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/ . Embryol. exp. Morph. Vol. 20, 3, pp. 329-41, November 1968 3
2 9With 4 plates
Printed in Great Britain
The development of teratomas from intratesticulargrafts 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 usuallyresult 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 tubalmouse ova to the
anterior chamber of the eye and one developed the threeprimary germ
layers and then regressed, probably because the host and donorwere
histo-incompatible. This is the only report of an ectopically
graftedpre-uterine egg that developed intra-embryonic derivatives.
Kirby (1962&, 1965)grafted oviducal segmenting mouse eggs to
the kidney and obtained only tropho-blast 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
blastocystsis higher in the testis than for any other extra-uterine
site that he tried. He men-tioned the ease and certainty with which
the blastocyst can be introduced intothe testis, and that the
vascularity of the testis is such that it affords a particu-larly
good bed for blastocyst development. I have also found (Stevens,
1964)that embryonic tissues develop well as intratesticular grafts,
and that the testicu-lar environment exerts a strong
teratocarcinogenetic influence on 12-day genitalridges of strain
129/Sv mice. This influence results in the initiation of
develop-ment of male primordial germ cells. They proliferate and
give rise to undif-ferentiated embryonic cells which in turn give
rise to the primary germ layers.The primary germ layers
differentiate into disorganized mixtures of many kindsof tissues
characteristic of teratomas.
Testicular teratomas develop spontaneously from primordial germ
cells inabout 10 % of the males of strain 129 mice (Stevens, 1967
a) during the 13th dayof gestation. They never develop in females
of this strain. It is possible thatteratomas do not develop from
female primordial germ cells because they mayhave 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.
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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 develop-ment of ovarian
teratomas.
The initiation of development of strain 129 male primordial germ
cells by thetesticular environment suggested the possibility that
the development of femalegerm cells might be similarly initiated
after meiosis is complete, i.e. afterfertilization. To explore this
possibility, fertilized strain 129 single- and two-cellova were
grafted into the testes of adult males. The influence of the sex
hor-mones on teratocarcinogenesis is not yet understood, but the
results presentedhere demonstrate that strain 129 two-cell eggs can
develop in a disorganizedmanner in the testis. Cells in these
growths may remain as proliferating un-differentiated embryonic
cells for remarkably prolonged periods of time. In onecase 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 preventsome cells from
differentiating. Alternatively, the undifferentiated cells mayhave
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 micecongenic with inbred strain 129/Sv. This stock, 129/Sv-SFCP,
was developedby introducing the genes, C, P, and SlJ into the
strain 129 genome, and ischaracterized by its relatively high
incidence (10 %) of spontaneous testicularteratomas. Hereafter
these mice will be referred to as strain 129. Other inbredstrains
used were obtained from colonies maintained at The Jackson
Laboratoryby other investigators. They include strains A/HeJ,
C57BL/10Sn, C57BL/6J,and C57BL/6K.S. An Fx hybrid, 129 x A/He, was
also used.
Eggs were removed from the oviduct 0-2 days after the mothers
were foundwith 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 hourswithout deleterious effects. Follicular
cells adhering to the zygotes were alsografted and the zona
pellucida was left intact. A single egg was grafted to
eachtestis.
Hosts ranged in age from about 1 month to fully mature males.
They wereanesthetized with Avertin, and testes were exposed through
a median ventralincision in the skin and body wall. Eggs were drawn
into a micropipette attachedto rubber tubing with a mouthpiece and
were expelled through a tear in thetunica albuginica into the
testis. All grafts were isogenic and were recoveredfrom 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 somegrafts were retransplanted subcutaneously
to male hosts.
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Egg graft teratomas 331
RESULTS
The results of grafting eggs to adult testes are summarized in
Table 1. Onlyfour of 250 grafts of single cell eggs were recovered.
Some of the two-cell strain129 eggs underwent development, but none
from strains other than 129 exceptfor one from a 129 x A/He
hybrid.
