Biomedical subjects

H Kurachi

Publications and source records attributed to H Kurachi.

At least 91 records · Page 5Linked to original sources

PubMed1994

Involvement of epidermal growth factor (EGF)/EGF receptor autocrine and paracrine mechanism in human trophoblast cells: functional differentiation in vitro.

We have studied the expression of epidermal growth factor (EGF) and EGF receptors (EGF-R) in isolated human trophoblast cells at various stages of differentiation and also the biological significance of the EGF/EGF-R autocrine and paracrine mechanism. Cytotrophoblast cells were isolated from human placental tissues of 6-9 weeks of gestation. Trophoblast cells underwent morphological and functional differentiation during in vitro culture. The expression of EGF and EGF-R protein and mRNA was studied in trophoblast cells cultured for 0-5 days, using immunocytochemical staining, and reverse transcription and polymerase chain reaction. Monoclonal antibodies (mAbs) against EGF and EGF-R showed specific staining in trophoblast cells at all stages of differentiation. Both EGF and EGF-R gene transcripts were detected in RNA samples isolated from trophoblast cells at all stages. These data suggest the presence of an EGF/EGF-R autocrine and paracrine mechanism in human trophoblast cells. Next, we examined the biological significance of this mechanism on trophoblast cell differentiation in vitro. EGF added to the culture medium significantly increased human chorionic gonadotrophin-beta (hCG-beta) secretion and, more importantly, anti-EGF neutralizing mAbs significantly reduced both hCG-beta and human placental lactogen secretion from trophoblast cells in culture. All these results suggest that human trophoblast cells express both EGF and EGF-R, and that EGF may play an important role in the functional differentiation of human trophoblast cells.

Base Sequence↗

PubMed1994

[Growth factors in gynecology].

In this lecture, importance of growth factors in reproductive functions and cancer growth are discussed. Among many kinds of growth factors, epidermal growth factor (EGF) and transforming growth factor (TGF)alpha are mentioned; both of these are called as EGF family because these share a common receptor (EGF receptor) and show similar biological activities. It is known that growth factors are important in reproductive fields. They play vital roles in the follicle development and the endometrial proliferation in response to estrogen. We studied the expression and role of EGF and TGF alpha in human fallopian tube. Immunohistochemical and RT-PCR studies revealed the menstrual stages specific synthesis and expression of EGF and TGF alpha in tubal epithelial cells. They were abundant at the late follicular and luteal stages, were little at the early follicular stage, suggesting that these growth factors are expressed in response to estrogen. We, next, examined the role of these factors in early embryo development using mouse embryos. Embryo development was significantly improved when embryos were co-cultured with human tubal epithelial cells. However, the favorable effects of the tubal epithelial cells were almost completely abolished by the addition of anti-EGF and -TGF alpha neutralizing antibodies. These facts suggest that EGF and EGF alpha expressed in human epithelial cells promote embryo development in a paracrine manner. Autocrine mechanisms by growth factors are known to be important on cancer cell growth. Among many kinds of autocrine mechanisms TGF alpha/EGF receptor autocrine mechanism is the most commonly expressed in many kinds of cancers such as lung cancer, esophageal cancer, pancreas cancer and liver cancer.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1993

Interleukin 6 inhibits mouse placental lactogen II but not mouse placental lactogen I secretion in vitro.

The mouse placenta produces several polypeptides belonging to the prolactin-growth hormone gene family, including mouse placental lactogen (mPL) I and mPL-II. The present study was undertaken to determine whether the secretion of mPL-I and mPL-II is regulated by interleukin 6 (IL-6), which is present in the placenta and has previously been reported to stimulate the secretion of pituitary members of this gene family. Effects of human and mouse IL-6 on mPL-I and mPL-II secretion were examined in primary cultures of placental cells from days 7, 9, and 12 of pregnancy. IL-6 caused a dose-dependent reduction in the mPL-II concentration in the medium of cells from days 9 and 12 of pregnancy but did not affect the mPL-II concentration in the medium of cells from day 7 of pregnancy or the mPL-I concentration in the medium of cells from days 7 or 9 of pregnancy. The lowest concentration of human IL-6 that significantly inhibited mPL-II secretion was 250 pM. The effect of IL-6 on the mPL-II concentration in the medium was due primarily to inhibition of mPL-II synthesis, which resulted at least partly from a decrease in the steady-state level of mPL-II mRNA. These data raise the possibility that IL-6 may regulate mPL-II production after midpregnancy in vivo.

