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B Westermark

Publications and source records attributed to B Westermark.

At least 163 records · Page 9Linked to original sources
PubMed1987

Expression of the c-myc proto-oncogene in growth stimulated porcine thyroid follicle cells.

In the present investigation the expression of c-myc in porcine thyroid follicle cells following the incubation with TSH or EGF was studied. EGF did not only have a mitotic effect on thyroid follicles but also increased c-myc mRNA. The increase in c-myc RNA is an early event in EGF stimulated thyroid cells. TSH had no effect on c-myc RNA levels of porcine thyroid cells. EGF-binding and sensitization to the mitotic activity of EGF, however, is increased by TSH. While TSH stimulates cAMP production and functional activity but not growth or c-myc expression of porcine thyroid cells, EGF produces cell division accompanied with c-myc expression.

Animals↗
PubMed1987

Cultured human endothelial cells express platelet-derived growth factor A chain.

Four principal cell types involved in the pathophysiologic response of the vessel wall--endothelial cells, smooth muscle cells, platelets, and monocyte/macrophages--secrete platelet-derived growth factor-like (PDGF-like) mitogenic activities. Extensive structural data on these activities exist only for the mitogen produced by platelets, which is a 30-kd dimeric protein composed of structurally related A and B polypeptide chains encoded by different genes. It was previously demonstrated that normal cultured endothelial cells transcribe mRNA encoding the B chain of PDGF from the c-sis gene. Here several new structural features of the mitogen produced by cultured vascular endothelial cells are shown. Hybridization analysis of RNA from normal cultured human umbilical vein endothelial (HUVE) cells revealed that they contain three PDGF A chain transcript species. These RNA species comigrated with and appeared to have the same relative abundance as the three RNA species previously identified in RNA from two human tumor cell lines. A chain transcripts were not identified in RNA from a strain of bovine aortic endothelial cells or in human dermal fibroblasts. The A chain transcripts in HUVE had the same relative abundance as the B chain transcripts. Immunoprecipitation of metabolically labeled endothelial conditioned medium with anti-PDGF antiserum revealed a 31-kd species which was split by reduction and alkylation into two species of 16.5 and 17 kd. Thus, endothelial cells secrete a dimeric mitogen antigenically related to PDGF, with a structure identical to previously isolated PDGF A-chain homodimer. These findings are consistent with the possibility that secretion of PDGF by human endothelial cells may be regulated independently of B-chain expression.

Cells, Cultured↗
PubMed1986

The phenotypic characteristics of simian sarcoma virus-transformed human fibroblasts suggest that the v-sis gene product acts solely as a PDGF receptor agonist in cell transformation.

Previous studies have indicated that the oncogene v-sis of simian sarcoma virus (SSV) encodes a growth factor that is structurally and functionally similar to platelet-derived growth factor (PDGF). In the present investigation we have analysed the phenotypic characteristics of human foreskin fibroblasts transformed by SSV. It was found that the PDGF receptors were extensively down-regulated. This finding is consistent with a high, local, extracellular concentration of a PDGF-like factor, synthesized by the transformed cell. The receptors were up-regulated by suramin, a drug that is known to dissociate PDGF and the v-sis product from the PDGF receptors. A cell-associated v-sis product of mol. wt 24,000 was identified by immunoprecipitation with PDGF antibodies; release of this component was induced by a high concentration of exogenous PDGF, indicating that a fraction of the product is associated with the PDGF receptors. SSV was not found to be an immortalizing virus; when serially passaged, SSV-transformed cells had essentially the same life-span as their non-transformed counterparts. Moreover, SSV did not induce growth in soft agar beyond the level afforded by exogenously added PDGF. Thus, the present study favors the notion that SSV transformation is mediated by a growth factor that mimics PDGF but has no further cellular effects.

Cell Division↗
PubMed1986

Growth factors induce early pre-replicative changes in senescent human fibroblasts.