Table 1. Results of grafting early mouse embryos to adult
testes
Stage
1-cell
2-cell
4- to 8-cell
Strain
129
A/Hex 129
129
A/HeJ
129 x A/He
C57BL/10Sn
C57BL/6JC57BL/KS
129
A/HeJ
Age ofgraft(days)
—
—
714172130405060
3060
3060
3060
20
20
203060
30
Numbergrafted
230
20
144
1211490
14151
243
13096
7827
459
14
10
3315057
27
Extraembryonic
deriva-tivesonly
3
1
2022148
105
00
00
00
0
0
6200
2
Em-bryonicderiva-tives
0
0
13178
1458
00
10
00
0
0
1171
0
Withundifferen-
tiatedembryonic
cells
0
0
23165522
00
00
00
0
0
1101
0
The histologic composition of the grafts varied according to the
length oftime the graft was left in the testis. As would be
expected, grafts of embryosrecovered a short period of time after
transplantation contained embryonic andimmature cells. Most grafts
which contained embryonic derivatives and wererecovered after long
residence in the testis were composed of adult-type
tissues.Unexpectedly, grafts of embryos left for as long as 30-60
days containedundifferentiated embryonic cells and immature
tissues. The undifferentiatedembryonic cells in these long-term
grafts were still proliferating and giving rise
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332 L. C. STEVENS
to undifferentiated as well as to differentiated cells. They
retained the capacityto grow progressively for long periods like
the stem cells of teratomas.
The embryonic cells that developed from the graft were not
confined byReichert's membrane, but were free to migrate through
the interstitial areasaway from the original graft site (Plate 1,
fig. E). This migratory activity dis-rupted normal intercellular
relationships.
The behaviour of the grafts of eggs to the testis is described
below accordingto their age. Since all were isografts, they did not
succumb to the homograftreaction, but were able to survive
indefinitely.
Single-cell eggs (day 0)
Two hundred and fifty zygotes were removed from the oviducts of
mice onthe day a copulation plug was found and they were
transplanted into adulttestes (Table 1). Only four of these were
recovered 14-20 days later, and theywere 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 aftertransplantation. Fifty-two of these were composed of
extraembryonic deriva-tives only, but 47 had embryonic shield
derivatives as well. Twenty-six of thosewith embryonic derivatives
contained undifferentiated embryonic cells, evenafter 21-60 days,
when all cells might be expected to be differentiated.
Nineteenlooked grossly like teratomas that can be serially
transplanted indefinitely, andthey were regrafted into other hosts.
One (LS5364) behaved as a transplantableteratoma 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)
issurrounded by ectoderm (EC) which is enveloped by proximal
endoderm (PE). Reichert'smembrane (R) has been secreted by distal
endoderm (DE). The embryo is surrounded bytrophoblastic 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) andendoderm (PE) are separated by mesodermal
cells (M). The distal yolk sac has secreted athick 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),
andtrophoblastic 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 repre-sented in Fig. D. The
darkly stained cells at the right are undifferentiated cells that
havemigrated away from the original graft site, x 100.
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/ . Embryol. exp. Morph., Vol. 20, Part 3 PLATE 1
L. C. STEVENS facing p. 332
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/ . Embryol. exp. Morph., Vol. 20, Part 3 PLATE 2
L. C. STEVENS
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Egg graft teratomas 333grafted into the adult testis, and only
one graft was recovered (Table 1). Thiswas 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 graftsof strain 129 eggs are described according to the length
of time they were allowedto remain in the host.
(1) 7-day-old grafts of two-cell eggs. Three grafts of two-cell
eggs were re-covered after 7 days of development in the testis. Two
were composed of a fewtrophoblastic 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 andextra-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
wasattached to Reichert's membrane which was secreted by a single
layer of distalyolk sac cells. A cavity separated the distal from
the proximal endoderm. Theproximal endoderm enveloped the
ectodermal epithelium which surrounded theproamniotic cavity. There
were a few cells of the ectoplacental cone. The embryowas well
organized, and the relationship of the ectoderm and endoderm
wassimilar 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 wererecovered 14-17 days after grafting. One
consisted of trophoblastic giant cellsin a pool of blood. Another
consisted of trophoblastic giant cells and a fewdistal yolk sac
cells embedded in a blob of Reichert's membrane. Four
otherscontained both extra-embryonic and embryonic immature tissues
in variousdegrees of organization. In some areas of the grafts,
ectodermal, mesodermal,and endodermal components were arranged in
the normal relationship to oneanother. In other areas, on the other
hand, the cells were arranged as dis-organized chaotic mixtures.
One contained an ectodermal vesicle surrounding aproamniotic 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, mesenchymalcells, 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 andpart 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
testiculartubules. 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
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334 L. C. STEVENS
separated from proximal endoderm by a sheet of mesodermal cells.
