0013-7227/78/1031-0223$02.00/0 Endocrinology Copyright © 1978 by The Endocrine Society
Vol. 103, No. 1 Printed in U.S.A.
Growth Response in Athymic Nude Mice to Transplanted Growth Hormone-Secreting Rat Pituitary Tumor Cells* SEUNG-IL SHIN.f ANDRE L. BROWN, AND F. CARTER BANCROFT Department of Genetics, Albert Einstein College of Medicine (S.S., A.L.B.), Bronx, New York 10461; Memorial Sloan-Kettering Cancer Center (F.C.B.), New York, New York 10021 ABSTRACT. Clonal cell lines originally derived from a somatotropic rat pituitary tumor were grafted sc into athymic nude mice, and the growth response induced in the graft carriers was examined. When at least 10s rat tumor cells were injected per mouse, virtually all mice developed slow growing, nonmetastasizing tumors at the injection site. In immature nude mice, the somatotropic effect of the rat GH secreted by the growing tumor was clearly evident, as body weight continued to increase well beyond that of control littermates. Nude mice with a final body weight of over 60 g were regularly produced in this manner. The action of the rat GH in the nude mouse carriers was generalized, as shown by the fact that the gigantic mice resulting from the xenografts maintained apparently normal balanced physical pro-
S
INCE the first demonstration by Rygaard and Povlsen (1) that the genetic absence of thymus-dependent immunity can lead to successful xenografts of human tumors in the nude mouse, this mouse mutant has emerged as a useful tool for the development of endocrine tumor lines from a wide variety of tissues and species (2). In addition to providing a convenient model system for analysis of hormone-mediated control mechanisms in vivo, stable endocrine tumor lines are valuable sources for the isolation of functional cell culture lines for quantitative studies on hormone synthesis, secretion, and regulation (3-5). In a previous report, it has been shown that
portions. Exceptionally, however, the islets of Langerhans in the host pancreas showed even greater hypertrophy, presumably due to the known anti-insulin activity of GH. The rat pituitary tumor cells were shown to synthesize and secrete a protein which was precipitated by an antiserum against rat GH and which comigrated with authentic rat GH when analyzed by sodium dodecyl sulfate-acrylamide electrophoresis. Through serial transplantation in nude mice, the tumor cells retained their functional and tumorigenic properties. These results demonstrate that rat GH is active in the mouse, and suggest also that the athymic nude mouse should be useful for the development of other xenogeneic tumor lines that secrete GH. (Endocrinology 103: 223, 1978)
a highly functional Syrian hamster tumor of the pancreatic islet cells that secretes insulin can be maintained as a serial transplant in nude mice without the loss of the insulin-secreting capacity (6). However, the development of reliable procedures for the establishment of highly differentiated human endocrine tumor lines in nude mice has yet to be achieved (7). A major goal in our work with nude mice has been the establishment, through serial xenografts, of a functionally stable pituitary tumor line of human origin that would continue to synthesize human GH. In an effort to optimize the technical procedures necessary for this purpose, we have investigated the host response in nude mice to xenografts of a highly malignant, GH-secreting rat pituitary tumor. Part of these results have been presented recently in two meetings on nude mice (7, 8).
Received October 21, 1977. Address requests for reprints to: Dr. S. Shin, Department of Genetics, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, New York 10461. * This work was supported by research grants from the National Science Foundation (PCM-76-07573 and PCMMaterials and Methods 74-19373), a National Cancer Institute Core Grant (CA08748) to Memorial Sloane-Kettering Cancer Center, a Rat pituitary cell lines research grant from the NIH (CA-08748), a Genetics Center Grant (GM-19100), and a Diabetes Research and The rat pituitary tumor cell line, GH3, was origTraining Center Grant (AM-20541) to Albert Einstein inally established by Tashjian and coworkers (9, College of Medicine. 10) from a transplantable somatotropic tumor t Recipient of a Faculty Research Award from the maintained in Wistar/Furth rats (11). GH3 cells American Cancer Society. 223
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224
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Endo • 1978 Vol 103 • No 1
synthesize both rat GH (rGH) and PRL in culture (10). Cell line GC is a subclone isolated from GH3, which synthesizes increased amounts of rGH but little or no detectable PRL (12). Extensive analysis has shown that the material produced by GH3 and GC cells in culture is indistinguishable from authentic rGH (9, 10, 12). Both cell lines were maintained in suspension culture in Joklik's modified Eagle's medium (Grand Island Biological Co., NY), supplemented with 15% horse serum and 2.5% fetal bovine serum, plus penicillin (100 U/ml) and streptomycin (100 /xg/ml).
