/. Embryol. exp. Morph. Vol. 20, 3, pp. 329-41, November 1968
With 4 plates
Printed in Great Britain
329
The development of teratomas from intratesticular
grafts of tubal mouse eggs
By LEROY C. STEVENS1
The Jackson Laboratory, Bar Harbor, Maine
Grafts of cleaving tubal ova from non-inbred mice to ectopic sites usually
result in growths composed of extra-embryonic but not embryonic tissues
(Fawcett, Wisloki & Waldo, 1947; Fawcett, 1950; Jones, 1951; Whitten 1958;
Kirby, 1960, 1962a; Billington, 1965; and others). Runner (1947) grafted tubal
mouse ova to the anterior chamber of the eye and one developed the three
primary germ layers and then regressed, probably because the host and donor
were histo-incompatible. This is the only report of an ectopically grafted
pre-uterine egg that developed intra-embryonic derivatives. Kirby (1962&, 1965)
grafted oviducal segmenting mouse eggs to the kidney and obtained only trophoblast and extra-embryonic membranes. He concluded that a 'uterine factor'
is necessary for the development of intra-embryonic structures from mouse eggs.
Kirby (1965) and Billington (1965) grafted morulae and blastocysts to the testis,
and the morulae never gave rise to embryonic shield derivatives.
Kirby (1963) found that the proportion of successful grafts of blastocysts
is higher in the testis than for any other extra-uterine site that he tried. He mentioned the ease and certainty with which the blastocyst can be introduced into
the testis, and that the vascularity of the testis is such that it affords a particularly good bed for blastocyst development. I have also found (Stevens, 1964)
that embryonic tissues develop well as intratesticular grafts, and that the testicular environment exerts a strong teratocarcinogenetic influence on 12-day genital
ridges of strain 129/Sv mice. This influence results in the initiation of development of male primordial germ cells. They proliferate and give rise to undifferentiated embryonic cells which in turn give rise to the primary germ layers.
The primary germ layers differentiate into disorganized mixtures of many kinds
of tissues characteristic of teratomas.
Testicular teratomas develop spontaneously from primordial germ cells in
about 10 % of the males of strain 129 mice (Stevens, 1967 a) during the 13th day
of gestation. They never develop in females of this strain. It is possible that
teratomas do not develop from female primordial germ cells because they may
have already entered into meiosis. Fetal female germ cells remain in meiotic
1
Author's address: The Jackson Laboratory, Bar Harbor, Maine 04609, U.S.A.
330
L. C. STEVENS
prophase, and do not undergo mitotic division until fertilization (Borum, 1961).
The prenatal cessation of mitosis in female germ cells may prevent the development of ovarian teratomas.
The initiation of development of strain 129 male primordial germ cells by the
testicular environment suggested the possibility that the development of female
germ cells might be similarly initiated after meiosis is complete, i.e. after
fertilization. To explore this possibility, fertilized strain 129 single- and two-cell
ova were grafted into the testes of adult males. The influence of the sex hormones on teratocarcinogenesis is not yet understood, but the results presented
here demonstrate that strain 129 two-cell eggs can develop in a disorganized
manner in the testis. Cells in these growths may remain as proliferating undifferentiated embryonic cells for remarkably prolonged periods of time. In one
case the growth from a two-cell egg was serially transplanted for five generations,
and it retained its embryonic nature, like a malignant teratoma, for 5£ months.
The disorganized nature of the cells and tissues within the grafts may prevent
some cells from differentiating. Alternatively, the undifferentiated cells may
have arisen from cells with characteristics of primordial germ cells.
MATERIALS AND METHODS
Most of the animals used in this investigation were from a stock of mice
congenic with inbred strain 129/Sv. This stock, 129/Sv-SFCP, was developed
by introducing the genes, C, P, and SlJ into the strain 129 genome, and is
characterized by its relatively high incidence (10 %) of spontaneous testicular
teratomas. Hereafter these mice will be referred to as strain 129. Other inbred
strains used were obtained from colonies maintained at The Jackson Laboratory
by other investigators. They include strains A/HeJ, C57BL/10Sn, C57BL/6J,
and C57BL/6K.S. An F x hybrid, 129 x A/He, was also used.
