[CANCER RESEARCH 41, 4154-4161
0008-5472/81
/0041-OOOOS02.00
, October 1981]
Metabolic Host Reaction in Response to the Proliferation of Nonmalignant
Cells versus Malignant Cells in
Ingvar Karlberg, Staffan Edström,Lars Ekman, Sonny Johansson, Tore Schersten, and Kent Lundholm2
Surgical Metabolic Research Laboratory, Departments of Surgery [I. K.. L. £., T. S., K. L.], Oto-Rhino-Laryngology
Sjukhuset, University of Göteborg, Göteborg, Sweden
[S. E.], and Pathology II [S. J.J, Sahlgrenska
ABSTRACT
INTRODUCTION
This study was aimed at elucidating whether proliferation of
malignant cells is a prerequisite for the development of the
multiform relationship between a malignant tumor and its host.
A methylcholanthrene-induced
sarcoma (MCG 101) trans
planted s.c. in nongrowing weight-stable mice (C57BL/6J) was
used as the tumor model. Injection i.p. of Corynebacterium
parvum vaccine in C57BL/6J mice stimulated normal cells to
exponential cell growth. This experimental model was consid
ered suitable for elucidating the metabolic host response to
proliferation of nonmalignant cells.
C. parvum-stimulated animals showed an exponential growth
rate of the spleen and liver leading to a more than doubled
tissue mass over 9 days. This was compared with the expo
nential growth rate of the tumor over 10 to 12 days in tumorbearing mice. Tissues other than the tumor and the spleen
showed unchanged or decreased dry weight in tumor mice,
indicating in terms of weight a proliferation mainly of tumor
tissue, which was confirmed by histological examination and
biochemical analyses in various tissues. Metabolic balance
studies, body composition, evaluation of hepatic and muscle
protein synthesis, as well as energy metabolism and measure
ment of the plasma concentration of various substances, were
performed in C. parvum-stimulated animals. All together 29
different parameters were selected for measurement in C.
parvum-stimulated animals. These parameters which reflected
the overall and intermediary metabolism were compared with
the results obtained in previous studies in tumor-bearing mice.
Twenty-one of the 29 parameters showed the same pattern of
alteration in C. parvum-stimulated animals as in tumor-bearing
animals. Frequency analyses showed no significant difference
between the metabolic host reactions in the C. parvum-stimu
lated animals compared with the "tumor-stimulated"
animals,
Exponential cell growth in vivo has been established in
experimental tumors (25, 26). This rapid proliferation of malig
nant cells kills the tumor host in cachexia and is associated
with a multiform metabolic relationship between the tumor and
the host (36). The mechanism behind the metabolic response
of the host to the proliferation of malignant cells leading to
cancer cachexia is as yet unclear (30, 41, 43). Experimental
studies suggest that tumor-associated factors may in part be
responsible for this tumor host reaction (8, 9, 11, 13, 33, 35,
39, 40). On the other hand, it has been postulated that there is
no need to generate specific hypothesis for the development
of the tumor-induced cachexia. The contribution of inadequate
suggesting a similar response in these models. These experi
mental models seem therefore suitable for comparison of, and
may be useful for mechanistic analyses of, the metabolic host
reaction in response to proliferation of malignant versus non-
Mice. Nongrowing, weight-stable 3-month-old C57BL/6J mice bred
in this laboratory and weighing 22 to 25 g were used (19-23). They
were housed in ordinary cages, if not otherwise stated, and were
maintained on Purina laboratory chow diet and tap water ad libitum. In
any particular experiment, study and control mice were matched for
age, sex, and size. In some experiment, pair-fed control mice were
used. Pair-fed animals received the same amount of food day by day
as the tumor-bearing animals ate spontaneously. These experiments
malignant cells.
We conclude that several metabolic host reactions found in
tumor-bearing animals may be directly coupled to cell prolif
eration per se but not necessarily to the proliferation of malig
nant cells.