As human fibroblasts in culture senesce their response to platelet-derived growth factor (PDGF) becomes attenuated. To clarify at which level such cells are blocked in the pre-replicative part of the cell cycle, we have analysed PDGF-induced pre-replicative events in senescent (phase III) cultures. We found that phase III cells retain a normal number of PDGF receptors and that these are functional with regard to PDGF-induced receptor autophosphorylation. Phase III cells also respond to PDGF by rapid actin reorganization and increased levels of c-fos and c-myc mRNA, similar to growth-arrested phase II fibroblasts. However, the expression of the nuclear antigen K-67, which in phase II cell is induced in S-phase and continues to be expressed throughout the cell cycle, is not induced in phase III cells in response to PDGF. We conclude that phase III human fibroblasts, although blocked with regard to proliferation, still retain a functional growth factor receptor system, and display early responses when exposed to growth factors, such as changes in the cytoskeleton and the expression of proto-oncogenes.

Cell Division↗
PubMed1986

Translocation of human glial and glioma cells in culture.

Two normal glial lines and four selected malignant glioma lines--considered as representative of the varying morphologies previously found among these lines--were studied quantitatively as to their translocatory ability on glass under standard culture conditions. The two glial lines showed very similar characteristics with almost identical results for total and net translocation, as well as in their directional persistence. The glioma lines gave values for these criteria which were either greater or lower than those of the glial cells. These values could to a certain degree be related to phenotypic traits, such as degree of cell polarization and amounts of cytoplasmic microfilament bundles. The findings once again display the heterogeneity of the human malignant glioma lines and raise doubts as to the general applicability of findings in studies of single lines of tumor cells, or of experimentally transformed cells, to all malignant cells.

Cell Movement↗
PubMed1986

Platelet-derived growth factor as a mediator of normal and neoplastic cell proliferation.

Human platelet-derived growth factor is the major mitogen in serum for connective-tissue-derived cells in culture. The factor is 30,000 mol. wt protein composed of two disulphide-linked polypeptide chains, named A and B. The B-chain is virtually identical to part of the transforming protein of simian sarcoma virus (SSV), implying that SSV-transformation is mediated by a PDGF-like growth factor. This notion is supported by the finding that specific as well as nonspecific inhibitors of PDGF-action (PDGF antibodies and suramin, respectively) are efficient inhibitors of SSV-transformation and revert the transformed phenotype of SSV-transformed cells. Expression of the genes encoding the PDGF subunits and production of PDGF-like growth factors is a common feature of human sarcoma cell lines, suggesting a role of PDGF in the pathogenesis of sarcomas, although direct support in favor of this notion is lacking. An involvement of PDGF in autocrine and paracrine stimulation of normal cell growth is suggested by the finding that responsive (arterial smooth muscle cells and placental cytotrophoblasts) as well as nonresponsive (endothelial cells and macrophages) cells produce PDGF-like growth factors. In conclusion, PDGF-like growth factors may be widely implicated in normal as well as neoplastic growth processes.

Amino Acid Sequence↗
PubMed1986

Efficient reversion of simian sarcoma virus-transformation and inhibition of growth factor-induced mitogenesis by suramin.

Simian sarcoma virus, an acutely transforming primate retrovirus with capacity to induce gliomas and sarcomas in experimental animals, has acquired its transforming properties by transducing the cellular gene sequences that encode one of the constituent chains of platelet-derived growth factor. Suramin, a drug used in the treatment of trypanosomiasis and onchocerciasis, has previously been reported to inhibit the interaction of platelet-derived growth factor with its cell surface receptor. We show here that suramin efficiently reverts the simian sarcoma virus-induced transformed phenotype in human and rat fibroblasts and propose that this is due to neutralization of an externalized v-sis product. Moreover, we show that suramin inhibits the action of a broad spectrum of growth factors.

