features may not be demonstrated by autoradiographic methods.
The labeling indices under these conditions were similar to
those in the control in which the tissue was labeled in vivo,
indicating that the level of radioactivity used in vitro did not
produce necrosis or inhibit cycling. Differences were also not
seen between the fragments subjected to the dThd block and
those not synchronized by this technique. This is empirical
evidence that, in this tumor cell system, the toxic and cytokinetic effects of high doses of dThd reported for other models
under other conditions (4) do not occur.
There has been concern about whether cells treated with
excess dThd are arrested at the Gt-S interface or are just
slowed substantially in their movement through the S phase
(15). We conclude that this tumor population was arrested at
the G,-S interface because there was a plateau between 2 and
8 hr and a sharp decline of the labeling index by 16 hr after
release from synchronization. In addition, the study of Ts further
substantiates the good degree of synchrony attained by the
double dThd arrest. If cells had only been slowed in their
movement through S phase as a result of the dThd block, the
cells would have continued to be distributed throughout S
phase, and, after release, there would be a gradual decline of
labeling indices with time rather than a sharp drop observed.
Tumors have been known to vary with regard to cell cycle
kinetics from the normal tissue from which they arose (1, 3, 5,
23). The durations of cell cycle phases might themselves be
characteristics useful in recognizing tumor cells. Heterogeneity
with regard to the generative cycle could be an indication of
instability of the tumor population while consistent deviations
of subpopulations might be indicative of clonal origins of tu
mors. Previous studies in which variations have been detected
involved the averaging technique of Quastler and Sherman (19)
or modifications thereof. This method does not reveal whether
the observed differences from normal tissue are limited to a
specific cellular subpopulation or are true for the entire tumor.
In tumors where variations in cycle time might exist, only gross
changes can be observed because small groups of cells have
minor effects on the populational averages. Synchrony of tumor
populations allows analysis of cell cycle kinetics while avoiding
the averaging methods required with randomly distributed pop
ulations.
We measured the duration of the phases of the cell cycle
looking for differences from normal in a specific phase for the
of Culture Synchronized
Rat Trachea! Tumor
entire tumor cell population or for deviant clones. 7S of 8 to 9
hr, TG?of 2.5 to 3.5 hr, TG, of 14.5 to 16.5 hr, and Tc of 28 hr
were determined for the tumor population. The phase durations
we have found for the tumor cells are the same as those of the
proliferative cells in healing mild wounds of the trachéalepithe
lium (10). We have therefore concluded that cell cycle kinetics
is not one of the characteristics which will distinguish these
tumor cells from normal cells. On the other hand, this similarity
between cancer cells and normal cells will facilitate study of
cycle-specific features which may be useful markers of cancer,
since the homogeneity of the tumor cell population increases
the predictability that, once synchronized, all cycling cells are
in the same phase and comparisons can be made between
normal and tumor cells without corrections for differences in
cycle time or phase durations in the 2 populations.
ACKNOWLEDGMENTS
We are grateful for the technical assistance of Chantal Weinhold in the tumor
induction aspects of this work.
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RESEARCH
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VOL. 40
Duration of Cell Cycle and Its Phases Measured in Synchronized
Cells of Squamous Cell Carcinoma of Rat Trachea
Ronald E. Gordon and Bernard P. Lane
Cancer Res 1980;40:4467-4472.
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