Animals↗

PubMed1993

Autocrine growth mechanism by transforming growth factor (TGF)-beta 1 and TGF-beta 1-receptor regulation by epidermal growth factor in a human endometrial cancer cell line IK-90.

Transforming growth factor-beta 1 (TGF-beta 1) enhanced cell proliferation in a concentration-dependent manner in a human endometrial cancer cell line, IK-90. Scatchard analysis of TGF-beta 1 receptor in IK-90 cells, using 125I-TGF-beta 1 as a ligand, revealed the presence of a class of high-affinity TGF-beta 1 receptors (2,000 sites per cell, KD = 74pM). Moreover, IK-90 cells produced and secreted TGF-beta 1: TGF-beta 1 messenger RNA was detected at 2.5 and 4.0 kb by Northern-blot analysis using 32P-labeled TGF-beta 1 cDNA as a probe, and TGF-beta 1 activity in conditioned medium by the inhibition of 3H-thymidine uptake into CCl 64 mink lung epithelial cells. We investigated the regulation of TGF-beta 1 receptor by 4 kinds of growth factor: epidermal growth factor (EGF) but not TGF-beta 1, insulin or insulin-like growth factor-1 increased the level of TGF-beta 1 binding sites in a concentration- and time-dependent manner. These findings suggest that TGF-beta 1 may be a potential autocrine growth factor in a human endometrial cancer cell line IK-90 and that this autocrine mechanism may be affected by EGF.

Adult↗

PubMed1993

Ovariectomy increases the level of estrogen receptor mRNA and estrogen receptor binding sites in female rat adipose tissue.

The roles of estrogen in the changes in estrogen receptor (ER) mRNA and ER binding sites in rat adipose tissue were studied in female rats. To elucidate the mechanism(s) behind ovariectomy (OVX)-induced obesity, the levels of ER mRNA and ER binding sites in adipose tissue were analyzed three weeks after OVX using Northern blot analysis of ER mRNA and the [3H]E2 binding assay, respectively. OVX induced an increase in body weight, and replacement of estradiol (E2) prevented that increase. Significant increases in the amounts of ER mRNA and in [3H]E2-specific binding were observed after OVX, and E2 replacement reduced both of those increases. These results suggest that E2 may regulate rat obesity directly through ER in adipose tissues.

Adipose Tissue↗

PubMed1993

Involvement of epidermal growth factor in inducing obesity in ovariectomized mice.

Ovariectomy (Ovx) of mice significantly increases the epidermal growth factor (EGF) concentration in the submandibular gland. To elucidate the role of this elevated EGF in obesity of Ovx mice, we examined the effects of sialoadenectomy (Sx) and anti-EGF rabbit antiserum administration on the body weight (BW) gain and carcass fat deposition in Ovx animals. Studies were performed in four groups of mice consisting of control, Ovx, Ovx+Sx, and Ovx+anti-EGF groups. Ovx increased the BW gain compared with the control animals, whereas Sx and anti-EGF significantly reduced it. Although the relative weights (weight ratio to BW) of the liver and kidney were not significantly changed by Ovx, Sx, or anti-EGF treatment of Ovx mice, the relative weights of mesenteric, parametrial, and subcutaneous fat tissues were increased in Ovx mice, and this increase was significantly reduced by Sx or anti-EGF administration. Ovx induced adipocyte hypertrophy, and this effect was eliminated by Sx and anti-EGF. Moreover, acyl-CoA synthetase mRNA level was increased by Ovx, and this increase was reduced by Sx and anti-EGF in mesenteric fat tissue. These findings suggest that elevation of EGF may play a role in the induction of obesity in Ovx mice.

Acetate-CoA Ligase↗

PubMed1993

Intraventricular administration of histidyl-proline-diketopiperazine [Cyclo(His-Pro)] suppresses prolactin secretion and synthesis: a possible role of Cyclo(His-Pro) as dopamine uptake blocker in rat hypothalamus.