Animals↗
PubMed1986

Rat skeletal myoblasts and arterial smooth muscle cells express the gene for the A chain but not the gene for the B chain (c-sis) of platelet-derived growth factor (PDGF) and produce a PDGF-like protein.

It is shown here that the myogenic cell line L6J1, primary skeletal myoblasts, and primary adult arterial smooth muscle cells express the gene for the A chain but not the gene for the B chain (c-sis) of platelet-derived growth factor (PDGF). It is further demonstrated that conditioned media from L6J1 cultures contain material that (i) competes with 125I-labeled PDGF for binding to human fibroblasts, (ii) is specifically precipitated by antibodies against PDGF, and (iii) has a relative molecular mass comparable to that of PDGF and, after reduction, its constituent subunit chains. The secretion of PDGF-receptor-competing activity was at a maximum in exponentially growing cultures but remained at a high level also after the cells had become confluent, stopped dividing, and fused to form multinucleate myotubes. Similarly, it was previously demonstrated that adult rat arterial smooth muscle cells in primary culture produce a mitogenic protein with immunological and structural properties similar to PDGF. In accordance with these findings, it was recently shown that secretion of PDGF-like mitogens by a number of human tumor cell lines correlates with expression of the gene for the A chain rather than the B chain of PDGF. The results suggest that production of homodimers of PDGF A chains may stimulate proliferation of skeletal myoblasts and arterial smooth muscle cells in an autocrine or paracrine manner. This could fulfill important functions during myogenesis in the embryo as well as in tissue repair and atherogenesis in the adult.

Animals↗
PubMed1986

Human melanoma cell lines of primary and metastatic origin express the genes encoding the chains of platelet-derived growth factor (PDGF) and produce a PDGF-like growth factor.

Normal human melanocytes and five human melanoma cell lines were analyzed for production of platelet-derived growth factor (PDGF)-like activity. Three of the melanoma cell lines released an activity that inhibited binding of 125I-labeled PDGF to human foreskin fibroblasts and stimulated [3H]thymidine incorporation in such cells. These activities were inhibited by the addition of anti-PDGF antibodies. All three factor-producing cell lines were derived from the same patient--one originated from the primary tumor (WM 115), and two were from individual lymph-node metastases (WM 239A and WM 266-4). The factor produced by WM 266-4 cells was characterized biochemically in detail. Immunoprecipitated, the metabolically labeled factor migrated in NaDod-SO4/gel electrophoresis as a homogeneous Mr 31,000 species, which under reducing conditions was resolved into two species of Mr 16,500 and Mr 17,000, implying a dimeric structure of the molecule. The factor was purified to homogeneity. Analysis by reverse-phase high-pressure liquid chromatography of reduced and alkylated factor revealed an elution pattern identical to that of PDGF A chains. Thus, the native molecule appears to be a homodimer of PDGF A chains. Blot-hybridization analysis of poly(A)+ RNA from the cell lines with 32P-labeled PDGF A chain and B chain (SIS product) cDNA probes revealed a relative abundance of B chain transcripts in the cell line originating from the primary tumor tissue only but expression of A chain in all three cell lines. We conclude that the two structural genes encoding each of the subunit chains of PDGF can be expressed in human melanoma cells and that the two genes can be independently expressed in such cells.

Cell Line↗
PubMed1986

Location of epidermal growth factor receptors on porcine thyroid follicle cells and receptor regulation by thyrotropin.

By using a direct electron microscopic autoradiographical technique, we were able to demonstrate that the epidermal growth factor (EGF) receptor is asymmetrically located on polarized porcine thyroid follicle cells cultured on a collagen gel matrix. More than 80% of autoradiographical grains from [125I]EGF were associated with the basolateral cell surface compared to only about 2.5% at the apical cell margin. EGF stimulated growth only when the collagen gels were floating, indicating a basolateral location of functional EGF receptors. Preincubation with TSH or another stimulator of cAMP synthesis, forskolin, increased binding of [125I]EGF to the cells. Moreover, the subsequent mitogenic response to EGF was potentiated by such pretreatment. The results are in accordance with the hypothesis that TSH potentiates the response to EGF in the stimulation of thyroid growth.

