Vol.
177,
No.
May
31, 1991
GENE
1, 1991
BIOCHEMICAL
AND
BIOPHYSICAL
RQEARCH
COMMUNICATIONS Pages
EXPRESSION
RECIEPTORS
OF IN
FIBROBLAST
THE
TISSUES
AND
MENINGIOMAS
GROWTH OF
HUMAN
l-7
FACTOR GLIOMAS
Jun A. Ta:kahashil,', Hiroshi Suzui112, Yosiko Yasuda3, Nobuyuki Mitsuhiro Ohta5, Michael Jaye6, Manabu Fukumoto7, Yoshifumi Oda', Haruhiko Kikuchi' and Masakazu Hatanakall*
Ito',
'Human Cancer Laboratory, Department of Molecular Virology, Institute for Virus Research, Kyoto University, Kyoto 606, of Neurosurgery, Faculty of Medicine, Kyoto JAPAN: 'Department University, Kyoto 606, JAPAN: 3Department of Anatomy, Kinki University School of Medicine, Osaka 589, JAPAN; 4Department of Biological Chemistry, Faculty of Pharmaceutical Sciences, Kyoto University, Kyoto 606, JAPAN; 5Clinical Research Center, Utano National Hospital, Kyoto 606, JAPAN; 6Rorer Biotechnology, King of Prussia, PA 19406;7DepartmentofPathology,FacultyofMedicine, Kyoto University, Kyoto 606, JAPAN Received April
9, 1991
Northern blot analysis showed transcripts of two types of the fibroblast growth factor (FGF) receptor genes, flg and bek, in almost all the tissues samples of 18 human gliomas and 22 human meningiomas, which produced abundant basic and/or acidic FGF. From immunohistochemistry, FGF receptors were expressed in the tumor cells of a glioma and a RNA expression of these FGF receptors was also meningioma. detectable in normal human brains and normal bovine meninges. The expression level of either FGF receptor gene was not significantly different between tumor tissues and normal tissues. Summary;
(FGFs) are and acidic fibroblast growth factors mi togens and differentiation factors for neuroectoderm-derived cells as well as potent cells or mesoderm-derived (1) It has been suggested that FGFs angiogenic factors (1,2). in neoplastic growth from the following are involved Basic
a group evidences; FGF-5 and hst-2/FGF-6, *To whom addressed. Abbreviations: 15mM
sodium
all
of
oncogenes, such as encodes FGF-related
correspondence FGF, citrate;
and
reprint
hst/K-fgf,
int-2,
proteins requests
(X-8);
should
be
fibroblast growth factor; SSC, 0.15M NaCl, SDS, sodium dodecyl sulfate. 0006-291X/91$1.50 1
Copyright 0 1991 by Academic Press. Inc. All rights of reproduction in any form reserved.
Vol.
177,
No.
BIOCHEMICAL
1, 1991
certain
cells
can
AND
BIOPHYSICAL
RESEARCH
transformed
acquire
phenotypes
transfection of the basic FGF gene (9-12); cells produce FGFs as autocrine growth hybridization demonstrated using Northern acidic
FGF
human
gliomas
were and
abundantly produced meningiomas, which (13).
human brain tumors bear FGF receptors with FGF
after
many kinds of tumor (2). We factors that basic and/or in more than comprise about
cells
are
also
90% of 50% of known
to
from the in vitro study by cross-linking the cDNAs for two types of human (14). Recently, (15-17). In flg and bek, were isolated also demonstrate the capacity these FGF receptors
1251-FGFs receptors,
addition, for
Some glioma
COMMUNICATIONS
binding
(16). expressed analysis as were genes
In
both
order to in brain
basic
and acidic
determine tumor
whether tissues
FGFs
with
high
affinity
FGF receptor genes are in vivo, Northern blot
for flg and bek was performed using the same samples previously examined as to the expression of the FGF Furthermore, (13). immunohistochemical study showed the
localization
Materials
with
of
and
FGF receptors
in
tumor
tissues.