1This work was supported by grants from the Swedish Cancer Society (Project
93), the Swedish Medical Research Council (Project 536). the Assar Gabrielsson
Foundation, the Swedish Society of Medical Sciences, and the Serena Ehren
strömFoundation.
2 To whom requests for reprints should be addressed, at Department of
Surgery I, Sahlgrenska Sjukhuset, University of Göteborg, S-413 45 Göteborg,
Sweden.
Received November 7, 1980; accepted June 16, 1981.
4154
food intake and extra caloric demands by the tumor coupled
with frequent, vigorous therapy can usually explain the classi
cal outcome of cancer disease (cited from Ref. 42).
Monoexponential growth and rapid proliferation of normal
tissues occur in response to the stimulus of the killed anaerobic
bacterial vaccine, Corynebacterium parvum (6, 7, 27, 28).
Studies of the host organ response and the rapid normal cell
growth in such an experimental model may provide insight into
the growth of untransformed animal cells and its relationship to
the metabolic consequences of the intact host organism (27).
The aim of this study was to compare the metabolic altera
tions of the animal host in response to the rapid proliferation of
untransformed cells versus that of proliferation of malignant
cells in vivo. The stimulus of the killed bacterial vaccine, C.
parvum, was used for this purpose, as compared to the stimulus
of a methylcholanthrene-induced
sarcoma in C57BL/6J mice
(19-23).
MATERIALS
AND METHODS
were performed in metabolism cages, and in all experiments the
animals had free access to tap water. The experimental details in pairfeeding experiments have been described elsewhere (20, 22) and were
as described under "Determination
of Food and Water Intake and
Nitrogen Excretion."
Vaccine. C. parvum was provided by Burroughs Wellcome Labora
tory, Triangle Park, N. C. A formalin-killed
vaccine was used as
homogeneous suspension with the organisms (7 mg/ml, dry weight) in
0.9% NaCI solution. The solution contained 0.1% merthiolate as a
preservative. A dose of 0.05 ml or 350 ftg was given i.p. as reported
previously (27, 28). This dose provides strong stimuli to the C57BL
CANCER
RESEARCH
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VOL. 41
Metabolie Host Reaction of Nonmalignant
mouse. Control animals were given 0.9% NaCI solution as a sham
procedure. Merthiolate was not included, since this has been reported
to produce no detectable effects (14).
Tumor Model. A methylcholanthrene-induced sarcoma was used in
nongrowing 3-month-old mice. The results on metabolic tumor host
reactions, as referred to in "Discussion,"
have been reported else
where (19-23).
This sarcoma (MCG 101) has been transplanted
into
C57BL/6J
mice for more than 4 years in our laboratory. The tumor
grows extraperitoneally
per continuation when implanted s.c. in the
flanks of the animals and does not metastasize. The tumor-bearing
animals die with cachexia in 14 ± 1 (S.D.) days after tumor implanta
tion. The tumor dry weight is 10 to 14% of the carcass dry weight at
death. Tumor mass corresponded to about 7 to 8% of the body weight
of the sham-implanted controls (20).
Determination of Food and Water Intake and Nitrogen Excretion.
Metabolism cages (Jacoby, Stockholm, Sweden) slightly modified by
us for mouse experiments were used. One animal was kept in each
cage. All registrations were started, the tumor was implanted, and the
C. pan/urn vaccine was injected after the animals had spent 3 to 5 days
in the cages and reached constant body weight. During balance stud
ies, food was supplied at 8 a.m., 3 p.m., and 10 p.m. every day. The
survival time of tumor animals in the metabolism cages did not differ
from that of the tumor animals in ordinary cages with more than one
animal in each. Food and water intake were determined by weighing.
Urine was collected in the presence of NaN3 to prevent microbial
growth. Urine was pooled from Days 1 to 6 and from Days 7 to 12 in
the tumor-bearing animals, as described previously (20). Urine was
pooled over 9 days in the C. para/m-stimulated
mice along the prolif
eration of the spleen (Charts 1 and 2). Urea and ions were determined
as described previously (20).
