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J Blay

Publications and source records attributed to J Blay.

27 records · Page 2Linked to original sources
PubMed1989

Heterologous regulation of EGF receptor function in cultured aortic smooth muscle cells.

A10 cultured smooth muscle cells from rat embryonic thoracic aorta bound 125I-labelled epidermal growth factor (125I-EGF), and responded to EGF by an increase in DNA synthesis. Scatchard analysis of binding data obtained at 4 degrees C showed curvilinearity consistent with there being two affinity classes of binding site. The amount of 125I-EGF that bound was decreased by treatment of the A10 cells at 37 degrees C with [Arg8]vasopressin or with 5-hydroxytryptamine (5-HT). Scatchard analysis of binding (at 4 degrees C after pretreatment at 37 degrees C) revealed this effect to be due to a loss of the high-affinity component of 125I-EGF binding, with no change in total receptor number. The presence of vasopressin or 5-HT raised the concentration of EGF required for the stimulation of DNA synthesis. Cultured A10 aortic smooth muscle cells therefore have receptors for EGF that mediate an increase in cell proliferation. EGF receptor function is modified by vasopressin and 5-HT, probably as a consequence of their effects on EGF receptor affinity.

Animals↗
PubMed1989

Epidermal growth factor stimulation of prostacyclin production by cultured aortic smooth muscle cells: requirement for increased cellular calcium levels.

We have examined the ability of epidermal growth factor (EGF) to regulate prostacyclin production by cultured A10 smooth muscle cells. EGF by itself had no effect on prostacyclin production, but it augmented the response to arg8-vasopressin. An AGF stimulation of prostacyclin production was also observed in the presence of the calcium ionophore A23187; it therefore seemed likely that the key event required for EGF to stimulate prostacyclin production might be an increase in the available cellular Ca2+. Studies with 45Ca2+ showed that vasopressin both mobilised Ca2+ from intracellular stores and increased the influx of extracellular Ca2+ into A10 cells. The increase in prostacyclin production caused by vasopressin and the augmentation by EGF were both abolished by TMB-8, an antagonist of Ca2+ mobilisation, by EGTA, a chelator of Ca2+ ions, or by incubating cultures in the absence of added Ca2+. These results were consistent with a central role for Ca2+ in the responses and showed that both intracellular and extracellular sources of Ca2+ were important for the triggering of prostacyclin production. The increases in prostacyclin production were only marginally affected by nifedipine, and no responses were seen (either in the absence or presence of EGF) when KCl was used to depolarise the cell membrane. These data indicated that uptake of Ca2+ ions via voltage-dependent channels was unlikely to be a major factor in the stimulation of prostanoid production. We conclude that the ability of EGF to stimulate prostacyclin production in A10 smooth muscle cells depends upon a concurrent stimulus that will increase available intracellular Ca2+ levels.

6-Ketoprostaglandin F1 alpha↗
PubMed1986

Contradistinctive growth responses of cultured rat intestinal epithelial cells to epidermal growth factor depending on cell population density.

The growth response of cultured rat intestinal epithelial (RIE-1) cells to epidermal growth factor (EGF) depends on the cell population density. EGF stimulated the proliferation of RIE-1 cells in dense cultures, but inhibited the proliferation of cells growing at low population densities. In contrast, insulin enhanced RIE-1 cell growth irrespective of the population density. The tumour promoter 12-0-tetradecanoylphorbol 13-acetate (TPA), like EGF, inhibited the proliferation of low-density RIE-1 cells, but differed from EGF in that it did not stimulate the growth of dense cultures.

Animals↗
PubMed1985

Functional receptors for epidermal growth factor in an epithelial-cell line derived from the rat small intestine.