Methods
Tissue samples of brain tumors were obtained Tissue Samoles. from 45 patients operated on at Kyoto University Hospital and affiliated hospitals. Tumor samples consisted of 18 gliomas, 22 meningiomas and five metastatic brain tumors (Table). Normal human brain tissues were obtained from two patients who required lobectomy. These samples were described previously (13). Northern Blot Analysis. Northern blot analysis was performed as described (13). Briefly, Total RNA was isolated by the Guanidinium thiocianate/cesium chloride method. Twenty micrograms of total RNA were denatured in 1M glyoxa1/50% dimethyl sulfoxide, fractionated by electrophoresis in 1% agarose and transferred to diazophenylthioether gels paper (Schleicher & Schuell, Inc., Keene, NH). The following cDNA probes were used for hybridization: fig, [a 2.8-kb EcoR I fragment (15) ] and bek, [a 2.25-kb EcoRI fragment (16)1. These probes were labeled with [a-32P]dCTP by random priming and hybridization was carried out. The final washes were twice performed under stringent conditions using 0.1X SSC and 0.5% SDS at 65OC for 30 minutes each time Filters (13). were then autoradiographed for two days at -7OOC. Densitometric measurement of band intensities was performed by a Zeineh soft laser scanning densitometer (Biomed). Immunohistochemistrv. Polyclonal antibodies against a synthetic peptide from human FLG [residues 782-805 (1611 were raised in rabbits according to the previous report (18). The antibodies crossreacted with a synthetic FLG fragment on a Western blot analysis. Frozen sections were prepared from tissue samples of a glioblastoma and a meningioma. Six-micron-thick sections were treated with the polyclonal anti-human FLG antibodies 2
Vol.
177,
No.
1, 1991
BIOCHEMICAL
AND
BIOPHYSICAL
RESEARCH
COMMUNICATIONS
(1:4000), biotynilated anti-rabbit antibody IgG (1:200:1 (Vector), followed by an avidin-biotin procedure (Vector) (13). The sections were counterstained with methylgreen. The staining specificity of antibody against FGF was assessed by comparing the staining pattern after receptc#r absorption by antigen.
Results
and
Discussion
Northern
blot
analvsis.
in
18 gliomas
all
metastatic
brain
bek
for
was
A 4.2-kb and
tumors
detected
(Fig. at
1-A kb
4.4
flg
band for
22 meningiomas
was detectable
and
three
and Tablel).
(Fig.
1-B)
of
five
A transcript
in all
18
gliomas,
19 of 22 meningiomas and three of five metastatic brain tumors (Table I). Both types of the FGF receptor genes were also expressed in normal human brains and bovine no significant difference in the expression or bek addition,
gene between tumor tissues there was no significant
levels
of
(assessed
the
FGF receptor
genes
to
Immunohistochemistrv. antibody
in
was observed (Fig.
2-A
3 12
8
1s
those
C).
In
the
7
2
22
35
34
flg
28 24
the
FLG
and meningioma
control
experiment,
meatastatic
42 41
44
43
no
8
brahtumors 26
FGF genes
P >O.Ol). anti-human
cells
menhgknna 3
of
test, with
glioblastoma
gbma
B
2
and
was
and normal tissues. In correlation of expression
rank correlation Dense staining
cells
by Spearman's
meninges. There levels of the
45
f
E
.z .s h
2
-28s -4.2kb -18s
C
actn
Fiq.
-
1.
Northern
hybridization
of
FGF
receptors.
Twenty micrograms of total RNA were loaded in each slot except in the bovine hypothalamus (2Opg of poly (A)+ RNA). The number of each lane indicates the patient number in Tablel. 3
Vol.
177,
No.
