..-
THE EFFECTS OF FATTY ACIDS IN THE DIET ON THE RATE OF
DEDIFFERENTIATEON OF SERIALLY TRANSPLAN1'ED MAMMARY GLAND TUMORS
AN HONORS THESIS (10 499)
JILL RENEE MURRELL
DR. ALICE S. BENNETT
BALL STATE UNIVERSITY
I ND I f~t\lf;
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TABLE OF CONTENTS
J !"',rrHUDLJCT I Uh!"
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, • " .•. " " ••... " " . " " ..• " " " " " " " • " " " " " " " . " " " " " " " " • " '-.'
FELI~'rED LITEf~:p!rUPE"."."."." .• ""."""".""""".".".".,,~.'.i
Effects of fatty acids on tumorigenesis
Pi" 0!3 t. I:~g 1 C":"lnd i n~;
!-'!or-mone c,;;
Membrane fluidit.y
Cell communicat.ion
IrnmunE' systE,>m
FEI)JB\I [iF
DI··"(.::I
dam"'tqE~
Effects of fat.t.y acids on differentiat.ion
Het.erogeneit.y of tumors
Differentiating agent.s
Physiological changes
Progression of t.he mammary tumor
Histological rating and growth rates
MATEFIALS AND METHODS .. " .• "." .. " ... ,," •• ,,"" """"""."."."" ..•. ,,12
Transplant.ation procedure
Slide preparation
Fat.t.y ac~d ext.raction
Lipid ex~ract.ion
Cholesterol ext.ract.ion
F:1::::SUL..·Th (.:iND D I ~3Cl..Jf:;h I UN" " " " " " " • " " • " " • " " " " " " " • " " " " " " " " " " " " " • " " :\1:.;
Histolog:cal analysis
[iF'ot.-,lth rate
Lipid and Cholesterol
Sl..JI';II"1f.:iHY" " •• " • " " •• " • " " " " " " •• " " " • " • " " " • " " • " ••
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REFERENCES CITED .. "" .. """"""" .. """.""" .". ,," ""." ."" ".""." .".,,20
-
-
Research has determined that dietary fat has a definite
effect on the development of spontaneous mammary tumors in mice.
Current research involves identifying the effects of
different level and types of fat on tumorigenesis.
the
Results
indicate that a hiqh fat diet promotes tumor development when
iii
1 Cl~'J -fat di (:,t"
ln addition linoleic acid, a
pCllyunsaturated fatty acid and its products are essential -for
t umot'-
-f 0(' rOd t :i. (:In"
In our IBboratory, certdin conclusions can be made from
results obtained.
First, a diet high in stearic acid, a
saturated fatty dcid, delays tumor development.
The latency
period for spontaneous mammary tumor formation is longer for
Stronq Strdin A mice fed a high fat stearic acid rich diet than
for mice fed a low fat stock dIet.
stearic acid or safflower
011
Second, the inclusion of
(rich in linoleic dcid)
in the diet
produces alterations in fatty acid compClsition of mammary gland
tumors when compared to those from animals Cln a stock diet.
NUl"!"'"
tumorous tissue such as brown and white adipose tissue and
mammary gland tissue uf steric acid animals showed no increase in
stearic acid storage and significdnt reductions in the
percentages
hand,
o~
polyunsaturated fats were evident.
large stores of linoleic acid were observed in tissues of
Finally. the role of fatty acids in early
tumorigenesis is being investigated.
.-
nodules (HAN)
Hyperplastic alveolar
which are considered to be a prelude to mammary
adenocarcinomas were observed in the mice.
Preliminary results
show distinct differences in eppeerence of the glends end number
of HAN
present in mice fed the different diets.
memmery glends hed the fewest HAN and resembled normal
on-
lactating glands.
The purpose
~f
this research project wes to determine the
effect of dietery fetty ecids on the rate of dedifferentiation of
serielly trensplented memmery gland tumors.
tissue becomes less differentiated or distinct and spectalized.
geared toward normal mammary gland function.
This research determined the
effect of the two different diets on this process.
IS
essential for better understanding of human tumor progression
as the tumor metastasizes and the mechanisms underlying the
change each metastasized tumor undergoes.
It may also suggest
a mechanism explaining how each of the fatty acids affect
tumor progression.