continued for 2 h and tumor cells were pelleted by centrifugation for 5 min at 800 X g. The supernatant was then subjected to immunoprecipitation with a baboon antiserum to rGH, as previously described (15). The recovered precipitate was dissociated in a buffer containing sodium dodecyl sulfate (SDS) and electrophoresed on a 13-cm-long discontinuous SDS-acrylamide gel containing 15% acrylamide (16). An internal 35S-labeled GH marker, prepared by incubation of GC cells with [35S]methionine followed by immunoprecipitation of the cytoplasm with anti-rGH serum (16), was run on the same gel.
Nude mice
Measurement of GH levels in mouse serum
The nude mouse colony used for this study was established originally from a breeding stock that had been backcrossed four generations with BALB/c (13). The colony was maintained in an isolated semiaseptic environment by the use of laminar-flow cage racks. Under our breeding conditions, nude mice generally survive longer than 8 months, and often over 18 months.
Sera, collected from control and tumor-carrying nude mice by heart puncture, were kindly assayed by Dr. Armen H. Tashjian, Jr., for immunoreactive rGH by the microcomplement fixation technique using a baboon antiserum to rGH (17).
Xenograft of rat pituitary cells in nude mice Cells were harvested from suspension culture by centrifugation, counted for viable cells with trypan blue, and resuspended in sterile physiological saline at a final concentration of 107 cells/ml. Using sterile 21-gauge needles fitted on 1-ml disposable sterile syringes, 0.2-ml aliquots (2 X 106 cells) were injected sc on the flank. Palpable tumor nodules usually appeared at the injection site within 2-4 weeks. For serial transplantation, the tumor was excised, freed of capsular material and any necrotic tissue, and minced with fine scissors after the addition of an equal volume of serum-free tissue culture medium. Aliquots containing 2 x 106 tumor cells were inoculated into nude mice, as above. Reestablishment of tumor cells in culture has been described previously (14). Measurement of GH synthesis by excised tumor cells A tumor grown in nude mice was excised, and the capsular material and any necrotic tissue were trimmed and discarded. A fragment of the tumor (0.2-0.5 g) was placed in 1.0 ml complete growth medium, containing 1/20 the normal concentration of leucine. After extensive mincing of the fragment with a scalpel, the mince was incubated for 15 min at 37 C in a small tube containing a rotating magnetic bar. After addition of 50 /xCi [3H]leucine (New England Nuclear Co.; 57 Ci/mmol), incubation was
Results Every nude mouse injected with GH3 or GC cells, or with tumor cell suspensions prepared from the first generation xenografts, developed progressively growing tumors at the injection site, regardless of the sex or the age of the mice (2 weeks to 5 months after birth; Table 1). Macroscopically detectable metastasis or spontaneous regression of the tumor was never observed. This observation is consistent with our earlier data regarding other highly tumorigenic cells (6, 13, 14). If left undisturbed, nude mice carrying GH3 tumors often survived for over 4 months until the tumor weight exceeded 5 g. The somatotropic effect of the rat pituitary tumor cells on the tumor carriers was dramatically evident when immature nude mice were used as recipients. The body weight of mice carrying a growing GH3 tumor continued to increase well beyond the normal adult limits, so that nude mice weighing over 60 g were regularly produced. The results of a matched series, in which 2 X 106 GH3 cells were injected into immature 21-day-old nude mice, are presented in Fig. 1. Control mice matched for age and sex were injected with saline. It can be seen that the control mice reached the normal adult body weight (32 g) by about 65 days of age, but
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GH-SECRETING RAT CELLS IN NUDE MICE
225
TABLE 1. Tumor induction in nude mice by rat pituitary tumor cells Recipient mice Exp no.