1, 1991
BIOCHEMICAL
AND
BIOPHYSICAL
RESEARCH
COMMUNICATIONS
Table 1 Sample lists and relative amounts of mRNA measured by a densitometer Patient
Gliomas
Meningiomas
Metastatic brain tumor
Control
I 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45
WHO classification
astrocytoma astrocytoma astrocytoma astrocytoma astrocytoma anplastic astrocytoma anplastic astrocytoma anplastic astrcqtoma glioblastoma glioblastoma glioblastoma glioblastoma glioblastoma glioblastoma oligodendroglioma malignant oligodendroglioma oligodendroglioma malignant ependymoma meningioma meningioma meningioma meningioma meningioma meningioma meningioma meningioma meningioma meningioma merdngioma meningioma meningioma meningioma meningioma meningioma meningioma meningioma meningioma meningioma menmgioma meningioma malignant lymphoma lung cancer sarcoma adenocarcinoma renal cell carcinoma bovine hypothalamus bovine meninges normal human brain normal human brain
basic FGF
acidic FGF
fig
bek
91 83 74 102 0 >274 154 124 270 167 152 150 120 61 52 176 98 191 46 167 59 80 54 0 0 120 141 133 117 85 96 48 57 85 37 59 178 170 41 87 0 0 0 0 0 100 0 0 0
128 0 >I42 108 0 85 >142 0 28 50 >142 87 25 137 0 69 >142 0 0 >I42 0 0 0 0 0 0 0 0 >142 0 0 0 0 0 0 0 0 0 0 130 0 0 0 0 0 100 0 0 0
24 35 22 20 114 60 21 100 78 89 29 100 68 20 31 100 33 120 89 94 36 85 35 32 13 60 74 32 28 63 34 30 21 73 79 26 57 81 37 78 0 0 96 79 5 100 35 71 15
14 60 23 10 23 19 25 13 19 12 12 11 12 4 13 8 11 40 32 61 0 19 4 21 10 23 12 22 14 6 5 0 0 13 14 17 19 14 12 19 0 0 10 10 6 100 24 14 7
AU values of mRNA of growth factor and receptor are expressed relative to the level of mRiVA in bovine hypothalamus poly (A)+ RNA, which was arbitrarily set at 100. Twenty micrograms of total RNA were used in all samples except in the bovine hypothalamus [2Oug of poly(A)+ RNA]. The values of FGFs were taken from reference 13. 4
Vol.
177,
No.
1, 1991
BIOCHEMICAL
AND
BIOPHYSICAL
RESEARCH
COMMUNICATIONS
Remarkable immunoreactivities 2 . Immunohistochemistry. Fiq. toward anti-FLG antibodies were detected in tumor cells of while glioblastoma tissues (Panel A, x160) they were
undetectable in a section of glioblastoma absorbed antibody (Panel B, x400). Dense protein was also detected in meningioma cells
immunoreactivity
was detected
when the
treated with the staining of FLG (Panel C, x160).
absorbed
applied to a glioblastoma section (Fig.2-B). Autonomous cell growth and tumorigenesis the
constitutive
interaction
of
cellular
antibody
may result
growth
factors
was from with
Vol.
177,
No.
their
1, 1991
BIOCHEMICAL
corresponding
receptors
AND
BIOPHYSICAL
RESEARCH
COMMUNICATIONS
We have demonstrated that FGFs in vivo while the human gliomas and meningiomas produce expression of FGFs was not detected in normal brain tissues Immunohistochemical study also and meningeal tissues (13) showed immunoreactivities toward anti-basic FGF in most of gliomas
and
biological demonstrated present basic FGFs
meningiomas
in
activity of in human
basic glioma
study
meningiomas and play
meningiomas Some
showed expressed
through of the of
basic
suggesting
or
cells
almost
all
either
flg
or bek
These in
an autocrine metastatic that
vivo (20,21). or acidic FGF
that
acidic FGF genes. a crucial role
transcript genes,
(19).
acidic FGFs
as a paracrine factor. The present study has and the meningiomas actually
in
vitro
of
the
gene
suggest
there was of the flg
and
It FGFs
of
normal
tissues.
tumor-derived
and meningiomas through cellular FGF receptors.
is
(14,22). gliomas
in
vivo
that
The and
the
as well
as
tumor-derived
tumorigenesis of gliomas mechanism (13,14,20-22). brain tumors, which showed
and
FGF,
bek
are
expressed
involved
shown that express
both
in
these
suggested
and
growths
of both FGFs were than in normal brains
no significant or bek gene between
is involved in the constitutive
flg tumor
no
almost all of the gliomas the FGF receptor genes in
vivo. However, while gene expressions elevated in gliomas and meningiomas more and meninges, expression levels
Moreover, the was reportedly
that
difference tumor
a significant
tumorigenesis interaction
of tissues amount
of
gliomas of with
Acknowledgments This work was supported by Grants Foundation and the Ministry of Culture of Japan to M.H.
from the Education,
Uehara Memorial Science and
References 1. 2. 3. 4.