Fatty acid composition, lipid content! and
amount of cholesterol in the tumors, white and brown adipose
tissue of later transplants were also determined.
4
REVIEW OF RELATED LITERATURE
Many conclusions, similar to those descussed in the
introduction, have been reached concerning the effect of dietary
fats on mammary tumorigenesis
(1).
Dietary fat definitely plays a
role in mammary tumor formation.
Tinsley
(2)
diet mixtures.
tested eleven different fats and oils in nine
Tumor incidence increased and the latency period
decreased with increases in linoleate
the other extreme,
(18:2)
concentration.
increasing levels of stearate (18:0)
At
decreased
incidence and increased the latency period for tumor formation.
Linolenate (18:3)
and oleate (18:1)
dId not have any effect.
This research siqnified that one cannot just classify fats as
polyunsaturated and saturated;
different effects
individual fatty acids have
(3-4).
Rao and Abraham
(5)
found that linoleic acid promoted the
growth of 5mg transplanted mammary tumor tissue whereas fat free
diets and diets rich in saturated fat detained growth.
linoleate tumors weighed three to four times more.
The
The
researchers hypothesized that the enhancement was due to the
production of arachidonic acid.
Linoleate is a precursor tn
arachodonic ac:id production which is used in the production of
prostaglandins (PG).
Prostaglandins are thought to depress the
immune system making tumor formation more probable.
Similar results were obtained by Hillyard and Abraham
In addition, :.ndomethacin
(.004%)
(6).
which inhibits prostaglandin
production was administered and resulted in reduced tumor qrowth
in animals fed the linoleate rich diet.
5
This suggested that the
immune system did playa part in the role of tumorigenesis.
studied the effect of indomethacin in rats on
2-5% and 10-20% fat diets on tumor growth.
inhibit qrowth in the 2-5% fat diet.
Indomethacin did
No prostaglandin E2
(PGE2)
was produced In the treated mice whereas PGE2 production by
mononuclear cells from spleens of untreated rats increased as fat
i ncrooec\s,oed.
Other research indicates that arachidonic acid
content of plasma lipids is significantly elevated in animals on
a safflower
011
diet
(8).
Physiologic levels of PGE2, as well as other hormones such
as insulin and estradiol which are normally found in the body,
increase stem cell differentiation and decrease stem cells' DNA
synthesis tumor formation in nude mice.
prostaglandins depends on the tIssue and cell
~ype.
induction of differentiation by PGs occurs only in early induced
tumcw<s
(7).
Much more work is needed to be done on
prostanglandin effects on tumorgenesis before sound conclusions
Ci::\n b\:? mac:le.
Besides prostaglandin production, dietary fats may induce
tumor formation by means of greater hormone responsiveness
(10)
Mammary hormones such as prolactin, growth hormone, estrogen,
progesterone, glucocorticoids, insulin, and thyroxine are thought
to be important in tumor development.
However, once the tumor is
established and advanced, it is no longer hormone-dependent
(11).
Evidence for the role of fat in increased hormone
-
responsiveness lies in the observations that animals on high fat
6
diets have longer estrous cycles and early puberty.
Also an
increase in the secretory activity of the endocrine system is
Dbser-vE~d
.
In
contrast~
responsive to hormones.
some fats stimulate tumors which are nonIn turn, hormones may provide critical
growth-promoting fatty acids (linoleic) to the tissues.
The
presence of hlgh levels of prolactin in tissues showed a shift
Dietary fats may also have an effect on membrane
fluidity.
Po:.yunsaturated fats have a lower melting point thus
producing a more fluid membrane.
This increase in fluidity
effects macromolecule mobility, receptor availability, enzyme
actibity,
prostaglandin~
virus invasion, and cAMP, cGMP, amino
acid and carbohydrate transport.
Chanqes in these cell
components may trigger cell division.
Cholesterol incorporated
into the membrane increases the viscosity (3).
suggest that the greater the viscosity of the membrane, the
greater resistance to malignant transformation
Fatty
ac~ds
(12).
may also promote tumorigenesis by inhibiting
cell-to-cell communication.
Cell-cell communication is essential
in the control of differentiation and growth.
of this may be a possible mechanism in tumor promotion.
promoters such as phorbol esters block cell communication.