Eggs were removed from the oviduct 0-2 days after the mothers were found
with mating plugs and were stored in Hank's solution. As pointed out by Kirby
(1963), eggs and embryos can be held in saline solution for several hours
without deleterious effects. Follicular cells adhering to the zygotes were also
grafted and the zona pellucida was left intact. A single egg was grafted to each
testis.
Hosts ranged in age from about 1 month to fully mature males. They were
anesthetized with Avertin, and testes were exposed through a median ventral
incision in the skin and body wall. Eggs were drawn into a micropipette attached
to rubber tubing with a mouthpiece and were expelled through a tear in the
tunica albuginica into the testis. All grafts were isogenic and were recovered
from the testis 7 to 60 days after transplantation, fixed in Vandegrift's solution,
serially sectioned at 7 fi and stained with hematoxylin and eosin. Parts of some
grafts were retransplanted subcutaneously to male hosts.
Egg graft teratomas
331
RESULTS
The results of grafting eggs to adult testes are summarized in Table 1. Only
four of 250 grafts of single cell eggs were recovered. Some of the two-cell strain
129 eggs underwent development, but none from strains other than 129 except
for one from a 129 x A/He hybrid.
Table 1. Results of grafting early mouse embryos to adult testes
Stage
1-cell
2-cell
Strain
129
—
A/Hex 129
—
230
20
3
1
0
0
0
0
129
7
14
17
21
30
40
50
60
30
60
30
60
30
60
20
14
4
12
114
90
141
51
243
130
96
78
27
45
9
14
2
0
2
21
4
8
10
5
0
0
0
0
0
0
0
1
3
1
7
8
14
5
8
0
0
1
0
0
0
0
2
3
1
6
5
5
2
2
0
0
0
0
0
0
0
20
10
0
0
0
129
20
30
60
33
150
57
6
20
0
1
17
1
1
10
1
A/HeJ
30
27
2
0
0
A/HeJ
129 x A/He
C57BL/10Sn
C57BL/6J
C57BL/KS
4- to 8-cell
Age of
graft
(days)
Extra
With
embryonic
undifferenEmderivabryonic
tiated
Number
tives
deriva- embryonic
grafted
only
tives
cells
The histologic composition of the grafts varied according to the length of
time the graft was left in the testis. As would be expected, grafts of embryos
recovered a short period of time after transplantation contained embryonic and
immature cells. Most grafts which contained embryonic derivatives and were
recovered after long residence in the testis were composed of adult-type tissues.
Unexpectedly, grafts of embryos left for as long as 30-60 days contained
undifferentiated embryonic cells and immature tissues. The undifferentiated
embryonic cells in these long-term grafts were still proliferating and giving rise
332
L. C. STEVENS
to undifferentiated as well as to differentiated cells. They retained the capacity
to grow progressively for long periods like the stem cells of teratomas.
The embryonic cells that developed from the graft were not confined by
Reichert's membrane, but were free to migrate through the interstitial areas
away from the original graft site (Plate 1, fig. E). This migratory activity disrupted normal intercellular relationships.
The behaviour of the grafts of eggs to the testis is described below according
to their age. Since all were isografts, they did not succumb to the homograft
reaction, but were able to survive indefinitely.
Single-cell eggs (day 0)
Two hundred and fifty zygotes were removed from the oviducts of mice on
the day a copulation plug was found and they were transplanted into adult
testes (Table 1). Only four of these were recovered 14-20 days later, and they
were all composed solely of trophoblastic giant cells (Plate 1, fig. A).
Two-cell eggs (day 1)
Of 669 grafts of strain 129 two-cell eggs, 99 were recovered 7-60 days after
transplantation. Fifty-two of these were composed of extraembryonic derivatives only, but 47 had embryonic shield derivatives as well. Twenty-six of those
with embryonic derivatives contained undifferentiated embryonic cells, even
after 21-60 days, when all cells might be expected to be differentiated. Nineteen
looked grossly like teratomas that can be serially transplanted indefinitely, and
they were regrafted into other hosts. One (LS5364) behaved as a transplantable
teratoma and retained undifferentiated proliferating cells for at least 165 days.
