Download The Role of Cell Differentiation as a Determinant

The Role of Cell Differentiation as a Determinant of
Susceptibility to Virus Carcinogenesis*
HENRY S. KAPLAN
(Department of Radiology, Stanford University School of Medicine, Palo Alto, Calif.)
Experimental analyses of the behavior of certain viruses in vitro, such as Dr. Dulbecco has presented to us on this memorable occasion, provide
an exquisitely powerful method with which to
dissect the intimate details of the cell-virus relationship. I t is to be hoped that such in vitro studies
with the polyoma (10, 1~, ~9, 34), the Rous (s
~8), the myeloblastosis (7), and other carcinogenic
viruses will soon yield new and fundamental information concerning the initial steps in the neoplastic transformation.
I t has been one of the fond hopes of the proponents of the "virus theory" of carcinogenesis
that the oncogenic viruses would provide a unifying conceptual framework for understanding the
complex process of carcinogenesis. Yet, despite
the admiration we may feel for the elegance of in
vitro investigations such as have just been described, we are also increasingly aware that the
advances of the past few years have provided us
with new complexities and difficulties. We now
know t h a t the oncogenic viruses are not a homogeneous family of viruses with similar special attributes. On the contrary, they exhibit as much
diversity as other animal viruses. Some propagate
in the cell nucleus (1), others in the cytoplasm (18,
18, $6); some contain D N A (11, 19, 31), others
R N A (4, 6, 9), and those that have been clearly
visualized by electron microscopy look disconcertingly like other animal viruses of comparable
intracellular location and nucleic acid composition
(5). Thus, we may begin to suspect that virus carcinogenesis will not prove to be a single process,
but a diverse set of responses, perhaps differing
characteristically for each group of oncogenic viruses.
* This paper is based on an invited discussion of the first
G. H. A. Clowes Memorial Lecture, entitled "Viral Carcinogenesis," presented by Dr. Renato Dulbecco at the annual
meeting of the American Association for Cancer Research,
Inc., Atlantic City, April 10, 1961.
Previously unpublished work cited herein was supported by
grant C-885~ from the National Cancer Institute, National
Institutes of Health, U.S. Public Health Service.
In the case of the family of viruses which elicit
lymphomas a n d leukemias in the mouse (15, 83),
we do not yet have the advantage of having an
in vitro assay system, since these viruses do not
produce cytopathogenic effects in any mammalian
cells on which they have been inoculated to date,
although they may be recovered from the culture
fluids (14, 17).
I t is therefore necessary to a t t e m p t to learn as
much as possible about their mode of action from
in vivo observations. Such investigations may yield
valuable new insight concerning the factors which
modify or determine cellular susceptibility to
these viruses and thus contribute to the continuing
search for quantitative in vitro methods.
Thymic lymphoid neoplasms in irradiated strain
C57BL mice have been shown to arise by a completely indirect process (~3). Cell-free filtrates and
centrifugates prepared from such neoplasms elicited significant increases in lymphoma incidence
when inoculated at birth into nonirradiated
C57BL mice. These filtrate-induced tumors then
yielded filtrates which exhibited enhanced potency on serial passage in newborn mice (~4).
Similar observations with filtrates from lymphomas in irradiated strain C3H mice were independently reported by Gross (16). I t thus became
apparent t h a t these supposedly radiation-induced
lymphomas were in reality induced by a virus-like
agent, and it seemed likely that irradiation set the
induction sequence in motion by causing some
change in the virus-host relationship.
A few years ago (8), it was noted t h a t the earliest morphological changes associated with the
induction of thymic implant lymphomas in irradiated C57BL mice bore a remarkable resemblance
to arrested maturation in the lymphoid cell differentiation sequence. Accordingly, attention was
focused on the process of differentiation in the
mouse thymus and its possible relation to this
tumor induction mechanism.
Sainte-Marie and Leblond (~0), studying the
rat thymus, traced a complex process of lymphoid
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98s
Cancer Research
cell differentiation which proceeds in a series of
steps from the undifferentiated reticulum or stem
cell through the large lymphocyte (lymphoblast)
and medium-sized lymphocyte stages to an end
stage, the small lymphocyte, which is generally
thought to be incapable of division. Although this
sequence has not been quantitatively studied in
the mouse, the same cell forms have been identified, and the great relative abundance of the small
lymphocytes in the normal adult mouse thymic
cortex makes it not unlikely that a similar pattern
exists in that species.