Unsaturated fatty acids, oleic and linoleic block cell-cell
communication in V79 Chinese hamster cells whereas saturated fats
h'::1Cj
-
no effect:"
(10)
Moreover, fatty acids are observed to suppress immune
':'"'Iett vi t·y.
This may be achieved through prostaglandin synthesis.
7
-
Research by Summerfield and Tappel
(13)
reveals that high levels
of polyunsaturates in the absence of vitamin E decreases DNA
template activity in hepatic cells suggesting peroxidative
damage to the DNA.
The different possible mechanisms by which fatty acids may
promote tumorIgenesis may be due to the heterogeneity of the
cells found in the mammary gland tumor
(14).
mammary tumors contain widely heterogeneous subpopulations.
Tumors are a mixture of cells.
Specific characteristics of a
tumor are the result of the selection of certain variant cells
within the tumor.
Histologically undifferentiated cells are seen
intermingled with completely differentiated cells.
In
spontaneous tumors, MMTV production and antigen expression may
vary singificantly among clones of cells.
to
characteri~e
Thus,
it is important
the various cells in order to understand the
biDlogicc:\1 Plr'()per'ties of the tumor-,
Not only is it important to know cell characteristics so
that tumor action can be predicted but it is also important in
planning
trea~ment.
cytodE-~=,tr'uct
i
'/i:.""! ..
(1 ~:!)
Drugs now used in chemotherapy are
Their use is danqerous because of a lack of
specificity and sometimes a result is not obtained.
that are non-destructive need to be developed.
currently
loo~(ing
maJ.:iqnancy.
for differentiating agents which may revert the
Hec£?nt E-\/i denc:e i ndi catE':s t.hat the mal i gr'I':,'Int.
~";ti::\tE:'
is not irreversible but represents a disease of altered
maturation. The findings that some tumors can be
chemical agents to differentiate to mature end-stage cells with
El
no proliferative potential is promising.
Oddly enough, these differentiating agents are also tumor
promoters which exemplifies the fine line between proliferation
and
For example,
different~ation.
linoleic acid has been shown
to stimulate differentiation in all types of myogenic cells.
In Allen's research
(16)
linoleic acid intake resulted in a hiqh
deqree of satellite cell differentiation and fusion without a
noticeable increase in cell number.
Differentiation results in many cellular changes not
phenotypically oriented.
Changes in cell cycle activity
changes in membrane receptor and responsiveness to
(17)
such
fact~rs
as hormones and dietary fats were just 50me of the observations
recorded in cell differentiation.
Some researchers used
these characteristics to label or find a differentiated or
nor··I-···d iff E.~I"f:·::-!t:. ated cE,ll
each of
~he
(:I.
7·····:i '-"1)
•
According to Paterson
(19)
morphologically intermediated cellular stages
1S
characterized by a unique polypeptide pattern, and novel
protein(s) are synthesized at each intermediate stage.
The study of mammary gland tumorigenesis lends itsel
to the study of the progression of morphologic
Premalignant leisions form
(HAN)
then it
F·'
1·
011_
neoplastic stage.
Abraham
20)
~DDked
transplanted
'Drm ___
~t
th0 effecrs of linole2te on the growth of
tissue, HAN tissue, neoplastic tissue
He found that normal mammary epithelium was regulated in qrowth
by other cells and the edge of the fat pad but growth was faster
in mice on the linoleate diet.
normal regulation
HAN tissue had lost some of the
mediated by other cells and the fat pad but
was not affected bv the linoleic diet or the addition of
i ndomet.I··!2i.c in"
However, transplanted HAN qrew at a faster rate in
mice fed the linoleic diet.
Aqain an increase in growth rate
and incidence of transplanted tumors in linoleate fed mice was
E·ncountered.
In response to spontaneous mammary tumors, they have been
shown to be s\ow growing, differentiated and responsive to fatty
As this tumor is transplanted, histological
and growth rate changes are observed which may result or be the
result of
die~ary
fat.
In Hurst's work
(21)
tumors were scaled from 0 to 5
according to the amount of ductal, and acine-alveolar structures,
stromal components, vascularity and necrosis.
She found that the
scores went succeedingly down as t.he transplant generation
Two main cell types were also found in 80% of the tumor
small basophilic cells wit.h barely visible nuclei and
cytoplasm and large lightly stained cells with several nuclei
(21) •
The small basophilic cells made up the larger population
in the earlier generations but switched places as the generation
of transplantat.ion increased.