Four hundred and nine two-cell eggs from mice other than strain 129 were
PLATE 1
Fig. A. Trophoblastic giant cells derived from a single-cell egg grafted to the testis for 20 days.
xlOO.
Fig. B. Growth from a two-cell embryo grafted for 7 days. The proamniotic cavity (P) is
surrounded by ectoderm (EC) which is enveloped by proximal endoderm (PE). Reichert's
membrane (R) has been secreted by distal endoderm (DE). The embryo is surrounded by
trophoblastic cells and it resembles a normal 6-day mouse embryo, x 430.
Fig. C. Growth from a two-cell embryo grafted for 14 days. An ectodermal vesicle (EC)
surrounds a proamniotic cavity (P) filled with cellular debris. The ectoderm (EC) and
endoderm (PE) are separated by mesodermal cells (M). The distal yolk sac has secreted a
thick Reichert's membrane (R). x 430.
Fig. D. Growth from a 2-cell embryo grafted for 14 days. It is composed of ectoderm (EC),
mesoderm (M), proximal endoderm (PE), distal endoderm, Reichert's membrane (R), and
trophoblastic cells; in approximately normal relationships to one another. x200.
Fig. E. Growth from a two-cell embryo grafted for 14 days. It is similar to the growth represented in Fig. D. The darkly stained cells at the right are undifferentiated cells that have
migrated away from the original graft site, x 100.
/. Embryol. exp. Morph., Vol. 20, Part 3
L. C. STEVENS
PLATE 1
facing p. 332
/. Embryol. exp. Morph., Vol. 20, Part 3
L. C. STEVENS
PLATE 2
Egg graft teratomas
333
grafted into the adult testis, and only one graft was recovered (Table 1). This
was a two-cell ¥1 hybrid, 129 x A/He, recovered 30 days after transplantation,
and it consisted of adult nerve.
The histological composition and the transplantation behavior of the grafts
of strain 129 eggs are described according to the length of time they were allowed
to remain in the host.
(1) 7-day-old grafts of two-cell eggs. Three grafts of two-cell eggs were recovered after 7 days of development in the testis. Two were composed of a few
trophoblastic giant cells like the graft of a single-cell egg represented in Plate 1,
fig. A. The third was much larger than the others and contained embryonic and
extra-embryonic cells arranged very much as in normally developing embryos
(Plate 1, fig. B). A network of trophoblastic cells bathed in a pool of blood was
attached to Reichert's membrane which was secreted by a single layer of distal
yolk sac cells. A cavity separated the distal from the proximal endoderm. The
proximal endoderm enveloped the ectodermal epithelium which surrounded the
proamniotic cavity. There were a few cells of the ectoplacental cone. The embryo
was well organized, and the relationship of the ectoderm and endoderm was
similar to that in 6-day embryos and in embryoid bodies of teratomatous origin
(Stevens, 1967 a).
(2) 14- to 17-day-old grafts of two-cell eggs. Six grafts of two-cell eggs were
recovered 14-17 days after grafting. One consisted of trophoblastic giant cells
in a pool of blood. Another consisted of trophoblastic giant cells and a few
distal yolk sac cells embedded in a blob of Reichert's membrane. Four others
contained both extra-embryonic and embryonic immature tissues in various
degrees of organization. In some areas of the grafts, ectodermal, mesodermal,
and endodermal components were arranged in the normal relationship to one
another. In other areas, on the other hand, the cells were arranged as disorganized chaotic mixtures. One contained an ectodermal vesicle surrounding a
proamniotic cavity filled with cellular debris (Plate 1, fig. C). The ectoderm was
PLATE 2
Fig. A. Growth from a two-cell egg grafted for 21 days. Embryonic epithelia, mesenchymal
cells, and immature neuro-epithelium are represented, x 200.
Fig. B. Growth from a two-cell egg grafted for 21 days. Embryonic ectodermal vesicles.
Testicular tubule of the host at lower left, x 430.
Fig. C. Growth from a two-cell egg grafted 30 days. Lower left, bone; top, adipose tissue and
part of a hair follicle; upper right, neural tissue; lower right, striated muscle, x 200.
Fig. D. Growth from a two-cell egg grafted 30 days. Alimentary epithelium and fetal testicular
tubules. x200.