There werenucleated red blood cells in primitive blood vessels. The
distal endoderm hadsecreted a thick Reichert's membrane, and
trophoblastic giant cells were attachedto it. Other
undifferentiated cells were located distal to the embryo. There
wereseveral areas of necrotic material. Plate 1, figs, D and E
illustrate growths com-posed of ectoderm, proximal endoderm, distal
endoderm, Reichert's membraneand 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 mesen-chymal cells, and a large area of fetal heart
muscle. The remaining two graftswere disorganized growths with
extra-embryonic and embryonic derivatives.
(3) 21-day-old grafts of two-cell eggs. Twenty-eight growths
from two-cell eggswere recovered 21 days after grafting. All
contained extra-embryonic derivatives.Twenty-one were composed
solely of extra-embryonic cells. Seven grafts con-tained embryonic
derivatives, and six of these had undifferentiated and imma-ture
cells. In addition to extra-embryonic derivatives and
undifferentiated cells,the following were represented: embryonic
ectoderm and endoderm, mesenchy-mal cells, immature
neuro-epithelium (Plate 2, figs. A and B), and fetal heartmuscle.
One graft contained a small nodule of cartilage.
(4) 30-day-old grafts of two-cell eggs. Twelve growths were
recovered 30 daysafter grafting. Four were composed solely of
extra-embryonic derivatives, andeight had embryonic derivatives.
The only embryonic shield derivative in threegrafts was immature
and adult neural tissue. Five other grafts were pleomorphicand
contained undifferentiated embryonic cells and immature and adult
neuraltissue, various types of epithelium, striated muscle,
cartilage, heart muscle,glandular tissues, bone, and adipose tissue
(Plate 2, figs. C and D). One graftwas very pleomorphic and the
tissues were highly differentiated. It contained animmature testis
(Plate 2, fig. D). This is the only growth that contained
gonadaltissue.
(5) 40- to 50-day-old grafts of two-cell eggs. Thirty-seven
grafts were recovered40-50 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
epithelialcysts. x200.Figs. D-F. First subcutaneous transplant
generation from a growth derived from a two-cellegg 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.
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/. Embryo!, exp. Morph., Vol. 20, Part 3
L. C. STEVENS facing p. 334
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/. Embryol. exp. Morph., Vol. 20, Part 3 PLATE 4
wmsm^
L. C. STEVENS
-
Egg graft teratomas 335derivatives 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
riseto 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. Fivewere composed of extra-embryonic derivatives only.
Six developed into tera-tomas composed of mature tissues. Two
others were pleomorphic, but hadundifferentiated embryonic cells
and immature tissues (Plate 2, fig. F, andPlate 3, figs. A-C). They
were large growths, approximately 25 mm indiameter, and
superficially they resembled transplantable testicular
teratomas.Portions of them were minced and grafted subcutaneously
to 12 male hosts. Mostof these grafts survived until the hosts were
killed 5-7 months later, and theywere 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
retransplantablesize 1 month after grafting. It was regrafted
subcutaneously to six males, andgrew to a large size in three of
them. These were pleomorphic, but, like trans-plantable teratomas,
they contained undifferentiated embryonic cells in additionto
immature and mature tissues. They were maintained as
transplantableteratomas (Plate 4, figs. B-E) and contained
undifferentiated and immatureelements for five subcutaneous
transplant generations (165 days). After thefifth generation, all
of the cells differentiated and the tumors failed to
growprogressively. The two-cell egg that gave rise to this teratoma
was grafted intothe testis August, 1966, and the last growth
derived from it was retransplantedin 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
afterfinding copulation plugs and were grafted to the testes of
adults (Table 1).Forty-seven growths were recovered 20, 30, and 60
days after transplantationand 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 egggrafted 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.
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336 L. C. STEVENS
after grafting two-cell eggs. Some growths were pleomorphic and
containedundifferentiated embryonic cells and immature tissues as
well as well-differen-tiated tissues. The grafts of A/HeJ eggs
yielded only two growths of extra-embryonic derivatives. The
descriptions below pertain only to strain 129eggs.
(1) 20 days after grafting. Seven grafts were recovered and six
of them con-tained extra-embryonic derivatives only. One, in
addition to trophoblast anddistal endoderm with Reichert's
membrane, had immature neuroepithelium andundifferentiated
cells.
(2) 30 days after grafting. Thirty-seven grafts were recovered.
Twenty hadonly 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 egggraft was recovered. It was highly pleomorphic,
containing immature and adultnerve, many kinds of epithelia,
striated muscle, cartilage, bone with marrow,adipose tissue,
pigment cells, and undifferentiated cells. There was a
metastaticgrowth composed of adult nerve in the left renal lymph
node.