-
given by Sherbet
(22)
This finding supports evidence
that in the dedifferentiated state, a loss
1 ()
cells to a particular type,
:j.nc::t-t2i::\~=.£·?
tn
!:3:~·ZE~
and also a
loss of cytoplasm and an
uf the nuclf.-?u!:;:. ;;:(r·le:! nuc] eol i
DCC:UI'-'::·
in t.hE:
cf.?ll ,:;:.•
These tumor changes occur by progression during serial
transplantation and does not depend on the host.
proven by tumor growth studies in vitro and in vivo
Not only do hist.ological
(21,23).
chanqes occur in transplanted
tumors but also an increase in tumor growth can be seen
shortentng of the mitotic cycle and of
wit.hcJut CI···;Eln(JE· :in "I":.I""·le pr·opol·-t.ion 0+
that
.
its Sand 81 phases but
timf.-~
':;:.pi~nt
:i.n Fl.
II
I··ie found
that the Sand 81 phases were progressively being shortened
wit.h no change in the 82 phase.
selection increased,
generation,
jt
cell
result.ing in the progressive select.ion 0+
cells with a greater potential
In conclusion,
As each generat.ion passed,
for rapid growth and growth in a
as a tumor is moved from generation to
becomes dedi++erentiated with the cells focusing
more on growth than the oriqinal
function 0+ mammary tissue;
mammary-like structures disappear,
replaced by large nucleus and
nucleolied cells which maintain and incredse cell
The significance of all
thu~
division.
of these findings is in their
application to the treatment of cancer patients.
progression of t.hese mouse tumors and the effects of dietary fat
should increase the understandinq of how the cells of similar
1 1.
MATERIALS AND METHODS
_..lr.~"~D"g~:p".1...i::.\IJ":t..s:.t..j,."g"!J..
Sets of three, four month old Strong Strain A mice (26)
diets containing 14% steric acid
(15% fat
were used for tumor transplantation.
fpd
and i5% safflower oil
Controls were mice fed a
A spontanpous mammary tumor was surgically
removed from a mouse fed the stock diet.
A cell suspension
prepared by m:ncing the tumor in RPMI with 5% fetal
calf serum
was injected subcutaneously into the experimental animals.
of this tumor was saved for slide preparation and lipid analysis.
When the transplanted tumor grew to the width of about a
centimeter and a half,
it was surgically removed, a cell
suspension prepared, and injected into the next generation.
Slides were prepared according to published procedures.
After removing the tissue, a small piece is place in 10% buffer
paraffin by soaking the tissue in 30% to 100% ethanol.
complete water removal, tissue was soaked in two changes of 100%
alcohol.
With the water removed, the tissue is ready to be set in
P':lf"c.'1pl i:'lst.
Th<:;:,
tis.~:;ue
\.-'Ji~S·
pl""cE?d in IIH2lted
~·)i::\rr.'\pli:~~::.t.
was placed in a vacuum oven to remove air bubbles in paraplast
The hot wax was then poured into a peel-away square.
When the paraplast hardens,
it was sectioned with a
1';;::
-
mi
Clr
'
The 4um ribbons were placed and arranged on clean
otomf:?"
The slides were placed on a warming tray
Tissues were stained with hematoxylin and eosin y.
o\iE'I'''ni qht.
Cells were examined microscopically to determine structural
characteristics of the tumors.
scaled from 0-5 (21).
Structural differences were
Dates were recorded to determine growth
rate and thess data were compared amoung tumors.
Tissues
(tumor, brown and white adipose tissue)
fatty acid analysis,
saved for
lipid content and cholesterol amounts were
stored in .025% acetic acid in the refrigerator until ready.
Tissues were then placed in 15ml conical test tubes with
approximately 2ml of 15% KOH in 85% methanol.
refluxed in a hot water bath
(85 C)
for one and a half hours.
Sufficient concentrated HCl was added to the cooled solution so
that it was
a~idic
to litmus.
"j"j"1 e
<:Ole: 1, CI'].
+",], f.0C1
extracted into an equal volume of hexane.
The water layer was
removed and an equal volume of saturated KCl
solution was added
Again, the aqueous later was removed.
full
of anhydrous Na2S04 was added to the sample to remove the
The solution was then transfered to another
centrifug~
A
tube and evaporated under N2.