Pituitary tumor cell clone
No. cells injected
Age when injected (days)
Incidence" of take
Latency* period (days)
3 x 10" 2/2 M 49 10 2/2 27 10 3 x 10" F 2/2 2 27 13 2 x 10" M 2/2 13 2X 10" 47 or 57 F GH 3 4/4 3 10 1 x 10" 28, 31, or 55 F M 2/2 4 30 2 X 107rf GH 3 /NuT c 34 and 51 In exp 1-3, washed single cells collected from suspension culture were injected sc as described in Materials and Methods. In exp 4, tumor cell preparations obtained from a tumor produced by injection of GH3 cells in exp 3 were used immediately after removal from the mouse. " Number of mice which developed growing tumors per number of mice inoculated. * Number of days postimplantation when palpable tumor first appeared. c GH3 tumor grown in nude mouse. rf Total cells including leukocytes but excluding red blood cells. 1
5
GC GC GH3 GH3
TABLE 2. Growth response to rat pituitary tumor cells transplanted in nude mice
60
£ 50 £ 40
Mouse no.
% 30 o m 20
Sex
< o
10 Ql
I I I I I I I I I I I I I I I I I I I I I I I I
0
10 20 30 40 50 60 70 80 90 100 110 120 AGE OF MICE (days)
FIG. 1. Somatotropic response in nude mice to a GHsecreting rat pituitary tumor cell transplant: time course of weight gain. The weight increases for the three control nude mice, which had reached the normal adult limits by day 65, were not followed after day 100. One of the three nude mice carrying the GH3 tumor died on day 110 with a large tumor. The two remaining tumor carriers (no. 1353 and 1354) were sacrificed on day 112 to measure the weights of various organs and to collect sera for rGH assay, as shown in Tables 2 and 3.
their littermates, carrying a growing GH3 tumor (average tumor weight, about 3 g), continued to grow until nearly twice the normal body weight was attained. The enormous increase in body weight in tumor-carrying nude mice was accompanied by proportional increases in the blood volume and in the sizes of all of the major organs that we have measured, so that even the largest mice appeared to retain normal physical proportions. These data are summarized in Table 2. In addition to the generalized somatotropic effects, the tumor-bearing nude mice showed
Control 1368 M None
Exp 1
Exp 2
1353
1354 M GH;,
M
Cells injected (2 x 10") GH., 21 21 21 Age at injection (days) 112 112 112 Age at termination (days) 0 4.1 5.0 Weight of tumor (g) Weight of mouse less 32.8 51.9 51.3 tumor wt (g) 1.0 2.6 2.6 Total recovered blood (ml)" Wet wt of organs (g) 0.9 0.7 1.2 Kidneys 5.0 2.0 4.9 Liver 0.2 0.5 0.4 Heart 0.2 Spleen 0.9 0.6 Body length (mm) 195 225 220 Tail tip to nose 75 90 90 Crown to rump " Total blood volume recovered from heart with a 26gauge needle.
another striking consequence of the heterografted endocrine tumor. It is known that GH has a strong anti-insulin activity, and as a result, human acromegalic patients often develop a compensatory hyperplasia of the islets of Langerhans (18). An identical response is induced in nude mice bearing a growing GH3 tumor. The entire pancreas is enlarged, but the islets of Langerhans are especially large and more numerous compared to the control littermates. The average size of the secretory cells within the islets is also much larger in
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226
SHIN, BROWN, AND BANCROFT
the tumor carriers. Low magnification micrographs of stained sections from the pancreas of a control and a GH3 tumor carrier at an identical magnification are shown in Fig. 2. It can be seen that the pancreatic islets of the mouse are specifically stimulated by the heterologous GH secreted by the rat pituitary tumor implant. That the somatotropic response demonstrated in the nude mice implanted with GH3 and GC cells is in fact due to rGH synthesized and secreted by the rat tumor cells has been supported more directly by the following observations. Blood samples (0.5-0.8 ml) were withdrawn from the heart of three nude mice carrying growing GH3 tumors and allowed to clot in glass test tubes for 4 h at room temperature; the sera were collected by centrifuga-
Endo • 1978 Vol 103 • No 1
tion in the cold. Aliquots of the frozen sera were stored at —20 C and subsequently analyzed for rGH levels by the microcomplement fixation technique. Serum collected from a nude mouse without a tumor served as a control. The results are given in Table 3. All of TABLE 3. Serum levels of immunoreactive GH in nude mice bearing rat pituitary tumor transplants
Mouse
1276 1353" 1354" 1565 Identical
Tumor transplant None GHa GH3 GH 3
Age of mouse when sacrificed (days)
Tumor wt (g)
Serum GH level (jig/ml)
42 112 112 107
0 4.1 5.0 3.5
Vol. 103, No. 1 Printed in U.S.A.