Histidyl-proline-diketopiperazine [Cyclo (His-Pro) (CHP)] was discovered to be one of the metabolites of TRH. To understand the specific role of CHP in rat hypothalamic dopamine neurons, we examined the in vivo effects of intraventricular (icv) infusion of CHP on the release and synthesis of PRL in the rat pituitary and the 3,4-dihydroxyphenylacetic acid (DOPAC)/dopamine ratio in the rat hypothalamus. We also examined the in vitro effects of CHP on the release of [3H]dopamine from dispersed tuberoinfundibular dopamine neurons, [3H]dopamine reuptake in hypothalamic membrane fractions, and PRL release from rat pituitary cultured cells. Female rats were treated by icv infusion of 1 microM CHP daily for 1, 3, and 7 days, using Alzet osmotic pumps. After 1 day of treatment, the serum PRL concentration was significantly decreased. Northern blot analysis of the total RNA isolated from the pituitary glands of control animals using 32P-labeled PRL cDNA as a probe indicated the presence of PRL gene transcript, 1.0 kilobase in size, and its amount was decreased by CHP treatment. CHP did not affect [3H]dopamine release from dispersed tuberoinfundibular dopaminergic neurons at any concentration up to 1 microM. CHP did not inhibit PRL release from cultured pituitary cells at low concentrations (1-100 nM), but it stimulated PRL release at high concentrations (1 and 10 microM). We also examined the concentrations of dopamine and DOPAC in the rat hypothalamus when CHP was administered icv for 1 or 7 days. There was a significant decrease in the DOPAC/dopamine ratio after CHP treatment for 1 day. Furthermore, CHP caused dose-dependent inhibition of [3H]dopamine uptake by the rat hypothalamus similar to other dopamine uptake blockers, such as benztropine and GBR12909. These data suggest that icv administration of CHP might decrease both PRL secretion and accumulation of PRL gene transcripts in the pituitary by decreasing the DOPAC/dopamine ratio and inhibiting dopamine reuptake in the rat hypothalamus.

3,4-Dihydroxyphenylacetic Acid↗

PubMed1993

Menstrual stage-specific expression of epidermal growth factor and transforming growth factor-alpha in human oviduct epithelium and their role in early embryogenesis.

We studied the expression of epidermal growth factor (EGF) and transforming growth factor (TGF)-alpha in human oviduct epithelium at various menstrual stages. Immunohistochemical stainings using anti-EGF and anti-TGF alpha antibodies showed a specific staining in ampullary oviduct epithelium at late follicular and luteal stages, but the stainings were very weak at early follicular stage. Quantitative reverse transcription and polymerase chain reaction, using beta-actin messenger RNA as an internal standard, revealed the menstrual stage-specific expression of EGF and TGF alpha gene transcripts: relative amounts of EGF and TGF alpha messenger RNA to those of beta-actin were 1.5 +/- 1.9% (mean +/- SD) and 1.4 +/- 0.6% (n = 3) at early follicular, 16.5 +/- 4.9% and 12.6 +/- 2.6% (n = 3) at late follicular, and 18.9 +/- 2.2% and 13.8 +/- 3.2% (n = 3) at luteal stages, respectively. The expression of these growth factors was in proportion to the increase in serum estradiol but not to progesterone levels. To clarify the biological significance of these growth factors, mouse two-cell embryos were cocultured with human oviduct epithelial cells with or without blocking the action of these growth factors. Cocultures significantly promoted blastocyst formation, but this promotive effect of the oviduct epithelial cells was completely abolished by the addition of anti-EGF and/or anti-TGF alpha monoclonal neutralizing antibodies to the coculture system. All these results showed that EGF and TGF alpha were synthesized and expressed in human oviduct epithelium specifically to menstrual stages, and that these growth factors may be involved in early embryonic development.

Actins↗

PubMed1993

Imaging of peritoneal pseudocysts: value of MR imaging compared with sonography and CT.

OBJECTIVE: Peritoneal pseudocysts are created when fluid arising from the ovary is trapped by peritoneal adhesions. We studied the value of MR imaging, compared with sonography and CT, for detecting these abnormalities. SUBJECTS AND METHODS: Nine women, 27 to 49 years old, who had surgically proved peritoneal pseudocysts were included in the study. All nine patients were premenopausal, and eight had a history of pelvic surgery. All patients were examined with MR imaging and sonography, and five were also examined with CT. Imaging findings were analyzed and compared with the surgical findings. RESULTS: Sonograms in all nine patients showed a thick-walled cyst that contained focal echoes indicating nodules, making it difficult to differentiate these lesions from cystic ovarian tumors. In eight of the nine patients, the fluid in the pseudocysts had low signal intensity on T1-weighted MR images and high signal intensity on T2-weighted spin-echo MR images, suggesting that the fluid was serous. The pseudocyst had an irregular shape on MR images in seven patients and on CT scans in three patients. However, MR provided more information than CT did about the nature of the fluid within the pseudocyst. CONCLUSION: Our study suggests that MR imaging is more useful than sonography and CT for determining the characteristic findings of peritoneal pseudocysts.