80~20
mixture of ether:methanol was added to redissolve
the fatty acids.
The samples were methylated with diazomethane
procedure by Schlenk and Gellermen
f.0VapOt"'a'i::ec! un(:iE':!!''' nitl'''O(::jE'n ar'iC! thf?'
(30).
int?th/latE~d
fatty ;:'.,c:::i.ci5;
stored at -20 C until gas liquid chromatoqraphic:: analysis
j ...":"..:,
VJE'!,,'E'
(GLC).
. . J::h ~~tS2.:t§~I:~. <;~L . _.en.!-;,,: . L.:'t.~2j:...??.
The same tumor tissue that was used for the fatty acid
extraction was used for cholesterol analysis.
saponification step applied.
The cholesterol was extracted
The cholesterol/hexane solution
into hexane before HCl was added.
was washed with saturated KCI and anhydrous Na2S04.
was then evaporated under nitrogen.
The cholesterol samples were
stored at -20 C until GLC analysis .
. ..1:.i.Q.tjL. ~~.;:.~ . t..X~~~~.:£:.:_t . ~l!l._..t~.t.!.. .
Weighed tumor tissues in 6X volume .025% acetic acid were
homogenized with a Vertis homogenizer at minium speed for one
Next~
the tissues stood under nitrogen for 15
followed by a 5 min.
minutes~
centrifugation at 3000 rpm.
which should have a pH of 4.0 was discarded.
repeated then C19 and cholestane standards
we~e
added.
The
pellet was slurried in a 40; volume of chloroform:methanol.
tissues were homoqenized and centrifuged.
contained the lipid was saved.
The
The supernatant which
These steps were repeated twice.
The supernatants were combined and flash evaporated at room
tE~mpF..'!ra.tur
e.
The moist residue was taken up in less than five
mililiters of 2:1 chloroform:methanol then filtered thru a
Whatman GFJA glass fiber filter.
The filter was rinsed and the
volume was brought up to five mililiters in a conical test tube.
The five mililiter solution was dried in weigh boats.
The
constant dried weight was subtracted from the original weight of
the bC)D.t.
This number represents the weight of lipid present in
that amount of tumor tissue.
RESULTS AND DISCUSSION
......Ij.L§:.tg.LE;:gl_~.;.§..L .....A.L").. ~~J.. y..??. L~2.
A differentiation index
(01)
which equals alveoli + ductual
ratings/2 + stromal rating was used to compare characteristics of
Overall, the tumors consisted of many types of cells,
thus beinq... heteroqeneous.
_.
Some areas of the tissue would be
differentiated whereas other areas showed a dedifferentiated
s:.tru.ctU!'··E' .
Ir general
l
the tumors did progressively become less
differentiated in animals on the stock diet
(Table 1).
pattern seemed to occur in transplanted tumors in animals on the
steric acid diet.
The safflower tumors seem to be more
differentiated than either the stock or steric transplants.
Various patterns and conclusions can be drawn from the
transplanted stock tumors.
(1)
As the tumors were serially transplanted, the normal
mammary gland structure was progressively lost thus becominq
dedifferentiated.
(2)
No predictable changes in the frequency of the ductal
structures were found but transplants were lower than the
spontaneous tumors in ductal ratings.
Appearance of acini/aveoli structures varied.
i'-..\u
stroma was observed aruund these structures.
(4)
Later transplants showed a marked decrease in the
appearance uf struma.
necrotic as compared with the spontaneous tumur and early
_
tl'- C\n~::;p 1 ant s.
(5)
Later transplants had a greater blood supply than early
(It has been observed in the lab that tumored
animals seem to have a larger volume of blood compared to nontumored animals).
(6)
Necrotic tissue appears around blood vessels where
possible lymphocyte filtration has occurred.
II}
The seventeenth stock transplant was extremely
dedifferentiated with no noticeable stroma and few ductal
It consisted of masses of cells interrupted
with blood vessels.
Patterns in the safflower and stearic acid transplants were
not as evident as in the stock transplants.
(1)
SAF transplants were rated relatively higher than
either the stock or steric acid transplants.
(2)
Overall, the SAF exhibited a higher frequency of ductal
s;t r" uc t ur-' (:?5·.
di
~::.t
i nqui
In addition, the structures were large and easily
~,.hE\b]' f2.