Growth Response in Athymic Nude Mice to Transplanted Growth Hormone-Secreting Rat Pituitary Tumor Cells* SEUNG-IL SHIN.f ANDRE L. BROWN, AND F. CARTER BANCROFT Department of Genetics, Albert Einstein College of Medicine (S.S., A.L.B.), Bronx, New York 10461; Memorial Sloan-Kettering Cancer Center (F.C.B.), New York, New York 10021 ABSTRACT. Clonal cell lines originally derived from a somatotropic rat pituitary tumor were grafted sc into athymic nude mice, and the growth response induced in the graft carriers was examined. When at least 10s rat tumor cells were injected per mouse, virtually all mice developed slow growing, nonmetastasizing tumors at the injection site. In immature nude mice, the somatotropic effect of the rat GH secreted by the growing tumor was clearly evident, as body weight continued to increase well beyond that of control littermates. Nude mice with a final body weight of over 60 g were regularly produced in this manner. The action of the rat GH in the nude mouse carriers was generalized, as shown by the fact that the gigantic mice resulting from the xenografts maintained apparently normal balanced physical pro-
S
INCE the first demonstration by Rygaard and Povlsen (1) that the genetic absence of thymus-dependent immunity can lead to successful xenografts of human tumors in the nude mouse, this mouse mutant has emerged as a useful tool for the development of endocrine tumor lines from a wide variety of tissues and species (2). In addition to providing a convenient model system for analysis of hormone-mediated control mechanisms in vivo, stable endocrine tumor lines are valuable sources for the isolation of functional cell culture lines for quantitative studies on hormone synthesis, secretion, and regulation (3-5). In a previous report, it has been shown that
portions. Exceptionally, however, the islets of Langerhans in the host pancreas showed even greater hypertrophy, presumably due to the known anti-insulin activity of GH. The rat pituitary tumor cells were shown to synthesize and secrete a protein which was precipitated by an antiserum against rat GH and which comigrated with authentic rat GH when analyzed by sodium dodecyl sulfate-acrylamide electrophoresis. Through serial transplantation in nude mice, the tumor cells retained their functional and tumorigenic properties. These results demonstrate that rat GH is active in the mouse, and suggest also that the athymic nude mouse should be useful for the development of other xenogeneic tumor lines that secrete GH. (Endocrinology 103: 223, 1978)
a highly functional Syrian hamster tumor of the pancreatic islet cells that secretes insulin can be maintained as a serial transplant in nude mice without the loss of the insulin-secreting capacity (6). However, the development of reliable procedures for the establishment of highly differentiated human endocrine tumor lines in nude mice has yet to be achieved (7). A major goal in our work with nude mice has been the establishment, through serial xenografts, of a functionally stable pituitary tumor line of human origin that would continue to synthesize human GH. In an effort to optimize the technical procedures necessary for this purpose, we have investigated the host response in nude mice to xenografts of a highly malignant, GH-secreting rat pituitary tumor. Part of these results have been presented recently in two meetings on nude mice (7, 8).
Received October 21, 1977. Address requests for reprints to: Dr. S. Shin, Department of Genetics, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, New York 10461. * This work was supported by research grants from the National Science Foundation (PCM-76-07573 and PCMMaterials and Methods 74-19373), a National Cancer Institute Core Grant (CA08748) to Memorial Sloane-Kettering Cancer Center, a Rat pituitary cell lines research grant from the NIH (CA-08748), a Genetics Center Grant (GM-19100), and a Diabetes Research and The rat pituitary tumor cell line, GH3, was origTraining Center Grant (AM-20541) to Albert Einstein inally established by Tashjian and coworkers (9, College of Medicine. 10) from a transplantable somatotropic tumor t Recipient of a Faculty Research Award from the maintained in Wistar/Furth rats (11). GH3 cells American Cancer Society. 223
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224
SHIN, BROWN, AND BANCROFT
Endo • 1978 Vol 103 • No 1
synthesize both rat GH (rGH) and PRL in culture (10). Cell line GC is a subclone isolated from GH3, which synthesizes increased amounts of rGH but little or no detectable PRL (12). Extensive analysis has shown that the material produced by GH3 and GC cells in culture is indistinguishable from authentic rGH (9, 10, 12). Both cell lines were maintained in suspension culture in Joklik's modified Eagle's medium (Grand Island Biological Co., NY), supplemented with 15% horse serum and 2.5% fetal bovine serum, plus penicillin (100 U/ml) and streptomycin (100 /xg/ml).