Fig. E. Growth from a two-cell egg grafted for 50 days. Proliferating undifferentiated cells.
x430.
Fig. F. Growth from a two-cell egg grafted for 60 days. Undifferentiated and immature cells.
x200.
22
! E K M 2Q
334
L. C. STEVENS
separated from proximal endoderm by a sheet of mesodermal cells. There were
nucleated red blood cells in primitive blood vessels. The distal endoderm had
secreted a thick Reichert's membrane, and trophoblastic giant cells were attached
to it. Other undifferentiated cells were located distal to the embryo. There were
several areas of necrotic material. Plate 1, figs, D and E illustrate growths composed of ectoderm, proximal endoderm, distal endoderm, Reichert's membrane
and trophoblastic cells, all in approximately normal relationships to one another.
Another 14-day graft of a two-cell egg developed proximal and distal endoderm,
Reichert's membrane, disorganized masses of immature epithelial and mesenchymal cells, and a large area of fetal heart muscle. The remaining two grafts
were disorganized growths with extra-embryonic and embryonic derivatives.
(3) 21-day-old grafts of two-cell eggs. Twenty-eight growths from two-cell eggs
were recovered 21 days after grafting. All contained extra-embryonic derivatives.
Twenty-one were composed solely of extra-embryonic cells. Seven grafts contained embryonic derivatives, and six of these had undifferentiated and immature cells. In addition to extra-embryonic derivatives and undifferentiated cells,
the following were represented: embryonic ectoderm and endoderm, mesenchymal cells, immature neuro-epithelium (Plate 2, figs. A and B), and fetal heart
muscle. One graft contained a small nodule of cartilage.
(4) 30-day-old grafts of two-cell eggs. Twelve growths were recovered 30 days
after grafting. Four were composed solely of extra-embryonic derivatives, and
eight had embryonic derivatives. The only embryonic shield derivative in three
grafts was immature and adult neural tissue. Five other grafts were pleomorphic
and contained undifferentiated embryonic cells and immature and adult neural
tissue, various types of epithelium, striated muscle, cartilage, heart muscle,
glandular tissues, bone, and adipose tissue (Plate 2, figs. C and D). One graft
was very pleomorphic and the tissues were highly differentiated. It contained an
immature testis (Plate 2, fig. D). This is the only growth that contained gonadal
tissue.
(5) 40- to 50-day-old grafts of two-cell eggs. Thirty-seven grafts were recovered
40-50 days after transplantation. Nineteen of them contained embryonic shield
PLATE 3
Fig. A. Growth from a two-cell egg grafted for 60 days. Immature neuro-epithelium. x 200.
Fig. B. Growth from a two-cell egg grafted for 60 days. Immature muscle fibers, x 200.
Fig. C. Growth from a two-cell egg grafted for 60 days. Notochord (center) and epithelial
cysts. x200.
Figs. D-F. First subcutaneous transplant generation from a growth derived from a two-cell
egg grafted to the testis for 60 days.
Fig. D. Mature neural tissue, x 430.
Fig. E. Adipose tissue, hair follicles, and epithelium, x 100.
Fig. F. Immature muscle fibers and epithelial cyst, x 200.
/. Embryo!, exp. Morph., Vol. 20, Part 3
L. C. STEVENS
facing p. 334
/. Embryol. exp. Morph., Vol. 20, Part 3
wmsm^
L. C. STEVENS
PLATE 4
Egg graft teratomas
335
derivatives and seven had undifferentiated embryonic cells (Plate 2, fig. E).
Among the tissue types represented were immature and adult nerve, notochord,
thyroid, muscle, and various types of epithelium. One of these grafts gave rise
to a metastatic growth of adult neural tissue in the left renal lymph node.
(6) 60-day-old grafts of two-cell eggs. Thirteen grafts were recovered. Five
were composed of extra-embryonic derivatives only. Six developed into teratomas composed of mature tissues. Two others were pleomorphic, but had
undifferentiated embryonic cells and immature tissues (Plate 2, fig. F, and
Plate 3, figs. A-C). They were large growths, approximately 25 mm in
diameter, and superficially they resembled transplantable testicular teratomas.