(3)
None of the SAF tumors exhibited alveoli structures.
(4)
SAF transplants had higher frequencies of stroma that
(5)
The stearic acid tumors closely resembled the stock
tumors in appearance and differentiation index.
Gross changes in appearance were also observed.
tumor was transplanted it transformed from a tiqht cohesive mass
The tissue fell
was made up
o~
individual separate cells.
1.6
apart as if it
This non-cohesiveness
-
may be due to the lack of stroma.
Time of growth was varied among the safflower and stearic
acid transplants (Table 3).
The safflower transplants took an
average of 32 days to grow while steric acid transplants took
days, not a
s~gnificant
difference.
The stock transplants only
took an average of 24 days which may be due to the more
dedifferentiated state of the tumor.
The growth period for the
stock was also pretty predictable and decreased from an average
of 30 days to the 27 day average observed after 19 passes.
The data obtained for growth rate study were not entirely
reliable and should not be used to compare among the different
dir=:ts..
The injection procedure did not entail countinq the
ceilis actually in the suspension.
Therefore each animal
receive the same number of tumor cells.
did not
The injected animals who
were on the same diet and of the same generation probably did
receive the same number of cells since the same suspension was
Since then, a procedure has been developed in
which the number of live cells injected is known.
High percentages of lipid were observed in the safflower and
stearate diets, but even these numbers were not consistant
3).
(Table
The percent of lipid in stock transplants was relatively
steady,
increasing slightly as the tumor was transplanted.
1··li gh
lipid content was observed in three tumors which took over 30
d,,:\y~:;
tD
dE:O'v'(;?1
op.
Cholesterol
levels varied in the stock transplants with two
:1.7
exhibiting high levels.
BAF transplants also showed high levels
of cholesterol but not if the high percent of lipid is taken in
con~:.i
dE!I'" at i on.
Only 5% of the total
BAF transplants.
the total
lipid was cholesterol in the
On the other hand, cholesterol made up 11% of
lipld in the stock tumors.
cholesterol, 2.5% of total
An even lower percent of
lipid, was found in the stearic
Fatty acid analysisi was done to determine what
fatty acids actually made up the rest of the lipid.
contraints, the raw data was not analyzed for this paper but will
18
HIS'rOLOGICAL RATINGS OF SERIALLY TRANSPLANTED
I"! PI i"! !"·I(.'I H VT U t"1 U r.~: ::;
NUMBER
DUCTS ALVEOLI STROMA VASCULAR NECROSIS
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CHARACTERISTICS OF TUMOR TRANSPLANTS
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DIFFERENTIATION INDEX.
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.I.
Histological and chemical studies revealed that the mammary
gland tumor studied in this paper was very heterogeneous.
The stock transplants dedifferentiated with
increasing passages.
Never-the-Iess, the tumor did not
dedifferentiate at a constant rate, giving the graph
irregular appearance.
(Table 2)
an
This also exemplifies the heterogeneous
nature of the tumor.
The amount of stroma seemed to be a good marker for
dedifferentiation.
It progressively decreased with each passage.
Alveoli structures were evident in stock and stearic acid tissue.
Present r&search is studying the abilities of differentiating
agents to differentiate stem (19)
cells to the specilized alveoli
cells which are evident during pregnancy and have a special
function.
The appearance of these structures in the transplants
may signify a differentiated state.
The high differentiation index of the SAF transplants may be
due to the effects of linoleic acid as in the myogenic cells
discussed in the literature review
(16).
In summary, the tumors were found to be heterogeneous,
agreement with reports in the literature (10,20).
in
Due tD thE:'
different types of cells, the different stage of development that
they are in and the specialized effects of the dietary fatty
acids, specific alterations were difficult to observe.
SUGGESTIONS FOR FURTHER RESEARCH
This research should be repeated with a spontaneous tumor
that would be transplanted in animals of the three diets,
of transplanting an already dedifferentiated tumor.
affect of each diet would be more easily determined.
instead
The true
Growth rate
would also be reliable if the new procedure for cell counting is
used.
Transplantations could also be done in animals on the corn
diet and the new sterie acid diet with a high level of
linoleate (to determine if steric acid is inhibitinu the
effect of linoleate).