Adult↗

PubMed1992

A new member of the insulin receptor family, insulin receptor-related receptor, is expressed preferentially in the kidney.

Shier and Watt isolated human and guinea pig genomic DNA encoding a putative protein, insulin receptor-related receptor (IRR), the primary structure of which is similar to that of other members of the insulin receptor family, the insulin receptor and type-I IGF receptor (J. Biol. Chem. 264, 14605-14608 (1989)). However, the expression of the IRR gene remained unknown. In this paper, we isolated the IRR cDNA from the rat brain and examined the expression of the IRR mRNA in a variety of rat tissues, including the brain, heart, lung, liver, small intestine, kidney, thymus, spleen, muscle, adipose tissue and cartilage by polymerase chain reaction. In contrast to the wide distribution of the insulin receptor and type-I IGF receptor mRNAs, the IRR mRNA is expressed preferentially in the kidney, which indicates that IRR has unique functions as a member of the insulin receptor family.

Amino Acid Sequence↗

PubMed1992

Activin A increases cytosolic free calcium concentration in rat pituitary somatotropes.

The effect of activin A on the cytosolic free calcium concentration ([Ca2+]i) in normal rat pituitary cells was examined using a calcium sensitive fluorescent dye, indo 1 AM, and a digital imaging fluorescent microscope system. The cells showing an increase in [Ca2+]i in response to activin A were then characterized by comparison with cells responding to growth hormone releasing hormone (GRH), thyrotropin releasing hormone (TRH), corticotropin releasing hormone (CRH), and gonadotropin releasing hormone (GnRH) in monolayer cultures of normal rat pituitary cells. Activin A increased [Ca2+]i in some cells in a mixed population of normal rat pituitary cells. The cells that responded to activin A also responded to GRH. Most of these cells were not affected by other tropic hormones (CRH, TRH, and GnRH), but a few cells responded to both GRH and TRH. None of the activin A-responding cells responded to CRH or GnRH, and none of the CRH- or GnRH-responding cells responded to activin A. In a preparation of somatotropes purified 80-90% by fluorescence-activated cell sorting, activin A increased [Ca2+]i in 30% of the cells that shows a [Ca2+]i-response to GRH. These findings suggest direct involvement of somatotropes in activin A-induced biological events in the rat pituitary gland.

Activins↗

PubMed1992

Binding sites for interleukin-6 in the anterior pituitary gland.

Interleukin-6 (IL-6) binding site in the rat anterior pituitary gland was characterized using radioiodinated human recombinant (hr) IL-6. Results showed that the anterior pituitary gland contained 170 binding sites per cell of a single class with a dissociation constant of 2.7 x 10(-9) M. The binding of 125I-hrIL-6 to the rat anterior pituitary gland was competitively inhibited by unlabeled hrIL-6, but not by hrIL-1 alpha, hrIL-1 beta, hrIL-2 or hr-interferon-gamma, indicating these binding sites are specific for IL-6. We also demonstrated mouse IL-6 receptor gene expression in the rat anterior pituitary gland by Northern blot analysis. Furthermore, the IL-6 receptor was detected on human gonadotrophs by the double immunofluorescence method. Our findings demonstrate the presence and expression of IL-6 binding site in the rat anterior pituitary gland and the presence of IL-6 binding site on human gonadotrophs, suggesting the important role of IL-6 binding site in pituitary hormone release in both species.

Animals↗

PubMed1992

Vasoactive intestinal peptide enhances dopamine accumulation in primary cell culture of rat hypothalamus.

The effect of vasoactive intestinal peptide (VIP) and PHI-27 on dopamine accumulation in cultured rat hypothalamic cells was investigated. VIP enhanced [3H]dopamine accumulation dose dependently. This effect was significant at 10(-8)-10(-5) M VIP with a concomitant increase in intracellular cyclic AMP (cAMP), and reached its plateau level at 10(-6) M VIP. VIP increased [3H]dopamine accumulation significantly within 15 min. PHI-27 and dibutyryl cAMP ((Bu)2-cAMP) also enhanced [3H]dopamine accumulation. These results suggest that VIP enhances dopamine accumulation in hypothalamic cells by increasing intracellular cAMP.

Animals↗

PubMed1992

Intraventricular administration of thyrotrophin-releasing hormone (TRH) suppresses prolactin secretion and synthesis: a possible involvement of dopamine release by TRH from rat hypothalamus.