Epidermal growth factor (EGF) regulates the proliferation of cells of a rat intestinal epithelial-cell line (RIE-1) in culture. Confluent RIE-1 cells were stimulated to proliferate by EGF with a half-maximal effect at 1-2 ng/ml. In contrast, the growth of sparse RIE-1 cells was inhibited by the growth factor. Binding studies at 4 degrees C with 125I-EGF identified two classes of binding sites for EGF on RIE-1 cells, one of high affinity (KD = 1.8 X 10(-10)M; 1.8 X 10(4) receptors/cell) and one of lower affinity (KD = 5.2 X 10(-9)M; 6.3 X 10(4) receptors/cell). After binding to the cells at 37 degrees C, 125I-EGF was rapidly internalized and subsequently degraded. Degradation products were released into the medium after a lag of 15-30 min. The degradation of 125I-EGF did not occur at 4 degrees C and was inhibited at 37 degrees C by chloroquine, methylamine or NH4Cl, but not by colchicine. Exposure of RIE-1 cells to EGF caused a time- and dose-dependent loss of EGF receptors from the cell surface. The recovery of receptors after the removal of EGF was retarded in the absence of serum and prevented by the presence of cycloheximide or actinomycin D. Sodium dodecyl sulphate/polyacrylamide-gel electrophoresis separation of the 125I-EGF-receptor complex from RIE-1 cells after covalently cross-linking with disuccinimidyl suberate indicated a receptor of Mr congruent to 160 000. The demonstration of functional EGF receptors in this cell line provides further evidence that EGF may regulate intestinal-epithelial-cell physiology.

Animals↗
PubMed1985

Epidermal growth factor promotes the chemotactic migration of cultured rat intestinal epithelial cells.

The RIE-1 cell line is an untransformed, epithelial cell line derived from the rat small intestine. We report that epidermal growth factor (EGF), which regulates the proliferation of RIE-1 cells, also directs their movement. We measured cell migration through gelatin-coated filters in blind-well Boyden chambers. The migration of RIE-1 cells was stimulated up to approximately 100-fold by EGF, with a half-maximal response at 1-2 ng/ml and a maximal effect at 10 ng/ml. Further analysis showed that the RIE-1 cells responded directionally to a gradient of EGF in solution. Other growth factors tested did not stimulate RIE-1 cell migration, and EGF did not stimulate the migration of fibroblasts in this assay. We conclude that EGF is a potent and specific chemo-attractant for RIE-1 intestinal epithelial cells and suggest that EGF might influence epithelial cell migration in vivo.

Animals↗
PubMed1984

Reduction of epidermal growth factor receptor affinity by heterologous ligands: evidence for a mechanism involving the breakdown of phosphoinositides and the activation of protein kinase C.

The tetradecapeptide bombesin converts epidermal growth factor (EGF) receptors on Swiss 3T3 cells from a high affinity state (KD = 9.8 X 10(-11)M) to a lower affinity state (KD = 1.8 X 10(-9)M). This conversion occurs when the cells are incubated with bombesin at 37 degrees C but not when incubated at 4 degrees C. Previously, a number of other (chemically unrelated) cell growth-promoting peptides and polypeptides have been shown to induce a similar indirect, temperature-dependent reduction of EGF receptor affinity. We have now demonstrated that hormones and growth factors which cross-regulate EGF receptor affinity in Swiss 3T3 cells have a common ability to stimulate the breakdown of phosphoinositides in these cells. We propose that the reduction of EGF receptor affinity is a consequence of the activation of protein kinase C by the diacylglycerol generated by this breakdown. In support of this proposal we have found that exogenously added diacylglycerol reduces the affinity of the Swiss 3T3 cell EGF receptor.

Animals↗
PubMed1984

Characterization of an epithelioid cell line derived from rat small intestine: demonstration of cytokeratin filaments.

A cell line (RIE-1) of epithelioid morphology was established from rat small intestine. RIE-1 cells possess the normal rat diploid number of chromosomes (42) up to at least passage 20. Both early (less than 10) and late (greater than 20) passage cells are non-tumourigenic and do not form colonies in soft agarose. RIE-1 cells bind two epithelial-specific monoclonal antibodies. In addition, the cells exhibit a network of cytoskeletal filaments which are labelled by an antiserum to keratin. These findings provide strong evidence for an epithelial origin of the RIE-1 cell line.

Animals↗