DISCUSSION
Two main findings are reported here: (1) Tubal two-cell strain
129 mouseeggs 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 remarkablylong periods of time.
Single-cell mouse eggs did not develop embryonic derivatives
when graftedto adult testes. Growths from only four of 250 zygotes
grafted to the testis wererecovered, and they were composed solely
of trophoblastic cells. Similarly,intratesticular grafts of
two-cell (1-day) and four- to eight-cell (2-day) eggs fromstrains
A/HeJ, C57BL/Ks, and C57BL/10Sn failed to develop
embryonicderivatives. In contrast, several growths containing
embryonic derivativesdeveloped 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 alwaysnecessary
for the development of grafted strain 129 eggs.
The donors and hosts were genetically identical, except for
Y-linked genes, sothat the growths from grafts of two-cell eggs
were not rejected, and they couldbe observed over extended periods
of time. Seven days after transplantation, agraft was composed of
trophoblastic giant cells, Reichert's membrane, distaland proximal
endoderm, and a layer of ectoderm surrounding a proamnioticcavity.
The extra- and intra-embryonic derivatives were organized as in
normal6-day mouse embryos. After 7 days, the grafts became
disorganized, and theintra- and extra-embryonic components were
arranged as a chaotic mixture.
All of the cells in the growths were derived from the grafted
eggs. The growthswere readily distinguished from host cells, and
the two-cell eggs were free of
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Egg graft teratomas 337follicle cells. Approximately 10 % of
strain 129 males have spontaneous testicularteratomas. These tumors
are all detectable grossly in adults, and mice withteratomas were
not used as hosts.
The term ^ undifferentiated embryonic cells' is used here to
refer to cells whichcannot be identified by morphological criteria
as being derived from any of thethree primary germ layers. They are
indistinguishable from the stem cells ofteratocarcinomas which
Pierce (1967) and Stevens (1967 a), have shown to bepluripotent.
They give rise to differentiated tissues and to more
undifferentiatedproliferating cells like themselves.
After 20 to 60 days some of the growths derived from two-cell
egg graftsconsisted of mixtures of well-differentiated adult-type
tissues. Others containedadult tissues and in addition, immature
and undifferentiated cells, even after60 days. It was unexpected
that proliferating undifferentiated and immaturecells would be
found in such old grafts. In one case, a graft of a two-cell
egggave rise to a growth that grew subcutaneously for five
transplant generations,and retained undifferentiated elements for
at least 165 days. Many of the 20- to60-day grafts looked
histologically like, and one behaved like the
transplantableteratomas derived from the testes of strain 129 mice
(Stevens, 1958).
We are currently maintaining transplantable teratomas derived
from 3- to6-day embryos grafted to the testes of adults. One is
still growing progressivelyafter 2 years of serial transplantion,
and several others continue to grow pro-gressively after 1-1/2
years. The transplantation behavior of these tumors willbe the
subject of a later article.
Kirby (1963) found that when mouse blastocysts were grafted to
the adulttest is, development usually ceased at the morphological
stage at which thedefinitive placenta should develop. He thought
that failure of the placenta todevelop is presumably a contributory
cause of death of the graft. Our resultsshow that the grafts of
two-cell mouse embryos will develop and survive in-definitely.
Apparently an adequate blood supply develops along with the
growthof the graft as it does for transplantable tumors.
The embryonic cells that developed from the grafts were not
confined byReichert's membrane. They were free to migrate through
the interstitial areasaway from the original graft site. This
migratory activity disrupted the normalintercellular relationships,
and may have resulted in the delay in the onset ofdifferentiation.
Possibly some of the cells which remained
morphologicallyundifferentiated for prolonged periods of time
failed to receive stimuli fromother cells they would normally be in
contact with. An alteration of histogeneticinteractions may
underlie the delay in differentiation.
Abbot & Holtzer (1966) found that if differentiated
chondrocytes are estab-lished as monodispersed cultures, they cease
to synthesize chondroitin sulfateand collagen, and begin to
synthesize DNA and to proliferate. This change inbehavior is
reversible. They proposed that a chondrocyte whose cell membraneis
engaged in amoeboid movement cannot make chondroitin sulfate, but
that
-
338 L. C. STEVENS
DNA synthesis and proliferation is promoted. They advanced a
theory thatadherent chondrocytes reciprocally stabilize their cell
membranes which allowsthem to make chondroitin sulfate. They point
out that this theory and theconcept of contact inhibition proposed
by Abercrombie & Heaysman (1954)are obviously analogous. This
kind of alteration of intercellular relationshipmay be involved in
the delay of differentiation reported here for cells in growthsfrom
two-cell eggs.
