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Title: Characterization of epidermal growth factor receptors on plasma membranes isolated from rat gastric mucosa

Abstract

The binding of human epidermal growth factor (hEGF), beta-urogastrone, to plasma membranes isolated from rat gastric mucosa was studied to characterize gastric EGF receptors. The binding of ({sup 125}I)hEGF was temperature dependent, reversible, and saturable. A single class of binding sites for EGF with a dissociation constant of 0.42 nM and maximal binding capacity of 42 fmol/mg protein was suggested. There was little change in the binding of ({sup 125}I)hEGF upon addition of peptide hormones (secretin, insulin), antiulcer drugs (cimetidine), or an ulcer-inducing reagent (aspirin). Cross-linking of ({sup 125}I)hEGF to gastric plasma membranes with the use of disuccinimidyl suberate resulted in the labeling of a protein of 150 kDa. These results indicate the presence of EGF receptors on plasma membranes of rat gastric mucosa.

Authors:
; ; ;  [1]
  1. (Kyoto Univ. Hospital (Japan))
Publication Date:
OSTI Identifier:
6875205
Resource Type:
Journal Article
Resource Relation:
Journal Name: Pharmaceutical Research; (USA); Journal Volume: 7:6
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; GROWTH FACTORS; RECEPTORS; CHEMICAL COMPOSITION; ACETYLSALICYLIC ACID; ANTIPYRETICS; BIOCHEMICAL REACTION KINETICS; BIOLOGICAL HALF-LIFE; DOGS; EPIDERMIS; ESCHERICHIA COLI; GASTROINTESTINAL TRACT; IN VITRO; INSULIN; IODINE 125; MAN; MUCOUS MEMBRANES; PROTEINS; RATS; REAGENTS; SECRETIN; TEMPERATURE DEPENDENCE; TRACER TECHNIQUES; ANALGESICS; ANIMAL TISSUES; ANIMALS; BACTERIA; BETA DECAY RADIOISOTOPES; BODY; CARBOXYLIC ACIDS; CENTRAL NERVOUS SYSTEM DEPRESSANTS; DAYS LIVING RADIOISOTOPES; DIGESTIVE SYSTEM; DRUGS; ELECTRON CAPTURE RADIOISOTOPES; EPITHELIUM; HORMONES; HYDROXY ACIDS; INTERMEDIATE MASS NUCLEI; IODINE ISOTOPES; ISOTOPE APPLICATIONS; ISOTOPES; KINETICS; MAMMALS; MEMBRANE PROTEINS; MEMBRANES; MICROORGANISMS; MITOGENS; NUCLEI; ODD-EVEN NUCLEI; ORGANIC ACIDS; ORGANIC COMPOUNDS; ORGANS; PEPTIDE HORMONES; PRIMATES; RADIOISOTOPES; REACTION KINETICS; RODENTS; SKIN; TISSUES; VERTEBRATES 550201* -- Biochemistry-- Tracer Techniques

