cDNA cloning of guinea pig monocyte chemoattractant protein-1 and expression of the recombinant protein

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Abstract

Monocyte chemoattractant protein-1 (MCP-1) cDNA was cloned from guinea pig spleen cells stimulated with Con A. The cDNA comprised 647 bp with an open reading frame that encoded a 120 amino acid protein. The sequence similarity of the first 99 amino acids to human MCP-1 is 56%. Recombinant guinea pig MCP-1 was expressed in COS-7 cells, then purified by a three step procedure with orange A-agarose, carboxymethyl-HPLC, and reversed phase-HPLC. The purified protein was found around 25 kDa as a broad bard on a polyacrylamide gel under reducing conditions. Guinea pig MCP-1 attracted about 34% of input human monocytes at 5 x 10-9 M. Guinea pig peritoneal exudate macrophages migrated toward guinea pig MCP-1 dose dependently, but only 1% of input cells responded to guinea pig MCP-1 at its optimal concentration of 5 x 10-9 M. Human MCP-1 attracted 1% of input guinea pig peritoneal exudate macrophages at its highest concentration of 2.5 x 10-8 M. Neither human nor guinea pig MCP-1 attracted guinea pig peritoneal resident macrophages. These results suggest that monocytes lose responsiveness to MCP-1 after differentiating to macrophages. Finally, intradermal injection of the recombinant protein into guinea pigs caused marked macrophage infiltration. Cloned and expressed guinea pig MCP-1 will help in studying the role of MCP-1 in vivo.

Original languageEnglish
Pages (from-to)5025-5032
Number of pages8
JournalJournal of Immunology
Volume150
Issue number11
Publication statusPublished - Jan 1 1993
Externally publishedYes

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology

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