Blood cholesterol levels could also be measured in order to
determine whe":her the bulk of the cholesterol is located in the
tissue qr the blood.
determine if
~umored
Blood level studies may be done to
animals exhibited a hiqher blood volume than
non-tumored animals.
The next step in this study would be to switch the animal to
another diet once the tumor cells were injected.
Perhaps
transplanting a safflower tumor to a steric acid mouse and
observing any structural changes between the original tumor and
the transr,lanted one would provide further infromation on the
effect of fatty acids on cell differentiation.
Another
interesting project would be to quantify the number of estroqen
receptors on the transplanted tissues and correlate these
findings with the differentiation index.
20
F~EFE!:;;~ENCE:b
C I TED
Effects of dietary
Carrington, C.A. and H.L. Hosick (1985).
1..
fat on the growth of normal, preneoplastic and neoplastic mammary
epithelial cells in vivo and vitro.
Journal of Cell Science
7~5: 269---27E3.
2:u
-rin:::.lf?V"
luJu'}
\JaAn
St:hi'n:i.i:. . ~::.?
£:"tnc]
[),:f~1J
F' if2F'CE'
(lC?81)
II
Influence of dietary fatty acids on the incidence of mammary
Cancer Research 41~1460-1465.
tumors in the C3H mouse.
3.
Reiser, R., JuL. Probstfield, A. Silvers, L.W. Scott, M.L.
Shorney, R.D. Woed, B.C. O'Brian, A.M. Gatto Jr., O.Phil, and W.
Insull Jr. (1985).
Plasma lipid and lipoprotein response of humans
to beef fat, coconut oil, and safflower oil.
American Journal of
Clinical Nutrition 42:190-197.
4.
Erickson" K.L., D.S. Schlanger, D.A. Adams, D.R. Fregeau,
and J.S. Stern (1984).
Influence of dietary fatty acid
concentration and geometric configeration on murine mammary
tumorigenesis and experimental metastasis.
Journal of Nutrition
114: 1f:r31.~--1f::l42"
5.
Rao, G.A. and S. Abraham (1976).
Enhanced growth rate of
transplanted nammary adenocarcinoma induced in C3H mice by
dietary linoleate.
JNCI56(2):431-432.
Effect of dietary
Hillyard. L.A. and S. Abraham (1979).
pol yun~:::-i::<.tLll'"- Eltf:'cj f ;;:It ty i::<.C i cI~:- on qr-DI"lth clf InI3miT!.:~,r-y E:lr.:!f:-::nclca_r--c i r-Iom"'l~:­
in mice and rats.
Cancer Research 41:1460-1465.
7.
Kollmorgan, G.M., M.M. King, S.D. Kosanke, and C.Do (1983).
Influence of dietary fat and indomethacin on the growth of
transplantable mammary tumors in rats.
Cancer Research 43:47144719.
8.
Croft, K.D., L.J. Beilin, R. Vandonqen, and E. Mathews(1984).
Dietary modifications of fatty acid and prostaglandin synthesis
in the rat.
Biochimica et Biophysica Acta 795:196-207.
9.
Rudland, P.S., A.T. Davies, and M.J. Warburton (1982).
Prostaglandin-inducer.:! differentiation or dimethyl sulfoxideinduced differentiation:
Reduction of the neoplastic potential
of a rat mammary tumor stem cell line.
JNCI 69(5)~1083-1090.
10.
Welsch, C.W. and C.F. Aylsworth (19f::l3).
Enhancement of
murine mammary tumorigenesis by feeding high levels of dietary
fat:
A hormonal mechanism?
JNCI 70(2)~215-221.
-
11.
Gregorio~ 0.1., L.J. Emrich, S. Graham, J.R. Marchall, and
T. Nemoto (1985).
Moderate changes in linoleate intake do not
influence the systemic production of E prstaglandins.
L1Plds
20 (5) : :~:6B--2)'2.
12.
Inbar, M. and M. Shinitzky (1974).
Increase of cholesterol
level in the surface membrane of lymphoma cells and its
inhibitory effect on ascites tumor development.
Sci USA 71~2128-2130.
1 ::::;.
SummE'r·f:i.<o?ld.! F.vJ. and rCi.L. T,,~ppel (19f3 LI·).
dietary polyunsaturated fats and vitamin E on aging and
peroxidative damage to DNA.
Archives of Biochemistry and
Biophysics 233(2)~408-416.
14.