Bernfield & Fell (1967) found a delay in development to the
fully differen-tiated state in pancreatic re-aggregates and the
significance of this was unclearto them. They suggested the
possibility that the small size of the explant may beinvolved, but
that it seemed more likely that interactions between
differentiatingand proliferating cells may be a factor in the
control of the expression of genomicfunction during
development.
The disorganization of the embryo after the 6-day stage
disrupted normalcellular relationships and may have delayed the
determination of the cells sothat they remained in an
undifferentiated state for prolonged periods of time.This
interpretation could be used to support the theory, sometimes
referred toas the 'misplaced blastomere theory,' proposed by
Askanazy (1907), and upheldby Needham (1950), Nicholson (1950),
Willis (1962), Collins & Pugh (1964),and Pugh & Smith
(1964) that teratomas originate from embryonic totipotentcells that
have escaped the influence of embryonic organizers. It is the
onlyexperimental evidence to support this view, but I do not
believe that it isproof.
The spontaneous and experimentally induced testicular teratomas
of strain129 mice are derived from primordial germ cells during the
13th day of fetal lifeand not later (Stevens, 1966, 19676). All of
the spontaneous teratomas arisewithin the seminiferous tubules
(Stevens, 1962). When male genital ridges withprimordial 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 tera-tomatous foci, and, as in the early spontaneous tumors,
these foci also arisewithin the seminiferous tubules (Stevens,
1964).
Homozygous steel (57/57) mice have very few if any primordial
germ cells.When genital ridges from SI I SI mice are grafted into
adult testes, they developedinto testes without teratomas (Stevens,
19676). Their fertile littermates ( + / +and 5//+) have primordial
germ cells and approximately 75 % of intratesticulargenital ridge
grafts from them developed into testes with teratomas. These
de-velopmental and genetic studies demonstrate that testicular
teratomas arederived from primordial germ cells.
The development of teratomas from two-cell eggs may be explained
in twodifferent ways. They may develop directly from undetermined
disorganizedproducts of the two-cell egg. Alternatively, the
grafted two-cell egg may giverise to cells which become determined
and develop characteristics of primordialgerm cells, and it is
these that give rise to the teratomas. When genital ridges
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Egg graft teratomas 339
with primordial germ cells are grafted to the adult testis, as
were the two-celleggs, they do develop teratomas. We are attempting
to obtain evidence that willhelp decide between these alternative
explanations.
SUMMARY
1. A small proportion of two-cell (but not one-cell) strain 129
mouse eggsdevelop intra- and extra-embryonic derivatives when
grafted to adult testes. Thedevelopment of grafted one-cell zygotes
was extremely infrequent, and em-bryonic derivatives were never
observed.
2. Two-cell eggs develop in the testis quite normally for about
a week, butlater they become disorganized mixtures of many kinds of
embryonic and adulttissues.
3. Some growths derived from grafts of two-cell eggs have
undifferentiatedcells 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 fromtwo-cell egg grafts may be attributed to the
disruption of normal intercellularrelationships. Alternatively, the
undifferentiated and immature cells may havebeen 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 adultesse developpent des tissus
embryonnaires et des tissus extra embryonnaires.Les zygotes greffes
ne se developpent que tres rarement, et des tissus embryon-naires
n'y sont jamais observes.
2. Les oeufs du stade deux cellules se developpent normalement
dans letesticule pendant une semaine, mais au-dela, ils forment des
amas inorganisesdans lesquels on retrouve divers tissus
embryonnaires et adultes.
3. Certaines tumeurs issues de la greffe d'ceufs du stade deux
cellules contien-nent des cellules non differenciees, et des tissus
immatures, jusqu'a 60 jours, etmeme dans une cas, jusqu'a 165
jours.
4. Le retard de certaines cellules a se differencier peut etre
du a la rupture desrelations normales intercellulaires. Les
cellules non differenciees peuventegalement deriver de cellules
semblables aux cellules germinales primordiales.
This investigation was supported in part by Public Health
Service Research GrantCA-02662, from the National Cancer Institute,
and a grant from The William H. DonnerFoundation, Tnc.
The principles of laboratory animal care as promulgated by the
National Society forMedical Research, are observed in this
Laboratory.
I gratefully acknowledge the expert assistance of Don S.
Varnum.
-
340 L. C. STEVENS
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