Portions of them were minced and grafted subcutaneously to 12 male hosts. Most
of these grafts survived until the hosts were killed 5-7 months later, and they
were composed of adult type epithelium, cartilage, muscle, and hair follicles.
One, however, contained adult type tissues (Plate 3, figs. D-F, and Plate 4,
fig. A) and undifferentiated proliferating cells, and it grew to a retransplantable
size 1 month after grafting. It was regrafted subcutaneously to six males, and
grew to a large size in three of them. These were pleomorphic, but, like transplantable teratomas, they contained undifferentiated embryonic cells in addition
to immature and mature tissues. They were maintained as transplantable
teratomas (Plate 4, figs. B-E) and contained undifferentiated and immature
elements for five subcutaneous transplant generations (165 days). After the
fifth generation, all of the cells differentiated and the tumors failed to grow
progressively. The two-cell egg that gave rise to this teratoma was grafted into
the testis August, 1966, and the last growth derived from it was retransplanted
in July 1967. Seventy mice were recipients of grafts derived from that egg.
Four- to eight-cell eggs grafted to the testis
Strains 129 and A/HeJ eggs were removed from the oviduct 2 days after
finding copulation plugs and were grafted to the testes of adults (Table 1).
Forty-seven growths were recovered 20, 30, and 60 days after transplantation
and prepared for histological examination. Results were similar to those observed
PLATE 4
Figs. A-E. Subcutaneous transplant generations from a growth derived from a two-cell egg
grafted to the testis for 60 days.
Fig. A. First generation. Nodule of cartilage, primitive epithelium, and undifferentiated cells.
x200.
Fig. B. Third generation. Immature epithelium, mesenchyme, and undifferentiated cells.
x200.
Fig. C. Fourth generation. Immature neuro-epithelium and undifferentiated cells, x 200.
Figs. D-E. Fifth generation. Mesenchyme, primitive epithelium, immature neuro-epithelium,
and undifferentiated cells, x 200.
336
L. C. STEVENS
after grafting two-cell eggs. Some growths were pleomorphic and contained
undifferentiated embryonic cells and immature tissues as well as well-differentiated tissues. The grafts of A/HeJ eggs yielded only two growths of extraembryonic derivatives. The descriptions below pertain only to strain 129
eggs.
(1) 20 days after grafting. Seven grafts were recovered and six of them contained extra-embryonic derivatives only. One, in addition to trophoblast and
distal endoderm with Reichert's membrane, had immature neuroepithelium and
undifferentiated cells.
(2) 30 days after grafting. Thirty-seven grafts were recovered. Twenty had
only extra-embryonic derivatives. Seventeen had many types of adult tissues,
and ten of these had undifferentiated cells and immature tissues.
(3) 60 days after grafting. A single large growth resulting from a 2-day egg
graft was recovered. It was highly pleomorphic, containing immature and adult
nerve, many kinds of epithelia, striated muscle, cartilage, bone with marrow,
adipose tissue, pigment cells, and undifferentiated cells. There was a metastatic
growth composed of adult nerve in the left renal lymph node.
DISCUSSION
Two main findings are reported here: (1) Tubal two-cell strain 129 mouse
eggs are able to develop into disorganized growths in an extra-uterine site; and
(2) Differentiation of some cells in the growths may be delayed for remarkably
long periods of time.
Single-cell mouse eggs did not develop embryonic derivatives when grafted
to adult testes. Growths from only four of 250 zygotes grafted to the testis were
recovered, and they were composed solely of trophoblastic cells. Similarly,
intratesticular grafts of two-cell (1-day) and four- to eight-cell (2-day) eggs from
strains A/HeJ, C57BL/Ks, and C57BL/10Sn failed to develop embryonic
derivatives. In contrast, several growths containing embryonic derivatives
developed from grafts of two-cell and four- to eight-cell eggs of strain 129 origin.
Apparently the 'uterine factor' postulated by Kirby (19626, 1965) is not always
necessary for the development of grafted strain 129 eggs.