The administration of thyrotrophin-releasing hormone (TRH) causes a variety of dopamine-related biological events. To understand the specific role of TRH on rat hypothalamic dopamine neurones, we examined the in-vivo effects of intraventricular (i.c.v.) infusion of TRH on the release and synthesis of prolactin in the rat pituitary gland and on the changes in binding of [3H]MeTRH and dopamine turnover rates in rat hypothalamus. We have also examined the in-vitro effects of TRH on the release of [3H]dopamine from dispersed tuberoinfundibular dopamine neurones. Female rats were treated with i.c.v. infusions of 1 mumol TRH/1 daily for 1, 3 and 7 days using Alzet osmotic pumps. Following 7 days of treatment the serum prolactin concentrations were significantly decreased. A reduction in hypothalamic TRH-binding sites (Bmax) was also apparent but the dissociation constant (Kd) was unaffected. Northern blot analysis of total RNA isolated from the pituitary glands of control animals using 32P-labelled prolactin cDNA as a probe indicated the presence of three species of prolactin gene transcripts of approximately 3.7, 2.0 and 1.0 kb in size, and these were decreased by TRH treatment. We examined the turnover rate of dopamine in the rat hypothalamus when TRH was administered i.c.v. for 7 days. There was a significant increase in 3,4-dihydroxyphenylacetic acid/dopamine ratio with TRH treatment. Moreover, exposure to TRH stimulated [3H]dopamine release from rat tuberoinfundibular neurones in a time- and dose-dependent manner. Dopamine receptor antagonists such as SCH23390 and (-)sulpiride, and other neuropeptides such as vasoactive intestinal peptide and oxytocin did not affect TRH-stimulated [3H]dopamine release.(ABSTRACT TRUNCATED AT 250 WORDS)

3,4-Dihydroxyphenylacetic Acid↗

PubMed1991

Involvement of transforming growth factor alpha/epidermal growth factor receptor autocrine growth mechanism in an ovarian cancer cell line in vitro.

Although transforming growth factor (TGF) alpha and epidermal growth factor (EGF) receptor (EGFR) autocrine mechanism is widely demonstrated in many kinds of cancers, its biological significances still remain circumstantial. We critically assessed the significance of this mechanism on the growth of an ovarian cancer cell line. Northern blot analysis in polyadenylated RNA isolated from cells by using 32P-labeled pre-TGF alpha, EGRF, and prepro-EGF complementary DNAs as probes revealed that pre-TGF alpha and EGFR but not prepro-EGF gene transcripts were expressed in the cell. TGF alpha and EGFR but not EGF proteins were observed by immunocytochemical stainings, using monoclonal antibodies against human TGF alpha, EGFR, and EGF, respectively. This cell line possessed a class of high affinity EGF receptor by 125I-EGF binding studies; Kd being 2.9 x 10(-10) M and Bmax to be 7.7 x 10(4) sites/cell. As much as 1.12 +/- 0.14 ng (SD; n = 3)/10(7) cells/24 h of TGF alpha was secreted in the conditioned media. These results suggested the expression of a TGF alpha/EGFR autocrine mechanism in this cell line. We, therefore, assessed the biological significance of this mechanism on the growth of this cell line in serum-free monolayer cell cultures. Although 0.1, 1.0, and 10 nM concentrations of TGF alpha did not show significant growth promotion, monoclonal antibodies against TGF alpha and EGFR but not EGF significantly inhibited cell growth. All these data suggested the biological importance of a TGF alpha/EGFR autocrine mechanism on the growth of this cell line in vitro.

Antibodies, Monoclonal↗

PubMed1991

Importance of transforming growth factor alpha/epidermal growth factor receptor autocrine growth mechanism in an ovarian cancer cell line in vivo.