Citation Formats

Hori, R., Nomura, H., Iwakawa, S., and Okumura, K. Characterization of epidermal growth factor receptors on plasma membranes isolated from rat gastric mucosa. United States: N. p., 1990. Web. doi:10.1023/A:1015838816061.
Hori, R., Nomura, H., Iwakawa, S., & Okumura, K. Characterization of epidermal growth factor receptors on plasma membranes isolated from rat gastric mucosa. United States. doi:10.1023/A:1015838816061.
Hori, R., Nomura, H., Iwakawa, S., and Okumura, K. 1990. "Characterization of epidermal growth factor receptors on plasma membranes isolated from rat gastric mucosa". United States. doi:10.1023/A:1015838816061.
@article{osti_6875205,
title = {Characterization of epidermal growth factor receptors on plasma membranes isolated from rat gastric mucosa},
author = {Hori, R. and Nomura, H. and Iwakawa, S. and Okumura, K.},
abstractNote = {The binding of human epidermal growth factor (hEGF), beta-urogastrone, to plasma membranes isolated from rat gastric mucosa was studied to characterize gastric EGF receptors. The binding of ({sup 125}I)hEGF was temperature dependent, reversible, and saturable. A single class of binding sites for EGF with a dissociation constant of 0.42 nM and maximal binding capacity of 42 fmol/mg protein was suggested. There was little change in the binding of ({sup 125}I)hEGF upon addition of peptide hormones (secretin, insulin), antiulcer drugs (cimetidine), or an ulcer-inducing reagent (aspirin). Cross-linking of ({sup 125}I)hEGF to gastric plasma membranes with the use of disuccinimidyl suberate resulted in the labeling of a protein of 150 kDa. These results indicate the presence of EGF receptors on plasma membranes of rat gastric mucosa.},
doi = {10.1023/A:1015838816061},
journal = {Pharmaceutical Research; (USA)},
number = ,
volume = 7:6,
place = {United States},
year = 1990,
month = 6
}
  • Muscarinic cholinergic receptors have been classified into subtypes based on their high (M-1 subtype) or low (M-2 subtype) affinities for the nonclassic antagonist pirenzepine, and this classification has important experimental and therapeutic implications. Because muscarinic receptors are abundant in the airways where they mediate several different cellular responses, the goal of this study was to characterize the affinities of pirenzepine for the muscarinic receptors in bovine tracheal mucosa and smooth muscle. After isolating membrane particulates from mucosa and smooth muscle, as well as from bovine cerebral cortex (a known source of M-1 receptors), we used /sup 3/H-quinuclidinyl benzilate to labelmore » muscarinic receptors in the particulates and performed competition radioligand binding assays in the presence of either atropine or pirenzepine. Receptors from all 3 tissues (mucosa, smooth muscle, and cerebral cortex) were of a relatively uniform affinity for atropine (range of KI values: 0.8 +/- 0.4 X 10(-9) to 2.4 +/- 1.7 X 10(-9) M), as would be predicted for this classic muscarinic antagonist. By contrast, affinities for pirenzepine differed depending on the tissue. In cerebral cortex, the majority of receptors were of high affinity for pirenzepine (KI = 1.8 +/- 1.4 X 10(-8) M). In both mucosa and smooth muscle, receptors were of low affinity for pirenzepine (Kl = 4.8 +/- 0.4 to 6.9 +/- 3.8 X 10(-7) M). We conclude that muscarinic cholinergic receptors in bovine tracheal mucosa and smooth muscle are predominantly of the M-2 subtype.« less
  • Receptors for insulin-like growth factor I (IGF-I) and epidermal growth factor (EGF) were localized and characterized in eight samples of human meningioma (four fibrous, two meningothelial, and two angioblastic types), using quantitative autoradiographic techniques. Effects of both growth factors on deoxyribonucleic acid (DNA) synthesis in the cultured meningioma cells were examined. High numbers of specific binding sites for both IGF-I and EGF were homogeneously present in tissue sections derived from fibrous and meningothelial types of meningiomas, whereas binding sites for these growth factors were not detectable in adjacent leptomeninges. While relatively large numbers of IGF-I binding sites were located inmore » the wall of the intratumoral vasculature, the number of binding sites in the stromal component was lower in angioblastic-type meningiomas, including a low number of EGF binding sites detected only in the stromal portion. Scatchard analysis revealed the presence of a single class of high-affinity binding sites for both IGF-I and EGF in the meningiomas examined (dissociation constant (Kd) = 0.6 to 2.9 nM, and the maximum number of binding sites (Bmax) = 16 to 80 fmol/mg for IGF-I; and Kd = 0.6 to 4.0 nM, Bmax = 3 to 39 fmol/mg for EGF). Both growth factors increased the synthesis of DNA, in a dose-dependent manner, as measured by 3H-thymidine incorporation. The combination of IGF-I and EGF synergistically stimulated the synthesis of DNA, and the effects seen with 10% fetal bovine serum could be reproduced at a concentration of 10(-10) M. These observations can be interpreted to mean that both IGF-I and EGF may be involved in the growth modulation of meningiomas, possibly through paracrine or autocrine mechanisms.« less
  • Epidermal growth factor (EGF) is present in high concentrations in milk, salivary, and pancreaticobiliary secretions. EGF, delivered to the intestinal lumen by these fluids, appears to influence intestinal proliferation. Because EGF exerts its mitogenic effect through binding to specific membrane-bound receptors, binding studies of {sup 125}I-labeled EGF to purified microvillus membrane (MVM) preparations fetal, newborn, and adult rat small intestine were performed. Using the membrane filter technique, binding of {sup 125}I-EGF to adult MVM was specific, saturable, and reversible. Adult and fetal MVM binding was rapid and reached a plateau after 30 min at both 20 and 37{degree}C. No bindingmore » was detected at 4{degree}C. Specific binding increased linearly from 0 to 75 {mu}g MVM protein. Scatchard analysis revealed a single class of receptors in fetal and adult MVM with an association constant of 1.0 {+-} 0.35 {times} 10{sup 9} and 2.3 {+-} 1.6 {times} 10{sup 9} M{sup {minus}1}, respectively. Binding capacity was 435.0 {+-} 89 and 97.7 {+-} 41.3 fmol {sup 125}I-EGF bound/mg MVM protein for fetal and adult MVM, respectively. Newborn MVM binding was negligible. After binding, cross-linking utilizing disuccinimidyl suberate, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis, autoradiography revealed a 170-kDa receptor. These data demonstrate specific receptors for EGF on MVM of rat small intestine and, thus, suggest a mechanism for the intraluminal regulation of enterocyte proliferation by EGF.« less
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