Animals↗
PubMed1986

Clonal variation in the production of a platelet-derived growth factor-like protein and expression of corresponding receptors in a human malignant glioma.

Two cell lines (U-343 MG and U-343 MGa) with different phenotypic characteristics were established from the same human glioblastoma multiforme biopsy. Previous studies have shown that a clonal derivative (Cl 2) of the U-343 MGa line produces a PDGF-like growth factor. In the present investigation glioma PDGF production and 125I-PDGF binding were found to be differently expressed in U-343 MG, U-343 MGa, and U-343 MGa Cl 2 cultures, providing evidence for a clonal variation in these properties. In order to investigate this point further, several clones were derived from low (23 clones) and high (30 clones) passage U-343 MGa cultures, as well as from U-343 MGa Cl 2 cells (30 clones). The clones could be divided into 4 groups according to morphology and growth pattern. A determination of the amount of PDGF receptor competing activity in serum-free conditioned media gave evidence for a clonal variation in the production of glioma PDGF, corresponding to 0-87 ng of authentic PDGF per ml. There was also a considerable range in 125I-PDGF binding (0-44 fmol of tracer bound per 10(6) cells). Scatchard plots performed on two clones confirmed the presence of saturable, high affinity PDGF receptors. High passage cultures were found to give rise to a higher number of high producing clones than did low passage cultures. There appeared to be a negative correlation between production of glioma PDGF and binding of 125I-PDGF, probably due to the receptor blocking activity of the endogenous growth factor. However, the presence of clones, apparently devoid of both glioma PDGF production and 125I-PDGF binding, suggests a true clonal variation in these two parameters. The growth rate in serum-free medium was found to correlate fairly well to the extent of glioma PDGF production. Production of glioma PDGF was found to have a morphological correlate and be most prominent among clones of "immature" looking, tightly growing cells. Clones that had large star-shaped cells with some resemblance to normal glia-like cells in culture were found to have a low production and a high 125I-PDGF binding capacity.

Cell Cycle↗
PubMed1985

Expression of the c-sis gene and secretion of a platelet-derived growth factor-like protein by simian virus 40-transformed BHK cells.

SV40-transformed BHK cells were shown to express two transcripts, of 3.5 kb and 2.0 kb, that hybridised to a human c-sis probe. Antibodies directed against human PDGF specifically recognized a 31 kDa protein in SV40/BHK cell conditioned medium, which upon reduction was split into 16 kDa species. Unfractionated conditioned medium and one of two growth factors isolated from SV40/BHK cells competed with 125I-PDGF for binding to its receptor. The present communication thus provides compelling evidence that an SV40/BHK cell-derived growth factor is a hamster equivalent to human PDGF.

Animals↗
PubMed1985

Similar action of platelet-derived growth factor and epidermal growth factor in the prereplicative phase of human fibroblasts suggests a common intracellular pathway.

We have investigated the effects of platelet-derived growth factor (PDGF) and epidermal growth factor (EGF) in the prereplicative phase of human foreskin fibroblasts cultured under defined conditions in serum-free MCDB 105 medium. Specific antisera against PDGF and EGF were used to inhibit the stimulation after certain incubation times. It was found that PDGF or EGF had to be present during the major part of the G0/G1 phase (greater than 8 h) in order to cause any appreciable commitment to DNA synthesis; half maximal stimulations were obtained after 9 h and 11 h of incubations with PDGF and EGF, respectively. When tested during a suboptimal period of time (6 h), neither an increase in concentration of PDGF or EGF, nor the addition of both growth factors simultaneously caused any appreciable stimulation of DNA synthesis. However, a suboptimal pulse of PDGF, followed by a suboptimal pulse of EGF, or vice versa, led to commitment to DNA synthesis. This finding indicates that PDGF and EGF, at least in part, induce similar intracellular events that transmit the mitogenic signal.