Gospodarowicz, D., Neufeld, G., and Schweigerer, L. (1987) J. Cell. Physiol. supp. 5,15-26. Folkman, J. and Klagsbrun, M. (1987) Science. 235,442447. Sakamoto, B., Mori, M., Taira, M., Yoshida, T., Matsukawa, S Shimizu, K., Sekiguchi, M., Terada, M., and Sugimura, T. - ' (1986) Proc. Natl. Acad. Sci. USA. 83.3997-4001. Bovi, P.D., Curatola, A.M., Kern, F.G., Greco, A., Ittmann, M., and Basilica, C. (1987) Cell. 50,729-737. 6
Vol.
177,
No.
1, 1991
BIOCHEMICAL
AND
BIOPHYSICAL
RESEARCH
COMMUNICATIONS
and Zhan, X., Culpepper, A., Reddy, M., Loveless, J., Goldfarb, M. (1987) Oncogene. 1,369-376. 6. Moore, R., Casey, G., Brookes, S., Dixon, M., Peters, G., and Dickson, C. (1986) EMBO J. 5,919-924. 7. Sakamoto. H., Yoshida, T., Nakakuki, M., Odagiri, H., Miyagawa, K., Sugimura, T., and Terada, M. (1988) Biochem. Biophys. Res. Commun. 151,973-981. 8. Marics, I., Adelaide, J., Raybaud, F., Mattei, M-G., COUlier, F., Planche, J., de Lapeyriere, O., and Dirnbaum, D. (1989) Oncogene. 4,335-340. 9. Sasada, R., Kurokawa, T., Iwane, M., and Igarashi, K. (1988) Mol. Cell. Biol. 8,588-594. 10. Quatro, N, Talarico, D., Sommer, A., Florkiewics, R., Basilica, C., and Rifkin, D.B. (1989) Oncogene Res. 5,101-110. 11. Yayon A., and Klagsbrun, M. (1990) Proc. Natl. Acad. Sci. USA.. 87,5346-5350. 12. Neu:Eeld, G., Mitchell, R., Ponte, P., and Gospodarowica, D. (1988) J. Cell Biol. 106,1385-1394. 13. Takahashi, J-A., Mori, H., Fukumoto, M., Igarashi, K., Jayle, M., Oda, Y., Kikuchi, H., and Hatanaka, M. (1990) Proc. Natl. Acad. Sci. USA. 87,5710-5714. 14. Libermann, T.A., Friesel, R., Jaye, M., Lyall, R.M., Westermark, B., Drohan, W., Schmidt, A., Maciag, T., and EMBO J. 6,1627-1632. Schlessinger, J. (1987) 15. Itoh, N., Terachi, T., Ohta, M., and Seo, M.K. (1990) Biochem. Biophys. Res. Commun. 169,680-685. 16. Dianne, C.A., Crumley, G., Bellot, F., Kaplow, J.M., Searfoss, G., Ruta, M., Burgess, W.H., Jaye, M., and EMBO J. 9,2685-2692. Schlessinger, J. (1990) 17. Rut-a, M., Howk, R., Ricca, G., Drohan, W., Zabelshansky, M - I Laureys, G., Barton, D.E., Francke, U., Schlessinger, J and Givol, D. (1988) Oncogene. 3,9-15. 18. Ru;;a, M., Burgess, W., Givol, D., Epstein, J., Neiger, N., Crumley, G., Dionne, C., Jaye, M., and Kaplow, J., Schlessinger, J. (1989) Proc. Natl. Acad. Sci. USA. 86,8722-8726. 19. Sporn, M.B., and Roberts, A.B. (1985) Nature (London). 313,745-747. Grothe, C., Sensenbrenner, M., Janet, T., 20. Paulus, w., Bauer, L., Graf, M., and Roggendorf, W. (1990) Acta. Neuropathol. 79,418-423. 21. Zagzag, D., Douglas, C. M., Sato, Y., Rifkin, D. B., and Res. Burstein, D.E. (1991) Cancer 50,7393-7398. 22. Sate, Y., and Friesen, H.G. Murphy, P.R., Sato, R., (1.989) Mol. Endoclinol. 3,744-748. 5.