continued for 2 h and tumor cells were pelleted by centrifugation for 5 min at 800 X g. The supernatant was then subjected to immunoprecipitation with a baboon antiserum to rGH, as previously described (15). The recovered precipitate was dissociated in a buffer containing sodium dodecyl sulfate (SDS) and electrophoresed on a 13-cm-long discontinuous SDS-acrylamide gel containing 15% acrylamide (16). An internal 35S-labeled GH marker, prepared by incubation of GC cells with [35S]methionine followed by immunoprecipitation of the cytoplasm with anti-rGH serum (16), was run on the same gel.
Nude mice
Measurement of GH levels in mouse serum
The nude mouse colony used for this study was established originally from a breeding stock that had been backcrossed four generations with BALB/c (13). The colony was maintained in an isolated semiaseptic environment by the use of laminar-flow cage racks. Under our breeding conditions, nude mice generally survive longer than 8 months, and often over 18 months.
Sera, collected from control and tumor-carrying nude mice by heart puncture, were kindly assayed by Dr. Armen H. Tashjian, Jr., for immunoreactive rGH by the microcomplement fixation technique using a baboon antiserum to rGH (17).
Xenograft of rat pituitary cells in nude mice Cells were harvested from suspension culture by centrifugation, counted for viable cells with trypan blue, and resuspended in sterile physiological saline at a final concentration of 107 cells/ml. Using sterile 21-gauge needles fitted on 1-ml disposable sterile syringes, 0.2-ml aliquots (2 X 106 cells) were injected sc on the flank. Palpable tumor nodules usually appeared at the injection site within 2-4 weeks. For serial transplantation, the tumor was excised, freed of capsular material and any necrotic tissue, and minced with fine scissors after the addition of an equal volume of serum-free tissue culture medium. Aliquots containing 2 x 106 tumor cells were inoculated into nude mice, as above. Reestablishment of tumor cells in culture has been described previously (14). Measurement of GH synthesis by excised tumor cells A tumor grown in nude mice was excised, and the capsular material and any necrotic tissue were trimmed and discarded. A fragment of the tumor (0.2-0.5 g) was placed in 1.0 ml complete growth medium, containing 1/20 the normal concentration of leucine. After extensive mincing of the fragment with a scalpel, the mince was incubated for 15 min at 37 C in a small tube containing a rotating magnetic bar. After addition of 50 /xCi [3H]leucine (New England Nuclear Co.; 57 Ci/mmol), incubation was
Results Every nude mouse injected with GH3 or GC cells, or with tumor cell suspensions prepared from the first generation xenografts, developed progressively growing tumors at the injection site, regardless of the sex or the age of the mice (2 weeks to 5 months after birth; Table 1). Macroscopically detectable metastasis or spontaneous regression of the tumor was never observed. This observation is consistent with our earlier data regarding other highly tumorigenic cells (6, 13, 14). If left undisturbed, nude mice carrying GH3 tumors often survived for over 4 months until the tumor weight exceeded 5 g. The somatotropic effect of the rat pituitary tumor cells on the tumor carriers was dramatically evident when immature nude mice were used as recipients. The body weight of mice carrying a growing GH3 tumor continued to increase well beyond the normal adult limits, so that nude mice weighing over 60 g were regularly produced. The results of a matched series, in which 2 X 106 GH3 cells were injected into immature 21-day-old nude mice, are presented in Fig. 1. Control mice matched for age and sex were injected with saline. It can be seen that the control mice reached the normal adult body weight (32 g) by about 65 days of age, but
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GH-SECRETING RAT CELLS IN NUDE MICE
225
TABLE 1. Tumor induction in nude mice by rat pituitary tumor cells Recipient mice Exp no.
