Specific receptors for platelet-derived growth factor on cells derived from connective tissue and glia.

Abstract

A cellular receptor for platelet-derived growth factor (PDGF) was demonstrated by incubation of "MI-labeled PDGF with human foreskin fibroblast cultures followed by liberation of cell-bound radioactivity with Triton X-100. The cellular binding of labeled PDGF in the presence of increasing amounts ofunlabeled PDGF showed saturation; Scatchard analysis ofbinding data indicated a single class of receptors having kd = 1 x 10-9 M. The number of PDGF binding sites was -3 X 105/cell. Labeled PDGF binding reached an apparent equilibrium after 3 hr at 4TC. At 370C, it passed a maximum after 30 min and then decreased with time due to degradation of the tracer. A large excess of unlabeled PDGF reduced labeled PDGF binding by more than 90% whereas similar doses of epidermal growth factor, fibroblast growth factor, or insulin had no effect. It was concluded that PDGF did not share receptors with these factors. PDGF receptors were found on skin fibroblasts, normal and malignant glial cells, smooth muscle cells, and 3T3 cells but not on epithelial-derived cells, neuroblastoma cells, endothelial cells, or peripheral lymphocytes. As only the receptor-positive cells-i.e., the connective tissue-and glia-derived cells-are responsive to stimulation with PDGF, these findings imply a functional significance of the PDGF receptor.The human platelet-derived growth factor (PDGF) is a Mr 30,000 cationic protein, probably composed of two polypeptide chains linked by disulfide bonds (1, 2). PDGF is stored in the a-granules of platelets (3,4) and released during the plateletrelease reaction (3-5). The factor stimulates the proliferation of various cultured cells, such as human glial cells (1, 6, 7), and a number of connective tissue-derived cells, e.g., arterial smooth muscle cells (8, 9) human fibroblasts (10, 11), and mouse 3T3 cells (12)(13)(14)(15). For these cell types, PDGF actually constitutes the major part of the growth-promoting activity of serum.The access to pure "2I-labeled PDGF (1) has made it possible to study the interaction between PDGF and its target cells. In this paper, we describe a specific binding of "2I-labeled PDGF to cultured cells. A preliminary report has been presented elsewhere (16). MATERIALS AND METHODSGrowth Factors. PDGF was purified from fresh platelets as described (2). The final product was >90% pure as estimated from analytical NaDodSO4polyacrylamide gel electrophoresis.Epidermal growth factor (EGF) and fibroblast growth factor (FGF) were purchased from Collaborative Research (Waltham, MA). Insulin was obtained from Vitrum (Stockholm, Sweden).Radiolabeing of PDGF. PDGF was labeled with "5I according to Hunter and Greenwood (17) as described (1). Five micrograms of PDGF was labeled with 0.5 mCi (1 Ci = 3.7 X 101" becquerels) of '25I. 125I-Labeled PDGF was separated from unbound radioactivity on a Sephadex G-25 column eluting with 1 M acetic acid containing serum albumin at 1 mg/ml (1). The specific activity was 16,000 cpm/ng, corresponding to -0.2 atoms of iodine per molecule of PDGF. The present ...

Human breast carcinoma desmoplasia is PDGF initiated

“…PDGF was labeled with 125 I (Heldin et al, 1981) and PDGF receptor competing activity in fractionated W9 CM was measured as inhibition of binding of [ 125 I]PDGF to con¯uent cultures of CRL 1477 ®broblasts as described previously (Peres et al, 1987;Bronzert et al, 1987). Puri®ed PDGF and EGF served as competition controls.…”

Identification of Tyr-762 in the platelet-derived growth factor α-receptor as the binding site for Crk proteins

“…Platelet-derived growth factor (PDGF) is a potent mitogen for cells of mesenchymal origin, and has recently been shown to act also on other types of cells (Beits et al, 1991;Do et al, 1992;Heldin et al, 1981;Henriksen et al, 1993;Ra et al, 1988;Smits et al, 1991). It is implicated in normal embryonic development and wound healing, as well as in development of pathological conditions such as tumors, in¯ammatory diseases and atherosclerosis (Heldin and Westermark, 1994;Ross, 1993).…”

Kinetic Analysis of Platelet-derived Growth Factor Receptor/Phosphoinositide 3-Kinase/Akt Signaling in Fibroblasts

“…However, to our knowledge, this has not been observed for PDGF or other ligands that engage receptor tyrosine kinases. PDGF-BB binds to cell membranes or immobilized PDGF receptor extracellular domains with an observed K D ϳ 0.1-1 nM (11,39,(42)(43)(44), yet inhibition of cell responses at much higher concentrations has not been observed. NIH 3T3 cells were stimulated with 3 nM PDGF-BB for the times indicated.…”