Experimental
Procedures for in Vivo and in Vitro Studies.
[14C]Leucine (0.1 /¿Ci/gbody weight) was given i.p., and the incorpo
ration of label into hepatic and muscle proteins was determined as
described previously in detail (18, 21). Glycogen was extracted from
liver tissue and determined as described previously (24). Liver slices
(thickness, 0.4 mm) were prepared by use of a Mcllwain tissue chopper.
Incubations of liver tissue slices were performed at pH 7.4 at 37° in
Krebs-Ringer
bicarbonate
buffer solution, supplemented
formalin solution. Five-/mi sections were cut from paraffin-embedded
and methacrylate-embedded
material and stained with hematoxylin and
eosin.
Statistics.
U test was used for
RESULTS
The results given here refer to the results of the C. parvumstimulated animals, since most of the results of the tumor model
have been reported previously (19-23). Results from tumorbearing animals and their controls are referred to in "Discus
sion."
The time course of the food intake in C. parvum-stimulated
animals is shown in Chart 1. The time course of food intake in
pair-fed controls in relation to sarcoma-bearing mice has been
reported elsewhere (22). The body weight and the body com
position are shown in Table 1. C. pa/vum-treated animals had
decreased content of body lipids but unchanged lean body
mass (fat-free dry weight) and unchanged absolute content of
body water. The relative content of body water was, however,
increased. The liver and the spleen enlarged, which was due
to both increased dry weight and water content. The skeletal
muscle mass decreased with unchanged water content. These
alterations in organ weights were concomitant with increased
hepatic protein synthesis, increased hepatic synthesis of RNA
and DNA, but unchanged protein synthesis and content of RNA
in skeletal muscle (Table 2).
Increased activity of the lysosomal enzyme cathepsin D was
seen in the liver, skeletal muscle, and heart of C. parvumstimulated animals (Table 3). The altered protein metabolism in
12
Tumor
implantation
Chart 1. Time course of food intake in tumor-bearing mice, C. parvum-treated
mice, and controls. O, tumor-bearing mice (n - 14); •¿,
C. parvum-treated mice
(n - 7); A, controls (n - 14); oars. S.E.
Body composition was determined by drying the whole animal to
constant weight at 80°. Tbe lipids were then extracted by means of
chlorofornrmethanol
(1:1), ethanol:acetone
(1:1), and finally pure
ether. Chloroform, methanol, ethanol, acetone, and ether were of
analytical grade. The extracts obtained from different procedures were
pooled and then evaporated to dryness. The amount of lipids was
quantified by weighing with an accuracy of ±1 mg.
Histológica! Method. The livers and the spleens were fixed in 10%
1981
The nonparametric Mann-Whitney
statistical evaluation (38).
with glucose
(12 mmol/liters). The specific radioactivity in the incubation medium
was used for calculation of the incorporation rate of glucose carbon
into carbon dioxide. The incubation procedure and determination of
the radioactivity in carbon dioxide and in proteins were as described
previously (24). Determination of cathepsin D activity in liver, muscle,
and heart tissue has been described elsewhere (22). RNA was deter
mined according to the method of Munro and Fleck (31 ). Proteins were
determined according to the method of Lowry ef a/. (17) using bovine
serum albumin as the standard. Albumin concentration in serum was
determined by means of rocket immunoelectrophoresis
according to
the method of Laurell ef al. (15). Blood glucose was determined by the
glucose oxidase method (Boehringer Mannheim, Mannheim, West Ger
many). The tissue content of DMA was estimated according to the
method of Ceriotti (4), and the glycerol concentration in blood was
determined using commercially available kits from Boehringer Mann
heim.