I t has long been known that bone marrow
shielding or injection protects irradiated C57BL
mice against the development of thymic lymphomas (ee). I t seemed desirable to compare the
thymuses of shielded versus unshielded irradiated
mice of this strain in a search for meaningful morphological antecedents of lymphoma induction.
As noted elsewhere (20, ~1), these studies indicated
rapid repair and maturation to the small lymphocyte stage in the radiation-damaged thymuses of
the thigh-shielded animals. In contrast, there was
an apparent arrest of maturation in the regenerating thymuses of the unshielded group, and their
cortices remained filled with large and mediumsized lymphoid ceils, with a sustained high level
of mitotic activity. A very similar process was
noted during the course of regeneration and tumor
development in subcutaneous thymic implants in
thymectomized irradiated C57BL or hybrid hosts.
This state of maturation arrest merged imperceptibly with the development of frank lymphomas,
which remained confined to the thymic cortex for
sonic weeks before invading the medulla, the capsule, and adjacent structures.
I t was then noted that the normal thymus of
1-day-old C57BL mice differs strikingly from the
adult pattern, by virtue of having almost a "pure
culture" of large and medium lymphocytes, with
a very high mitotic index, occupying the outer
zone of the cortex and comprising from one-third
to one-half of its entire width (Fig. 1). During the
first days of life, this zone of immature cells differentiates rapidly and becomes indistinguishable
from the inner cortical zone, composed predominantly of mature small lymphocytes. By 10 days
Vol. ~1, S e p t e m b e r 1961
of age, the immature cell zone is only about one
cell layer thick, and by s
days it is gone (Fig.
2; 32). Since the only two hosts known to be susceptible to the strain C57BL lymphoma agent
were (a) newborn mice and (b) irradiated (unshielded) mice, and since both of these revealed a
similar and striking prevalence of immature
lymphoid cell forms, it was postulated (~0) t h a t
the viral agent may have a particular affinity or
requirement for these undifferentiated cells as
hosts and that it is not able to proliferate at an
equilibrium-sustaining level leading to tumor development except in the presence of an adequate
reservoir of such immature cells. If this view is
correct, then the role of x-radiation in the induction of these tumors in strain C57BL mice may
be plausibly attributed to its twofold action in (a)
injuring the adult thymus, thus exciting a stimulus
for regeneration which recapitulates the neonatal
thymic lymphoid cell differentiation sequence and
(b) injuring the bone marrow, thus interfering
with the capacity of marrow cells to promote rapid
regeneration (22) and maturation in the radiationinjured thymus.
Further support for the view that the lymphoma viruses may have a particular requirement for
immature lymphoid cells may now be found in the
observation by Arnesen (~) and Metcalf (~5) t h a t
the high-leukemia AK strain, in which irradiation
is unnecessary for lymphoma development, is constitutionally adrenal-insufficient, with a concomitantly long-sustained thymic hyperplasia, in
which immature cells are continuously abundant.
Finally-, the low-leukemia C3H strain, which is
susceptible to the freshly extracted AK lymphoma
virus only during the postnatal period, has now
been shown by the ingenious quantitative studies
of Axelrad and Van der Gaag (3) to exhibit a transient zone of immature cells in the outer cortex,
similar to t h a t in newborn C57BL mice, during a
brief postnatal period which correlates well with
the time of susceptibility to virus inoculation.
Although such evidence is based on correlation,
rather than direct experimental proof, these observations remain consistent under such diverse conditions as to make their chance occurrence highly
unlikely. I t may therefore be concluded that the
1~o. 1.--Normal thymus of 1-day-old C57BL mouse. Note
the zone of large, pale-stainingimmature lymphoid cells in the
subeapsular region of the cortex. Darker-staining, mature small
l~nphocytes are abundant in the inner cortex. X 1~5.
FIo. ~.~Normal thymus of ~0-day-old C57BL mouse. The
zone of large, pale-staining cells has disappeared. X 125.
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Research.
KAPLAN--Cell Differentiation and Virus Carcinogenesis
state of differentiation of the lymphoid cells of the
host is a significant determinant of susceptibility
to this family of viruses. It seems likely that differentiation and/or physiological activity of target
tissues will also prove to be important in the case
of other oncogenic viruses, and that systematic
investigations along these lines will be rewarding.
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The Role of Cell Differentiation as a Determinant of
Susceptibility to Virus Carcinogenesis
Henry S. Kaplan
Cancer Res 1961;21:981-983.
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