Cell Division↗
PubMed1985

Coexpression of the sis and myc proto-oncogenes in developing human placenta suggests autocrine control of trophoblast growth.

First trimester human placentas actively express the sis proto-oncogene, the structural gene for the B chain of platelet-derived growth factor (PDGF). Using the in situ hybridization technique, the 4.2 kb c-sis transcript has been localized to the cytotrophoblastic component, especially the highly proliferative and invasive cytotrophoblastic shell, paralleling the distribution of c-myc transcripts in early placenta. Explants of first trimester placenta release significant levels of PDGF-like activity into the medium under apparent developmental control. Moreover, cultured trophoblasts display abundant high-affinity PDGF receptors and respond to exogenous authentic PDGF by an activation of the c-myc gene and DNA synthesis. The developing human placenta may therefore represent a case of autocrine growth regulation in a normal tissue, in which cells bearing receptors for a growth factor can also synthesize and respond to that factor.

Cell Line↗
PubMed1985

Inhibition of cyclic AMP formation by iodide in suspension cultures of porcine thyroid follicle cells.

In the present study porcine thyroid cells in suspension cultures were employed to investigate the suppressive effect of iodide on adenylate cyclase under basal conditions and following incubation with TSH, PGE1, cholera toxin and forskolin. Within 30 min of incubation with iodide (half-maximal effect 10(-5) M), inhibition was established and remained unchanged up to 40 h of culture. The inhibitory action was abolished by methimazole. TSH, PGE1, cholera toxin and forskolin stimulated cAMP accumulation 10-, 3-, 24- and 22-fold, respectively. Iodide pretreatment reduced basal cAMP levels and also made the cells less sensitive to stimulation by the various agents. High concentrations of TSH or PGE1 could not overcome the suppressive influence of iodide, whereas with high concentrations of cholera toxin and forskolin the reduction in cAMP levels in iodide-treated cultures was less pronounced. Membranes isolated from iodide-treated cultures produced significantly lower amounts of cAMP compared to control membranes. Furthermore, iodide did not inhibit basal or forskolin-stimulated cAMP production in human fibroblasts. The results demonstrate that iodide via an iodination-dependent mechanism influences cAMP generation in thyroid cells. It is suggested that the inhibitory activity, which has a long half-life, involves stable modification of the membrane-localized catalytic unit of adenylate cyclase such that its activation by the regulatory unit is rendered less efficient.

Adenylyl Cyclase Inhibitors↗
PubMed1985

Thyrotropin modulates EGF receptor function in porcine thyroid follicle cells.

Porcine thyroid follicle cells in monolayer cultures were shown to contain one single class of high-affinity EGF receptors with Kd = 4.5 X 10(-10) M and approximately 20 000-25 000 receptors per cell. Suspension cultures of aggregated follicle cells, exposed to TSH for 3 days, showed a 3-fold increase in [125I]EGF binding. Scatchard analysis demonstrated that this was due to an increase in receptor number. Other cAMP-elevating agents (cholera toxin, dibutyryl cAMP, forskolin) induced a similar effect. In suspension cultures, preincubation with TSH or cholera toxin for 2 days reduced the subsequent [3H]thymidine incorporation. This inhibition was overcome by a low concentration of EGF (0.1 ng/ml). At higher concentrations of EGF (1-10 ng/ml) the incorporation of [3H]thymidine was potentiated 2-3-fold in cultures preexposed to TSH or cholera toxin. The results demonstrate the presence of a high-affinity EGF receptor in porcine thyroid follicle cells. Receptor expression, as well as responsiveness to the mitogenic action of EGF, is modulated in vitro by TSH, through a cAMP-dependent process.

Animals↗