Determination of Body Composition, Tissue Wet Weight, and
Tissue Dry Weight. Wet weight of the animals, the tumors, and the
spleen were determined in a standardized manner with an accuracy of
±1 mg. The water content of organs and tissues were determined by
drying to constant weight at 40°.
OCTOBER
versus Malignant Cells
Body composition
CPa
content
wt)69.9
(% of wet
±0.86' c
ControlsWater 65.8 ±0.9Dry
Table 1
in C. parvum-treated
mice
dry
(g)6.92wt
±0.1 5C
content(g)1.89
wt(g)5.03
±0.16C
±0.15
7.87 ±0.14Fat-free
5.23 ±0.16Lipid 2.64 ±0.19
CP. C pa/vum-treated mice.
b Mean ±S.E. of 11 animals.
c p < 0.01 (Mann-Whitney U test).
4155
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I. Karlberg et al.
Table 3
Cathepsin D activity in liver, skeletal, and heart muscle tissue from C. parvumtreated mice
IO0
Oó
¿imol/min/g protein
o•H
-HS
Skeletal muscle
N
O)O
tissueCP"Controls9.5 Liver tissue
±1.1b'c (10)"
Óssè^^co
5.1 ±0.5
(10)
2.4 ±0.
muscle
tissue8.8
±0.3°(20)
1.8 ±0.1C(10)
6.1 ±0.1 (20)
1 (10)Heart
" CP, C. parvum-treated mice.
6 Mean ±S.E.
c p < 0.001 (Mann-Whitney U test).
'' Numbers in parentheses, number of animals.
o•H
-HO
^Ti-
hepatic and skeletal muscle tissue (Table 4) was s"1Il£*QÂ
concomitant
i1a¥£è3.0.§w"i;S1i11'S1•S1o_2?i1Ã
with a decreased excretion of urea and decreased plasma
CJssCM
albumin concentration in the C. parvum-treated animals.
The hepatic incorporation rate of glucose carbon into carbon
dioxide was 3 to 4 times higher in C. parvum-treated mice
compared with controls, and the glycogen content was reduced
by 50%. The plasma concentration of blood glucose, glycerol,
and albumin was decreased in C. pa/vum-treated
animals
WCO
CO0
0•H
-HCM
r-.^
coh-
h-o
oo•»
(Table 5). Urinary excretion of urea, potassium, and sodium
was decreased (Table 6). The proliferation rate of the tumor
cells in sarcoma-bearing mice and that of the spleen cells in
the C. parvum-stimulated animals is shown in Chart 2.
The liver of the C. pa/vum-treated mice exhibited marked
histopathological changes. Small areas of focal necrosis of
liver parenchyma were seen along with signs of regeneration.
The regeneration was associated with a proliferation of hepatocytes showing an increased number of mitoses. A dense
inflammatory infiltrate composed mainly of lymphocytes and
histiocytes was seen, as well as small granulomas composed
of macrophages. The inflammatory infiltrate was more pro
nounced around the central veins and portal areas. There was
no apparent difference in the histological appearance of the
livers from the control animals and the tumor-bearing animals.
The histological appearance of the spleen from the C. pa/vumtreated and the tumor-bearing animals was similar, but the
increase in weight differed considerably. There was marked
proliferation of histocytes and macrophages in the red pulp,
along with frequent mitoses (Fig. 1). The C. pa/vum-treated
animals had abundant megakaryocytes and tumor-bearing an
imals had occasional megakaryocytes which was also seen in
the control mice. The size of the white pulp was markedly
reduced in the C. pa/vum-treated animals due to the prolifera
IO•H
-HSS^^
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KCM
hCM
0Ó•H
+((D
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tion of the macrophages and histiocytes. Such a reduction in
white pulp was not apparent among the tumor-bearing animals.
The main difference in the cellular morphology of the spleens
from C. parvum-treated and tumor-bearing animals was thus
quantitative (Fig. 1, A to D).