177,
No.
May
31, 1991
GENE
1, 1991
BIOCHEMICAL
AND
BIOPHYSICAL
RQEARCH
COMMUNICATIONS Pages
EXPRESSION
RECIEPTORS
OF IN
FIBROBLAST
THE
TISSUES
AND
MENINGIOMAS
GROWTH OF
HUMAN
l-7
FACTOR GLIOMAS
Jun A. Ta:kahashil,', Hiroshi Suzui112, Yosiko Yasuda3, Nobuyuki Mitsuhiro Ohta5, Michael Jaye6, Manabu Fukumoto7, Yoshifumi Oda', Haruhiko Kikuchi' and Masakazu Hatanakall*
Ito',
'Human Cancer Laboratory, Department of Molecular Virology, Institute for Virus Research, Kyoto University, Kyoto 606, of Neurosurgery, Faculty of Medicine, Kyoto JAPAN: 'Department University, Kyoto 606, JAPAN: 3Department of Anatomy, Kinki University School of Medicine, Osaka 589, JAPAN; 4Department of Biological Chemistry, Faculty of Pharmaceutical Sciences, Kyoto University, Kyoto 606, JAPAN; 5Clinical Research Center, Utano National Hospital, Kyoto 606, JAPAN; 6Rorer Biotechnology, King of Prussia, PA 19406;7DepartmentofPathology,FacultyofMedicine, Kyoto University, Kyoto 606, JAPAN Received April
9, 1991
Northern blot analysis showed transcripts of two types of the fibroblast growth factor (FGF) receptor genes, flg and bek, in almost all the tissues samples of 18 human gliomas and 22 human meningiomas, which produced abundant basic and/or acidic FGF. From immunohistochemistry, FGF receptors were expressed in the tumor cells of a glioma and a RNA expression of these FGF receptors was also meningioma. detectable in normal human brains and normal bovine meninges. The expression level of either FGF receptor gene was not significantly different between tumor tissues and normal tissues. Summary;
(FGFs) are and acidic fibroblast growth factors mi togens and differentiation factors for neuroectoderm-derived cells as well as potent cells or mesoderm-derived (1) It has been suggested that FGFs angiogenic factors (1,2). in neoplastic growth from the following are involved Basic
a group evidences; FGF-5 and hst-2/FGF-6, *To whom addressed. Abbreviations: 15mM
sodium
all
of
oncogenes, such as encodes FGF-related
correspondence FGF, citrate;
and
reprint
hst/K-fgf,
int-2,
proteins requests
(X-8);
should
be
fibroblast growth factor; SSC, 0.15M NaCl, SDS, sodium dodecyl sulfate. 0006-291X/91$1.50 1
Copyright 0 1991 by Academic Press. Inc. All rights of reproduction in any form reserved.
Vol.
177,
No.
BIOCHEMICAL
1, 1991
certain
cells
can
AND
BIOPHYSICAL
RESEARCH
transformed
acquire
phenotypes
transfection of the basic FGF gene (9-12); cells produce FGFs as autocrine growth hybridization demonstrated using Northern acidic
FGF
human
gliomas
were and
abundantly produced meningiomas, which (13).
human brain tumors bear FGF receptors with FGF
after
many kinds of tumor (2). We factors that basic and/or in more than comprise about
cells
are
also
90% of 50% of known
to
from the in vitro study by cross-linking the cDNAs for two types of human (14). Recently, (15-17). In flg and bek, were isolated also demonstrate the capacity these FGF receptors
1251-FGFs receptors,
addition, for
Some glioma
COMMUNICATIONS
binding
(16). expressed analysis as were genes
In
both
order to in brain
basic
and acidic
determine tumor
whether tissues
FGFs
with
high
affinity
FGF receptor genes are in vivo, Northern blot
for flg and bek was performed using the same samples previously examined as to the expression of the FGF Furthermore, (13). immunohistochemical study showed the
localization
Materials
with
of
and
FGF receptors
in
tumor
tissues.