Danielson, K.G., L.W. Anderson, and H.L. Hosick (1980).
Selection and characterization in culture of mammary tumor cells
with distinct~ve growth properties in vivo.
40~ IH:I.:?·····lF:l(}.
·1
r::
( :I. c.y'~3 ::.i) "
Malignant cell differentiation
British Journal of Cancer
j:jot",'nti,:;,l therapeutic approach.
.I. ._.! •
c:\
16.
Allen, RuE., L.S. Luiten, and M.V. Dodson (1985).
Effect of
insulin and l:noleic acid on satellite cell differentiation.
Journal of Animal Science 60(6):1571-1579.
17.
Decosse, J.J. ~ C.L. Gossens, and J.F. Kuzma (1973).
Induction of differentiation by embryonic tissue.
18.
Newman, R.A., P.J. Klein, and P.S. Rudland (1979).
Binding
of peanut lectin to breast epithelium, human carcinomas, and a
cultured rat mammary stem cell:
Use of the lectin as a marker of
JNCI63(6):1339-1346.
19.
Paterson, F.C. and P.S. Rudland (19H5).
Indentification of
nD\iel ~ st,;~,gE~-··~:.pec if i c: pol ypept i dE'~::. a,::.~:·oc:: i E'tt£0ci ~.,I:i. th i::.!··iE::!
differentiation of mammary epithelial stem cells to alveolar-like
cells in culture.
Journal of Cellular Physiology 124:525-53H.
20.
Abraham, 5., L.J. Faulkin, L.A. Hillyard, and D.J. Mitchell
(1984).
Effect of dietary fat Dn tumorigenesis in the mouse
mammary gland.
JNCI 72(6):1421-1429.
21.
Hurst~ J., L. Ferayorni, C. Scarantion, D.
Emma, and J.C.
Kovacs.
Mammary tumor progression:
Histology vs clonogenicity.
C,~:\ncer- He£,.eal·-·c:h.
-
G.V.
Neoplasia and Cell Differentiaiton.
S. 1< ":\f·· fj er-·
~
1 Cj7 4.
Neoplastic Development.
1 C?69 •
Foulcls, L..
P,cctclerni c Fr·£~~::.s:·'.1
2::~;.
'v' 0 1
In
:::<1-.
St<=:f::~l, C~., ~:::" (~,d,3.ms, ,J. Hod<:'.:Jf?tt, i.'tnd P • .J':'1nik.
CE,:t1
population kinetics of a spontaneous rat tumor during serial
transplantation.
British Journal of Cancer 25:802-811.
25.
McCredie, J, W. Inch, and R. Sutherland (1971).
Differences
in growth and morphology between the spontaneous C3H mammary
carcinoma in the mouse and its syngeneic transplants.
Cancer
27: 6::~;5·-·64~?
2f:..~
Journal
II
27.
(~irm,
I::;~es::.e''''.r·
D.
of Heredity 27:21-25.
( 1 (jiB2) .
c h p i.-:'.p E~r- ,
Histology of fats in mouse mammary t i
II"',eli ani:\ University-Purdue University.
s::·~·ue.
Humason, G.L. Animal Tissue Techniques.
Freeman anel Company, 1967.
2[3.
Essentials of Practical
29.
Galigher, A.E. anel E.N. Kozloff.
Lea and Febiger, Philadelphia, 1971.
Microtechnique 2nd ed.
~:;o
.
Schlenk, H.
and J.L.
Gellerman
( :I. 960) •
Anal.
Chem.
32:1412.
:31.
Phillips" F. and D.S. Pl'-iv;::,:,tt (1978).
A ~=.:implified pY"ocec:!ulre
for the quant:tative extraction of lipids from brain tissue.
Lipids 14(6);590-595.
32.
Ferretti! A., J.T Judd, M.W. Marshall, V.P. Flanagan, J.M.
Roman, and E.J. Matusik Jr.
(1985).
Moderate changes in
linoleate intake do not influence the systemic production of E
prostaglandins.
L.ipids 20(5):268-272.
33.
Piers, G.B., P.K. Nakane, A. Martinez-Hernandez, and J.M
Ward (1977).
Ultrastructural comparison of differentiation of stem
cells of murine adenocarcinomas of colon and breast with their
normal counterparts.
JNCI 58(5): 1329-1345.
-