We have elucidated the importance of a transforming growth factor (TGF) alpha and epidermal growth factor receptor autocrine mechanism on the growth of a human ovarian serous cystadenocarcinoma-derived cell line (SHIN-3) in vitro. In this study, we studied the biological significance of this autocrine mechanism in vivo using female athymic nude (nu/nu) mice. We measured the mouse plasma epidermal growth factor and TGF alpha levels by radioimmunoassay and enzyme-linked immunosorbent assay, respectively. Plasma epidermal growth factor concentrations were remarkably decreased by sialoadenectomy (Sx): 410 +/- 65 (SE) pg/ml (n = 10) in intact animals; and undetectable in Sx mice (n = 5). Plasma TGF alpha levels were 90 and 40 pg/ml in intact and in Sx animals, respectively. Ten million SHIN-3 cells inoculated into nu/nu mice formed tumors in 100% of mice, and tumors grew progressively. Implantabilities and tumor growth rates of inoculated cells were not affected by Sx and even by Sx and anti-mouse epidermal growth factor antibody treatment. However, anti-TGF alpha monoclonal antibody (mAb) administered to SHIN-3 cell-inoculated Sx animals drastically reduced the tumor growth. Although 10(7) SHIN-3 cells formed tumors in this group, tumor growth was significantly inhibited by 10 micrograms of anti-TGF alpha mAb given 3 times a week, and growth inhibitions were more by 20 micrograms of anti-TGF alpha mAb. Moreover, as aggressive tumor growth as that in Sx animals was resumed by the cessation of anti-TGF alpha mAb treatments. All these data suggested the biological importance of a TGF alpha/epidermal growth factor receptor autocrine mechanism on the growth of this cell line in vivo.

Animals↗

PubMed1991

Evidence for the involvement of transforming growth factor alpha and epidermal growth factor receptor autocrine growth mechanism in primary human ovarian cancers in vitro.

We examined 35 primary human ovarian adenocarcinomas for the presence of epidermal growth factor (EGF) receptor (EGFR) in plasma membranes from cancer tissues by using 125I-EGF as a ligand. Specific 125I-EGF bindings were observed in 20 (57%) of these 35 cases. Scatchard analysis showed a class of high affinity EGF receptor: Kd 5.0 +/- 1.0 x 10(-10) M; Bmax, 83.3 +/- 12.1 fmol/mg protein (mean +/- SE, n = 20). Northern analysis in polyadenylated RNA from 15 EGFR(+) cancers using pretransforming growth factor alpha (pre-TGF alpha), prepro-EGF complementary DNA, and pE7, a complementary DNA clone of human EGFR, as probes revealed that pre-TGF alpha and EGFR mRNAs but not prepro-EGF mRNA were expressed in all cases examined. Immunocytochemical studies using monoclonal antibodies (mAbs) against TGF alpha, EGF, and EGFR showed that TGF alpha and EGFR but not EGF proteins were present on ovarian cancer cells in all cases. These data suggested a possible TGF alpha/EGFR autocrine mechanism in EGFR(+) ovarian cancers. We, therefore, examined the biological significance of this autocrine mechanism by using primary monolayer cell cultures. In primary cultures from EGFR (+) cancers, TGF alpha added to the culture medium stimulated the [3H]thymidine incorporation dose dependently. Moreover, the addition of mAbs against TGF alpha and EGFR but not EGF inhibited [3H]thymidine incorporation dose dependently in EGFR(+) cancer cells. On the other hand, in primary cultures from EGFR(-) cancers, TGF alpha and anti-TGF alpha, -EGFR, and -EGF mAbs did not show any effects on [3H]thymidine incorporation. All these results suggested the possible crucial role of a TGF alpha/EGFR autocrine growth mechanism in primary human ovarian cancers which express EGFR.

Adenocarcinoma↗

PubMed1991

Epidermal growth factor attenuates cell proliferation by down-regulating the transforming growth factor-beta receptor in the osteoblastic cell line MC3T3-E1.

We investigated the role of transforming growth factor beta 1 (TGF-beta 1) in growth regulation of the murine osteoblastic cell line, MC3T3-E1. We detected TGF-beta activity in the conditioned medium of MC3T3-E1 cells and its activity was increased by acid treatment of the bioassay system using CC1 64 cells. Northern blot analysis revealed the expression of TGF-beta 1 precursor messenger ribonucleic acid (mRNA). MC3T3-E1 cells possessed a single class of high affinity TGF-beta 1 receptor (2.8 x 10(4) sites per cell, Kd = 196 pM). Cross-linking studies revealed three specific TGF-beta receptors with molecular masses of 65, 95 and 280 kDa. Both TGF-beta and epidermal growth factor (EGF) stimulated [3H]-thymidine incorporation into cells. EGF decreased the number of TGF-beta receptor dose-dependently, and pretreatment of cells with 10 nM EGF attenuated cell growth by TGF-beta. All these data suggested that TGF-beta might be an autocrine growth factor in murine osteoblastic MC3T3-E1 cells and that EGF might modulate the growth of cells by down-regulating the TGF-beta receptor level.

Animals↗