Pituitary tumor cell clone
No. cells injected
Age when injected (days)
Incidence" of take
Latency* period (days)
3 x 10" 2/2 M 49 10 2/2 27 10 3 x 10" F 2/2 2 27 13 2 x 10" M 2/2 13 2X 10" 47 or 57 F GH 3 4/4 3 10 1 x 10" 28, 31, or 55 F M 2/2 4 30 2 X 107rf GH 3 /NuT c 34 and 51 In exp 1-3, washed single cells collected from suspension culture were injected sc as described in Materials and Methods. In exp 4, tumor cell preparations obtained from a tumor produced by injection of GH3 cells in exp 3 were used immediately after removal from the mouse. " Number of mice which developed growing tumors per number of mice inoculated. * Number of days postimplantation when palpable tumor first appeared. c GH3 tumor grown in nude mouse. rf Total cells including leukocytes but excluding red blood cells. 1
5
GC GC GH3 GH3
TABLE 2. Growth response to rat pituitary tumor cells transplanted in nude mice
60
£ 50 £ 40
Mouse no.
% 30 o m 20
Sex
< o
10 Ql
I I I I I I I I I I I I I I I I I I I I I I I I
0
10 20 30 40 50 60 70 80 90 100 110 120 AGE OF MICE (days)
FIG. 1. Somatotropic response in nude mice to a GHsecreting rat pituitary tumor cell transplant: time course of weight gain. The weight increases for the three control nude mice, which had reached the normal adult limits by day 65, were not followed after day 100. One of the three nude mice carrying the GH3 tumor died on day 110 with a large tumor. The two remaining tumor carriers (no. 1353 and 1354) were sacrificed on day 112 to measure the weights of various organs and to collect sera for rGH assay, as shown in Tables 2 and 3.
their littermates, carrying a growing GH3 tumor (average tumor weight, about 3 g), continued to grow until nearly twice the normal body weight was attained. The enormous increase in body weight in tumor-carrying nude mice was accompanied by proportional increases in the blood volume and in the sizes of all of the major organs that we have measured, so that even the largest mice appeared to retain normal physical proportions. These data are summarized in Table 2. In addition to the generalized somatotropic effects, the tumor-bearing nude mice showed
Control 1368 M None
Exp 1
Exp 2
1353
1354 M GH;,
M
Cells injected (2 x 10") GH., 21 21 21 Age at injection (days) 112 112 112 Age at termination (days) 0 4.1 5.0 Weight of tumor (g) Weight of mouse less 32.8 51.9 51.3 tumor wt (g) 1.0 2.6 2.6 Total recovered blood (ml)" Wet wt of organs (g) 0.9 0.7 1.2 Kidneys 5.0 2.0 4.9 Liver 0.2 0.5 0.4 Heart 0.2 Spleen 0.9 0.6 Body length (mm) 195 225 220 Tail tip to nose 75 90 90 Crown to rump " Total blood volume recovered from heart with a 26gauge needle.
another striking consequence of the heterografted endocrine tumor. It is known that GH has a strong anti-insulin activity, and as a result, human acromegalic patients often develop a compensatory hyperplasia of the islets of Langerhans (18). An identical response is induced in nude mice bearing a growing GH3 tumor. The entire pancreas is enlarged, but the islets of Langerhans are especially large and more numerous compared to the control littermates. The average size of the secretory cells within the islets is also much larger in
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226
SHIN, BROWN, AND BANCROFT
the tumor carriers. Low magnification micrographs of stained sections from the pancreas of a control and a GH3 tumor carrier at an identical magnification are shown in Fig. 2. It can be seen that the pancreatic islets of the mouse are specifically stimulated by the heterologous GH secreted by the rat pituitary tumor implant. That the somatotropic response demonstrated in the nude mice implanted with GH3 and GC cells is in fact due to rGH synthesized and secreted by the rat tumor cells has been supported more directly by the following observations. Blood samples (0.5-0.8 ml) were withdrawn from the heart of three nude mice carrying growing GH3 tumors and allowed to clot in glass test tubes for 4 h at room temperature; the sera were collected by centrifuga-
Endo • 1978 Vol 103 • No 1
tion in the cold. Aliquots of the frozen sera were stored at —20 C and subsequently analyzed for rGH levels by the microcomplement fixation technique. Serum collected from a nude mouse without a tumor served as a control. The results are given in Table 3. All of TABLE 3. Serum levels of immunoreactive GH in nude mice bearing rat pituitary tumor transplants
Mouse
1276 1353" 1354" 1565 Identical
Tumor transplant None GHa GH3 GH 3
Age of mouse when sacrificed (days)
Tumor wt (g)
Serum GH level (jig/ml)
42 112 112 107
0 4.1 5.0 3.5