Methods
Tissue samples of brain tumors were obtained Tissue Samoles. from 45 patients operated on at Kyoto University Hospital and affiliated hospitals. Tumor samples consisted of 18 gliomas, 22 meningiomas and five metastatic brain tumors (Table). Normal human brain tissues were obtained from two patients who required lobectomy. These samples were described previously (13). Northern Blot Analysis. Northern blot analysis was performed as described (13). Briefly, Total RNA was isolated by the Guanidinium thiocianate/cesium chloride method. Twenty micrograms of total RNA were denatured in 1M glyoxa1/50% dimethyl sulfoxide, fractionated by electrophoresis in 1% agarose and transferred to diazophenylthioether gels paper (Schleicher & Schuell, Inc., Keene, NH). The following cDNA probes were used for hybridization: fig, [a 2.8-kb EcoR I fragment (15) ] and bek, [a 2.25-kb EcoRI fragment (16)1. These probes were labeled with [a-32P]dCTP by random priming and hybridization was carried out. The final washes were twice performed under stringent conditions using 0.1X SSC and 0.5% SDS at 65OC for 30 minutes each time Filters (13). were then autoradiographed for two days at -7OOC. Densitometric measurement of band intensities was performed by a Zeineh soft laser scanning densitometer (Biomed). Immunohistochemistrv. Polyclonal antibodies against a synthetic peptide from human FLG [residues 782-805 (1611 were raised in rabbits according to the previous report (18). The antibodies crossreacted with a synthetic FLG fragment on a Western blot analysis. Frozen sections were prepared from tissue samples of a glioblastoma and a meningioma. Six-micron-thick sections were treated with the polyclonal anti-human FLG antibodies 2
Vol.
177,
No.
1, 1991
BIOCHEMICAL
AND
BIOPHYSICAL
RESEARCH
COMMUNICATIONS
(1:4000), biotynilated anti-rabbit antibody IgG (1:200:1 (Vector), followed by an avidin-biotin procedure (Vector) (13). The sections were counterstained with methylgreen. The staining specificity of antibody against FGF was assessed by comparing the staining pattern after receptc#r absorption by antigen.
Results
and
Discussion
Northern
blot
analvsis.
in
18 gliomas
all
metastatic
brain
bek
for
was
A 4.2-kb and
tumors
detected
(Fig. at
1-A kb
4.4
flg
band for
22 meningiomas
was detectable
and
three
and Tablel).
(Fig.
1-B)
of
five
A transcript
in all
18
gliomas,
19 of 22 meningiomas and three of five metastatic brain tumors (Table I). Both types of the FGF receptor genes were also expressed in normal human brains and bovine no significant difference in the expression or bek addition,
gene between tumor tissues there was no significant
levels
of
(assessed
the
FGF receptor
genes
to
Immunohistochemistrv. antibody
in
was observed (Fig.
2-A
3 12
8
1s
those
C).
In
the
7
2
22
35
34
flg
28 24
the
FLG
and meningioma
control
experiment,
meatastatic
42 41
44
43
no
8
brahtumors 26
FGF genes
P >O.Ol). anti-human
cells
menhgknna 3
of
test, with
glioblastoma
gbma
B
2
and
was
and normal tissues. In correlation of expression
rank correlation Dense staining
cells
by Spearman's
meninges. There levels of the
45
f
E
.z .s h
2
-28s -4.2kb -18s
C
actn
Fiq.
-
1.
Northern
hybridization
of
FGF
receptors.
Twenty micrograms of total RNA were loaded in each slot except in the bovine hypothalamus (2Opg of poly (A)+ RNA). The number of each lane indicates the patient number in Tablel. 3
Vol.
177,
No.