The donors and hosts were genetically identical, except for Y-linked genes, so
that the growths from grafts of two-cell eggs were not rejected, and they could
be observed over extended periods of time. Seven days after transplantation, a
graft was composed of trophoblastic giant cells, Reichert's membrane, distal
and proximal endoderm, and a layer of ectoderm surrounding a proamniotic
cavity. The extra- and intra-embryonic derivatives were organized as in normal
6-day mouse embryos. After 7 days, the grafts became disorganized, and the
intra- and extra-embryonic components were arranged as a chaotic mixture.
All of the cells in the growths were derived from the grafted eggs. The growths
were readily distinguished from host cells, and the two-cell eggs were free of
Egg graft teratomas
337
follicle cells. Approximately 10 % of strain 129 males have spontaneous testicular
teratomas. These tumors are all detectable grossly in adults, and mice with
teratomas were not used as hosts.
The term ^ undifferentiated embryonic cells' is used here to refer to cells which
cannot be identified by morphological criteria as being derived from any of the
three primary germ layers. They are indistinguishable from the stem cells of
teratocarcinomas which Pierce (1967) and Stevens (1967 a), have shown to be
pluripotent. They give rise to differentiated tissues and to more undifferentiated
proliferating cells like themselves.
After 20 to 60 days some of the growths derived from two-cell egg grafts
consisted of mixtures of well-differentiated adult-type tissues. Others contained
adult tissues and in addition, immature and undifferentiated cells, even after
60 days. It was unexpected that proliferating undifferentiated and immature
cells would be found in such old grafts. In one case, a graft of a two-cell egg
gave rise to a growth that grew subcutaneously for five transplant generations,
and retained undifferentiated elements for at least 165 days. Many of the 20- to
60-day grafts looked histologically like, and one behaved like the transplantable
teratomas derived from the testes of strain 129 mice (Stevens, 1958).
We are currently maintaining transplantable teratomas derived from 3- to
6-day embryos grafted to the testes of adults. One is still growing progressively
after 2 years of serial transplantion, and several others continue to grow progressively after 1-1/2 years. The transplantation behavior of these tumors will
be the subject of a later article.
Kirby (1963) found that when mouse blastocysts were grafted to the adult
test is, development usually ceased at the morphological stage at which the
definitive placenta should develop. He thought that failure of the placenta to
develop is presumably a contributory cause of death of the graft. Our results
show that the grafts of two-cell mouse embryos will develop and survive indefinitely. Apparently an adequate blood supply develops along with the growth
of the graft as it does for transplantable tumors.
The embryonic cells that developed from the grafts were not confined by
Reichert's membrane. They were free to migrate through the interstitial areas
away from the original graft site. This migratory activity disrupted the normal
intercellular relationships, and may have resulted in the delay in the onset of
differentiation. Possibly some of the cells which remained morphologically
undifferentiated for prolonged periods of time failed to receive stimuli from
other cells they would normally be in contact with. An alteration of histogenetic
interactions may underlie the delay in differentiation.
Abbot & Holtzer (1966) found that if differentiated chondrocytes are established as monodispersed cultures, they cease to synthesize chondroitin sulfate
and collagen, and begin to synthesize DNA and to proliferate. This change in
behavior is reversible. They proposed that a chondrocyte whose cell membrane
is engaged in amoeboid movement cannot make chondroitin sulfate, but that
338
L. C. STEVENS
DNA synthesis and proliferation is promoted. They advanced a theory that
adherent chondrocytes reciprocally stabilize their cell membranes which allows
them to make chondroitin sulfate. They point out that this theory and the
concept of contact inhibition proposed by Abercrombie & Heaysman (1954)
are obviously analogous. This kind of alteration of intercellular relationship
may be involved in the delay of differentiation reported here for cells in growths
from two-cell eggs.
Bernfield & Fell (1967) found a delay in development to the fully differentiated state in pancreatic re-aggregates and the significance of this was unclear
to them. They suggested the possibility that the small size of the explant may be
involved, but that it seemed more likely that interactions between differentiating
and proliferating cells may be a factor in the control of the expression of genomic
function during development.
The disorganization of the embryo after the 6-day stage disrupted normal
cellular relationships and may have delayed the determination of the cells so
that they remained in an undifferentiated state for prolonged periods of time.