Twenty-one of 29 parameters analyzed in the C. parvum-
CM6
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Kn
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4156
stimulated animals showed a pattern of alteration to that found
in previous studies in sarcoma-bearing mice (Table 7). Statis
tically, there was no significant difference between the pattern
of alteration in the C. parvum model compared with appropriate
controls and the pattern found in tumor-bearing mice as com
pared to their appropriate controls.
In Table 8 results are shown from concomitant metabolic
studies in sarcoma-bearing mice, pair-fed mice, and controls
eating ad libitum. These experiments are given in order to
elucidate the possible role of decreased food intake in C.
parvum-treated animals. Results from pair-feeding experiments
CANCER
RESEARCH
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VOL. 41
Metabolie Host Reaction of Nonmalignant versus Malignant Cells
Table 4
Glycogen content, in vitro oxidation rate of ¡U-"C]glucose in liver tissue, and net in vivo incorporation of
l"C¡leucine into acid-precipitable proteins in liver and skeletal muscle tissue from C. parvum-treated mice
muscle
tissue[14C]Leucine
tissueOxidation
in
CP"
content
wt)10
(mg/g wet
±1.8ö-c(5)d
rate of
glucose (nmol/hr/
wt)1
g wet
33 ±7C (5)
incorpora
tion (dpm/mg protein/
4hr)1932
±168e (15)
37 ±3 (51("ClLeucine
1526 ±142
ControlsGlycogen 34 ±3.2
(5)Liver
a CP, C. parvum-treated mice.
6 Mean ±S.E.
0 p< 0.001 (Mann-Whitney U test).
d Numbers in parentheses, number of animals.
8 p < 0.05 (Mann-Whitney U test).
Plasma concentration
corporation
(dpm/
mghr)25
protein/4
(15)Skeletal
±3(15)
26 ±2(15)
Table 5
of glucose, glycerol, and albumin in C. parvum-treated
mice and in sarcoma-bearing mice
CPa
(mmol/liter)5.4±0.5b-c(11)d
(funol/liter)1.6
±0.1e (7)
(g/liter)15.4
±1.4'
ControlsMCG
7.3
(10)5.3
±0.6
21
(12)14.7
.4 ±0.9
±O.s'' 9 (12)
2.3
(7)1.1
±0.2
±0.1 "(9)
(12)
±0.5'- "(9)
ControlsGlucose 8.8 ±0.4
(17)Glycerol1.6 ±0.3 (8)Albumin
20.3 ±0.7
" CP, C. parvum-treated mice; MCG, sarcoma-bearing mice.
6 Mean ±S.E.
c p < 0.05 (Mann-Whitney U test).
" Numbers in parentheses, number of animals.
8 p < 0.01 (Mann-Whitney U test).
'p< 0.001 (Mann-Whitney U test).
9 Previously reported.
(8)
75
1 2 3456
Not previously reported.
Table 6
Urinary excretion of sodium, potassium, and urea from C. parvum-treated
over 9 days
mice
iimol/day/animal
Tumor
implantation
C.P»rvum
8 9 10 11 12 13 U 15 16 17
Days
injtction
Chart 2. Growth rate of the spleen in C. pa/vum-treated mice versus that of
the tumor in sarcoma-bearing mice. •¿,
spleen weight (n - 25; y - 7.85 x
eo46jo,; r = o 95). o tumor weigrlt („= so; y = 10.74 X e0"157", r = 0.97);
oars, S.E.
CPa
±17b'c
±18C
±155C
ControlsSodium254 320 ±13Potassium242 333 ±12Urea1468 2039 ±117
CP, C. parvum-treated mice.
0 Mean ±S.E. of 14 animals.
c p < 0.01 (Mann-Whitney U test).
on body composition, muscle dry weight, nitrogen balance,
oxidation rate of glucose carbons in vitro, and urinary excretion
of sodium, potassium, and urea among sarcoma-bearing mice,
pair-fed controls, and controls eating ad libitum have been
reported elsewhere (20, 22).
