Secretion by human fibroblasts of monocyte chemoattractant protein-1, the product of gene JE

Teizo Yoshimura, E. J. Leonard

Research output: Contribution to journalArticle

118 Citations (Scopus)

Abstract

We recently purified human monocyte chemoattractant protein-1 (MCP-1) from culture fluids of either human glioma cell lines or mitogen-stimulated human peripheral blood mononuclear leukocytes. It has now been shown that MCP-1 is the product of the gene JE, which was first recognized by its expression in fibroblasts stimulated with platelet-derived growth factor (PDGF). We therefore studied secretion of MCP-1 by three human fibroblast cell lines. Monocyte chemotactic activity was found in culture fluids of all three lines after growth to confluence in DMEM-10% FCS, and the amounts secreted per cell were comparable for the three lines. The MRC-5 line was chosen for further study. Monocyte chemotactic activity secretion by confluent MRC-5 cultures continued after a switch to serum-free medium and was not inhibited by anti-PDGF antibody, indicating that secretion may not have been caused by autocrine release of PDGF. When concentrated serum-free MRC-5 culture fluid was injected into an HPLC gel filtration column, only one chemotactic activity peak was observed, which was in the same location as glioma-derived MCP-1. The activity was completely absorbed out by an anti-MCP-1 affinity column, which indicates that all the chemotactic activity in MRC-5 culture fluid was accounted for by MCP-1. PDGF caused a marked increase in chemotactic activity over that found in serum-free culture fluid of MRC-5 or 501T cells. Immunoprecipitation by anti-human MCP-1 showed two bands, corresponding to the two forms of MCP-1 previously described (MCP-1α and β); and the amounts increased in response to PDGF stimulation. Thus, the reported in human fibroblast EJ mRNA in response to PDGF-containing serum stimulation is reflected in increased secretion of the MCP-1 gene product.

Original languageEnglish
Pages (from-to)2377-2383
Number of pages7
JournalJournal of Immunology
Volume144
Issue number6
Publication statusPublished - 1990
Externally publishedYes

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Chemokine CCL2
Platelet-Derived Growth Factor
Fibroblasts
Genes
Glioma
Monocytes
Serum
Mononuclear Leukocytes
Cell Line
Serum-Free Culture Media
human CCL2 protein
Mitogens
Immunoprecipitation
Gel Chromatography
High Pressure Liquid Chromatography
Messenger RNA
Antibodies
Growth
Proteins

ASJC Scopus subject areas

  • Immunology

Cite this

Secretion by human fibroblasts of monocyte chemoattractant protein-1, the product of gene JE. / Yoshimura, Teizo; Leonard, E. J.

In: Journal of Immunology, Vol. 144, No. 6, 1990, p. 2377-2383.

Research output: Contribution to journalArticle

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abstract = "We recently purified human monocyte chemoattractant protein-1 (MCP-1) from culture fluids of either human glioma cell lines or mitogen-stimulated human peripheral blood mononuclear leukocytes. It has now been shown that MCP-1 is the product of the gene JE, which was first recognized by its expression in fibroblasts stimulated with platelet-derived growth factor (PDGF). We therefore studied secretion of MCP-1 by three human fibroblast cell lines. Monocyte chemotactic activity was found in culture fluids of all three lines after growth to confluence in DMEM-10{\%} FCS, and the amounts secreted per cell were comparable for the three lines. The MRC-5 line was chosen for further study. Monocyte chemotactic activity secretion by confluent MRC-5 cultures continued after a switch to serum-free medium and was not inhibited by anti-PDGF antibody, indicating that secretion may not have been caused by autocrine release of PDGF. When concentrated serum-free MRC-5 culture fluid was injected into an HPLC gel filtration column, only one chemotactic activity peak was observed, which was in the same location as glioma-derived MCP-1. The activity was completely absorbed out by an anti-MCP-1 affinity column, which indicates that all the chemotactic activity in MRC-5 culture fluid was accounted for by MCP-1. PDGF caused a marked increase in chemotactic activity over that found in serum-free culture fluid of MRC-5 or 501T cells. Immunoprecipitation by anti-human MCP-1 showed two bands, corresponding to the two forms of MCP-1 previously described (MCP-1α and β); and the amounts increased in response to PDGF stimulation. Thus, the reported in human fibroblast EJ mRNA in response to PDGF-containing serum stimulation is reflected in increased secretion of the MCP-1 gene product.",
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