This interpretation could be used to support the theory, sometimes referred to
as the 'misplaced blastomere theory,' proposed by Askanazy (1907), and upheld
by Needham (1950), Nicholson (1950), Willis (1962), Collins & Pugh (1964),
and Pugh & Smith (1964) that teratomas originate from embryonic totipotent
cells that have escaped the influence of embryonic organizers. It is the only
experimental evidence to support this view, but I do not believe that it is
proof.
The spontaneous and experimentally induced testicular teratomas of strain
129 mice are derived from primordial germ cells during the 13th day of fetal life
and not later (Stevens, 1966, 19676). All of the spontaneous teratomas arise
within the seminiferous tubules (Stevens, 1962). When male genital ridges with
primordial germ cells from 12-day (for strain 129) or 13-day (for strain A/HeJ)
fetuses are grafted into the testes of adults, they develop into testes with teratomatous foci, and, as in the early spontaneous tumors, these foci also arise
within the seminiferous tubules (Stevens, 1964).
Homozygous steel (57/57) mice have very few if any primordial germ cells.
When genital ridges from SII SI mice are grafted into adult testes, they developed
into testes without teratomas (Stevens, 19676). Their fertile littermates ( + / +
and 5//+) have primordial germ cells and approximately 75 % of intratesticular
genital ridge grafts from them developed into testes with teratomas. These developmental and genetic studies demonstrate that testicular teratomas are
derived from primordial germ cells.
The development of teratomas from two-cell eggs may be explained in two
different ways. They may develop directly from undetermined disorganized
products of the two-cell egg. Alternatively, the grafted two-cell egg may give
rise to cells which become determined and develop characteristics of primordial
germ cells, and it is these that give rise to the teratomas. When genital ridges
Egg graft teratomas
339
with primordial germ cells are grafted to the adult testis, as were the two-cell
eggs, they do develop teratomas. We are attempting to obtain evidence that will
help decide between these alternative explanations.
SUMMARY
1. A small proportion of two-cell (but not one-cell) strain 129 mouse eggs
develop intra- and extra-embryonic derivatives when grafted to adult testes. The
development of grafted one-cell zygotes was extremely infrequent, and embryonic derivatives were never observed.
2. Two-cell eggs develop in the testis quite normally for about a week, but
later they become disorganized mixtures of many kinds of embryonic and adult
tissues.
3. Some growths derived from grafts of two-cell eggs have undifferentiated
cells and immature tissues for at least 60 days, and in one case, 165 days.
4. The marked delay in the onset of differentiation of some cells derived from
two-cell egg grafts may be attributed to the disruption of normal intercellular
relationships. Alternatively, the undifferentiated and immature cells may have
been derived from cells with characteristics of primordial germ cells.
RESUME
Formation de teratome a partir de greffe d'ceufs de Souris,
dans le testicule de Souris adultes
1. Une petite quantite d'ceufs de Souris de la souche 129, au stade deux cellules
(mais jamais au stade une cellule) greffes dans le testicule de Souris adultes
se developpent des tissus embryonnaires et des tissus extra embryonnaires.
Les zygotes greffes ne se developpent que tres rarement, et des tissus embryonnaires n'y sont jamais observes.
2. Les oeufs du stade deux cellules se developpent normalement dans le
testicule pendant une semaine, mais au-dela, ils forment des amas inorganises
dans lesquels on retrouve divers tissus embryonnaires et adultes.
3. Certaines tumeurs issues de la greffe d'ceufs du stade deux cellules contiennent des cellules non differenciees, et des tissus immatures, jusqu'a 60 jours, et
meme dans une cas, jusqu'a 165 jours.
4. Le retard de certaines cellules a se differencier peut etre du a la rupture des
relations normales intercellulaires. Les cellules non differenciees peuvent
egalement deriver de cellules semblables aux cellules germinales primordiales.
This investigation was supported in part by Public Health Service Research Grant
CA-02662, from the National Cancer Institute, and a grant from The William H. Donner
Foundation, Tnc.
The principles of laboratory animal care as promulgated by the National Society for
Medical Research, are observed in this Laboratory.
I gratefully acknowledge the expert assistance of Don S. Varnum.
340
L. C. STEVENS
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{Manuscript received 25 March 1968)