DISCUSSION
Exponential growth in vivo of transformed cells in an intact
animal has been established for experimental malignant tumors
(25, 26). It was reported previously that the killed anaerobic
bacterial vaccine from C. parvum induced rapid and exponen
tial proliferation of spleen and of various liver cells (6, 27). This
organ growth was due to increased DNA synthesis measured
by [3H]thymidine incorporation and an increase in DNA polymerase activity. The cell numbers of the spleen and the liver
increased exponentially over 10 days, and this condition was
completely reversible within approximately 4 weeks, which
differs from proliferation of malignant cells. The growth of
normal organ tissues in response to the challenge by a noxious
OCTOBER
1981
stimulus, in this case to the bacterial antigenicity, was thought
of as a potentially useful experimental model aimed at further
elucidating the degree of specificity and possibly the mecha
nisms behind cancer cachexia. This assumption was based on
the reported conclusion that "the C. parvum experimental
model may provide insight into the growth and proliferation
response for normal tissues in the intact organism" (27).
In this study, C. parvum vaccine was given i.p. in order to
evaluate the metabolic host reaction in response to exponential
growth of nonmalignant tissues as compared to the metabolic
host reaction in response to exponential growth of malignant
cells as illustrated in Chart 2. The rationale for this was to test
whether the host reactions in tumor-bearing organisms are
essentially due to a proliferation of malignant cells (19-23) or
to a proliferation of nonmalignant cells as well (10, 29, 37).
The tumor reactions in our animal model were almost identical
with corresponding changes found in studies on cancer pa
tients with various malignant tumors (19). The variables ana
lyzed in this study were selected among those found to be of
significance in the tumor-bearing mice, according to our pre
vious studies (19-23).
In this study, 29 parameters were analyzed, and 21 of these
showed the same pattern of alteration in C. parvum-treated
animals as that found in tumor-bearing animals. Statistical
evaluation showed that the pattern of reactions did not differ
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/. Karlberg et al.
between the 2 models. This shows that most metabolic altera
tions in our sarcoma-bearing mice are not specific for tumor
growth and can perhaps be explained by changes associated
with cell proliferation but not necessarily by proliferation of
malignant cells.
Table 7
Relative alterations in various metabolic parameters in C. parvum-treated
and in sarcoma-bearing mice compared with appropriate controls
mice
The pattern of alteration in C. parvum-treated animals did not differ statistically
significantly from that in sarcoma-bearing animals as evaluated by frequency
analysis. (Each parameter which was altered changed in the same direction in C.
parvum and sarcoma-bearing animals compared with control levels.)
Relative alteration vs. appropriate
controls
ParametersTotal
wtTotalbody
contentLipid
body water
storesLean
massFood
body
The metabolic alterations in C. parvum-treated animals could
hypothetical^
be explained by the decrease in food intake
similar to that found in the tumor animals (20). However, the
metabolic reactions in our tumor-bearing mice were essentially
not ascribed to the decrease in food intake (anorexia) itself, as
shown in previous studies (20, 22, 23) and in Table 8. Thus
far, we have found that the anorexia itself can to some extent
explain quantitatively and qualitatively the decreased protein
synthesis in skeletal muscles of sarcoma-bearing mice (23). In
contrast to this, C. parvum-stimulated
mice had increased
incorporation of [14C]leucine into muscle proteins. The precise
relationship between incorporation rate of [14C]leucine into
muscle proteins and the net protein imbalance in the skeletal
muscles of C. parvum-treated mice was not determined. We
conclude from pair-feeding experiments, in relation to sar
coma-bearing mice, that altered metabolic activity in C. par
vum-treated mice was not explained by their anorexia per se.