1, 1991
BIOCHEMICAL
AND
BIOPHYSICAL
RESEARCH
COMMUNICATIONS
Table 1 Sample lists and relative amounts of mRNA measured by a densitometer Patient
Gliomas
Meningiomas
Metastatic brain tumor
Control
I 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45
WHO classification
astrocytoma astrocytoma astrocytoma astrocytoma astrocytoma anplastic astrocytoma anplastic astrocytoma anplastic astrcqtoma glioblastoma glioblastoma glioblastoma glioblastoma glioblastoma glioblastoma oligodendroglioma malignant oligodendroglioma oligodendroglioma malignant ependymoma meningioma meningioma meningioma meningioma meningioma meningioma meningioma meningioma meningioma meningioma merdngioma meningioma meningioma meningioma meningioma meningioma meningioma meningioma meningioma meningioma menmgioma meningioma malignant lymphoma lung cancer sarcoma adenocarcinoma renal cell carcinoma bovine hypothalamus bovine meninges normal human brain normal human brain
basic FGF
acidic FGF
fig
bek
91 83 74 102 0 >274 154 124 270 167 152 150 120 61 52 176 98 191 46 167 59 80 54 0 0 120 141 133 117 85 96 48 57 85 37 59 178 170 41 87 0 0 0 0 0 100 0 0 0
128 0 >I42 108 0 85 >142 0 28 50 >142 87 25 137 0 69 >142 0 0 >I42 0 0 0 0 0 0 0 0 >142 0 0 0 0 0 0 0 0 0 0 130 0 0 0 0 0 100 0 0 0
24 35 22 20 114 60 21 100 78 89 29 100 68 20 31 100 33 120 89 94 36 85 35 32 13 60 74 32 28 63 34 30 21 73 79 26 57 81 37 78 0 0 96 79 5 100 35 71 15
14 60 23 10 23 19 25 13 19 12 12 11 12 4 13 8 11 40 32 61 0 19 4 21 10 23 12 22 14 6 5 0 0 13 14 17 19 14 12 19 0 0 10 10 6 100 24 14 7
AU values of mRNA of growth factor and receptor are expressed relative to the level of mRiVA in bovine hypothalamus poly (A)+ RNA, which was arbitrarily set at 100. Twenty micrograms of total RNA were used in all samples except in the bovine hypothalamus [2Oug of poly(A)+ RNA]. The values of FGFs were taken from reference 13. 4
Vol.
177,
No.
1, 1991
BIOCHEMICAL
AND
BIOPHYSICAL
RESEARCH
COMMUNICATIONS
Remarkable immunoreactivities 2 . Immunohistochemistry. Fiq. toward anti-FLG antibodies were detected in tumor cells of while glioblastoma tissues (Panel A, x160) they were
undetectable in a section of glioblastoma absorbed antibody (Panel B, x400). Dense protein was also detected in meningioma cells
immunoreactivity
was detected
when the
treated with the staining of FLG (Panel C, x160).
absorbed
applied to a glioblastoma section (Fig.2-B). Autonomous cell growth and tumorigenesis the
constitutive
interaction
of
cellular
antibody
may result
growth
factors
was from with
Vol.
177,
No.
their
1, 1991
BIOCHEMICAL
corresponding
receptors
AND
BIOPHYSICAL
RESEARCH
COMMUNICATIONS
We have demonstrated that FGFs in vivo while the human gliomas and meningiomas produce expression of FGFs was not detected in normal brain tissues Immunohistochemical study also and meningeal tissues (13) showed immunoreactivities toward anti-basic FGF in most of gliomas
and
biological demonstrated present basic FGFs
meningiomas
in
activity of in human
basic glioma
study
meningiomas and play
meningiomas Some
showed expressed
through of the of
basic
suggesting
or
cells
almost
all
either
flg
or bek
These in
an autocrine metastatic that
vivo (20,21). or acidic FGF
that
acidic FGF genes. a crucial role
transcript genes,
(19).
acidic FGFs
as a paracrine factor. The present study has and the meningiomas actually
in
vitro
of
the
gene
suggest
there was of the flg
and
It FGFs
of
normal
tissues.
tumor-derived
and meningiomas through cellular FGF receptors.
is
(14,22). gliomas
in
vivo
that
The and
the
as well
as
tumor-derived
tumorigenesis of gliomas mechanism (13,14,20-22). brain tumors, which showed
and
FGF,
bek
are
expressed
involved
shown that express
both
in
these
suggested
and
growths
of both FGFs were than in normal brains
no significant or bek gene between
is involved in the constitutive
flg tumor
no
almost all of the gliomas the FGF receptor genes in
vivo. However, while gene expressions elevated in gliomas and meningiomas more and meninges, expression levels
Moreover, the was reportedly
that
difference tumor
a significant
tumorigenesis interaction
of tissues amount
of
gliomas of with
Acknowledgments This work was supported by Grants Foundation and the Ministry of Culture of Japan to M.H.
from the Education,
Uehara Memorial Science and
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