The hepatic metabolism showed essentially the same bio
chemical alterations in C. parvum-treated animals and the
tumor-bearing animals, with the exception of one fundamental
difference. In C. parvum-treated animals, there was a marked
cell proliferation, leading to an increase in the organ weight,
whereas liver weight and its microscopic appearance did not
tissueGlucose
content in liver
show evidence of alterations in the tumor host. This difference
tissueProtein
oxidation in liver
is of major importance, since it suggests that the metabolic
contentLiver
response in tumor host was not due to any major proliferation
tissueSkeletal
tissueProteinmuscle
of the same tissues as in the C. parvum-treated animals with
vivoLiver
synthesis in
the exception of the spleen. However, in terms of weight, the
tissueSkeletal
proliferation was confined mainly to malignant cells in tumortissueRNA muscle
synthesisLiver
bearing mice (20). The hepatic metabolic alterations in C.
tissueSkeletal
parvum-treated animals are consistent with increased hepatic
tissueDNA muscle
tissueCathepsin
content in liver
protein turnover coupled to cell proliferation, as well as in
activityLiver
D
creased activity of reticuloendothelial cells (2). The decreased
tissueSkeletal
hepatic content of glycogen and the decreased blood glucose
tissueHeart
muscle
concentration in C. parvum-treated animals indicated that the
tissuePlasma
muscle
concentrationGlucoseGlycerolAlbuminUrinary
synthesis and the exogenous supply of glucose was insufficient
to meet the total energy demand of the proliferating cells in the
C. parvum-treated animals, as it was in various tumor animal
excretionSodiumPotassiumUreainCP"UnchangedIncreased"Decreased0UnchangedDecreased0Increased0Decreased0Increased0Increased0UnchangedIncreased0Decreased0Increased0Increas
models (19, 20, 36). This suggestion is further supported by
low plasma concentrations of glycerol in both C. parvumtreated and tumor animals but not in pair-fed controls and on
a CP, C. parvum-treated mice; MCG. sarcoma-bearing mice.
the finding that body lipid stores were decreased, indicating a
6 p < 0.01 (Mann-Whitney U test).
0 p < 0.001 (Mann-Whitney U test),
net lipolysis in both conditions.
p < 0.05 (Mann-Whitney U test).
The liver and the spleen were the organs that significantly
intakeLiver
wtSkeletal
dry
wtSpleen muscle dry
wtWater dry
contentLiver
tissueSkeletal
tissueSpleenGlycogen
muscle
Table 8
Plasma glucose and glycerol, hepatic content of glycogen and RNA, incorporation of [U-"C¡leucÃ-ne into hepatic proteins in vivo, and
cathepsin D activity in heart tissue from sarcoma-bearing mice, pair-fed controls, and controls eating ad libitum
These results were from strictly matched pair-feeding experiments as described in "Material and Methods," with one animal in each
metabolism cage.
("CJIeucine
glucose
(mmol/liter)4.64
±0.32°(11)°
1.2.3.MCG"Pair-fed
glycerol
Qimol/liter)1.13
±0.12(9)
6.31 ±0.40 (12)
1.71 ±0.29(7)
Controls
7.13 ±0.70 (10)
1.56 ±0.33(8)
1 vs. 2a
p < 0.01
p < 0.08
2 vs. 3°
NS
NS
1 vs. 3dPlasma
p < 0.001Plasma
NSHepatic
a MCG, sarcoma-bearing mice; NS. not significant.
6 Mean ±S.E.
0 Numbers in parentheses, number of animals.
" Mann-Whitney U test.
4158
glycogen
(mg/g)7.53
content
±2.89 (8)
22.5 ±5.2 (9)
21 .5 ±5.5 (8)
p < 0.025
NS
p < 0.025Hepatic
RNA con
(mg/g)12.1
tent
of
into
hepatic proteins
(dpm/mg protein/
4hr)1665
±0.3
(6)
±120(12)
1115 ±123(17)
8.92 ±0.17(20)
1502 ± 61 (13)
8.8 ±0.2 (23)
p < 0.001
p < 0.01
p < 0.025
NS
p < 0.001Incorporation p<0.10cathepsin
D activ
ity in heart tissue
(nmol tyrosine/
min/mg
protein)8.69
7.65
7.46
p
p
p
CANCER
±0.32 (4)
±0.08(7)
±0.05 (7)
< 0.01
< 0.05
< 0.01
RESEARCH
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VOL.
41
Metabolie Host Reaction of Nonmalignant
increased in weight in C. pa/vum-treated animals due to cell
proliferation. This corresponded to a net increase of 220 to
230 mg dry organ weight but was concomitant with a decrease
in total body dry weight, suggesting a negative energy balance
in the total animal. The skeletal muscle mass lost 180 to 190
mg, dry weight, in the C. pan/um-treated animals and in the
tumor-bearing animals (20). The protein imbalance of skeletal
muscles could thus supply essentially all the nitrogen neces
sary for cell proliferation in the liver and spleen in the C.
parvum-treated animals and about 50% of the tumor mass (20).
The fractional loss of the carcass dry weight was the same in
C. pa/vum-treated animals and tumor-bearing animals, while
pair-fed animals maintained proteins and lipids (20). The loss
of lipids concomitant with a pronounced cell proliferation sug
gests an increased energy demand in the C. parvum-treated
animals.
We do not think that this model, in its present form, is ideal
for studies of proliferating nonmalignant cells in vivo. The
antigenic burden may have been unnecessarily high (6), as
indicated by the more pronounced alteration of several vari
ables in the C. para/m-treated animals compared with tumor
mice and by the histological appearance of the liver of C.
pa/vum-treated
mice (Fig. 1). It may be desirable to make
adjustments in this respect, by modifying the administered
dose (6), when mechanistic studies are to be performed.
The role of C. parvum and of tumor in their selective effects
of cell proliferation cannot be decided from this study. They
may cause a metabolic alteration or product, which may cause
selective tissue response to release Gi-arrested cells into cell
proliferation, by as yet unidentified factors (5, 32). Our results
are consistent with the following hypothesis. Normal cells are
stimulated by some mechanisms, probably by an antigen(s), to
rapid proliferation when exposed to C. parvum vaccine (3, 10,
28). This cell proliferation may be associated with a change in
the enzyme equipment of various cells (39, 40) and with a
substrate demand as found in proliferating malignant and em
bryonic cells (12, 34, 44). This may produce adequate enzyme
alteration(s) to guarantee optimal cell growth. It was reported
previously that the growth state of adult hepatocytes in primary
monolayer culture was dependent on phenotypes (16). The Lisozymic form of pyruvate kinase and the glutathione S-transferase B decreased more than 70 to 80% during the restingto-growing transitions but increased abruptly as proliferative
rates declined. Conversely, the levels of fetal functions, such
as the K-isozymic form of pyruvate kinase and a-fetoprotein
production increased during the exponential growth (16).
Moreover, regenerating liver cells after partial hepatectomy
synthesized pyruvate kinase type III, a characteristic of fetal
liver or transformed cells, instead of type I. Regenerating tissue
decreased its content of the adult enzyme form glucokinase
but not of the Iow-Km hexokinases (1), which is a pattern in
common with transformed cells.
We suggest that several metabolic host reactions in tumorbearing animals, and perhaps also in cancer patients, may be
directly coupled to cell proliferation per se but not necessarily
to the proliferation of malignant cells.
ACKNOWLEDGMENTS
Dr. P. Wahlen. Department of Bacteriology, University of Göteborg, is acknowl
edged for valuable advice concerning the C. parvum vaccine.
OCTOBER
1981
versus Malignant Cells
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CANCER
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VOL.
41
Metabolie Host Reaction of Nonmalignant
versus Malignant Cells
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Metabolic Host Reaction in Response to the Proliferation of
Nonmalignant Cells versus Malignant Cells in Vivo
Ingvar Karlberg, Staffan Edström, Lars Ekman, et al.
Cancer Res 1981;41:4154-4161.
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