Purification and identification of mouse lung microvessel endothelial cell-derived chemoattractant for lung-metastasizing murine RAW117 large-cell lymphoma cells: identification as mouse monocyte chemotactic protein 1.
We reported recently that medium conditioned with mouse lung microvessel endothelial cells possessed chemotactic activity for a highly lung-metastatic variant (L17) of the RAW117 murine large-cell lymphoma cell line but not for the poorly metastatic parental cells (P) or a liver-metastasizing variant (H10). The chemotactic factor was purified to homogeneity by a five-step procedure involving hydrophobic interaction, Cibacron blue F3GA affinity, metal affinity, anion exchange, and reversed phase chromatography, followed by preparative gel electrophoresis. The purified material appeared as a single broad band when analyzed by SDS-PAGE, with an average molecular weight of 26,000. The factor was cleaved by cyanogen bromide treatment, and a partial amino acid sequence of one of the cleaved polypeptides proved identical to mouse monocyte chemotactic protein 1 (mMCP-1/JE). The amino acid composition of the factor also indicated similarity to mMCP-1/JE. Separately purified mMCP-1/JE significantly stimulated the chemotactic migration of RAW117 cells (L17 >> H10, P). When recombinant human monocyte chemotactic protein 1 was compared to the purified endothelial cell chemotactic factor as a chemoattractant, similar migratory responses were observed in the RAW117 sublines. The chemotactic activity for L17 cells was significantly reduced from lung microvessel endothelial cell-conditioned medium after treatment with anti-mouse MCP-1 antibody. In contrast, the migration-stimulating activity of liver sinusoidal endothelial cell-conditioned medium to H10 cells was not affected by anti-mouse MCP-1. A major function of mMCP-1/JE is to recruit monocytes to inflammatory sites, and our results suggest that mMCP-1/JE also facilitates lymphoma lung invasion and metastasis.
Wakabayashi
H
H
Department of Tumor Biology, University of Texas M. D. Anderson Cancer Center, Houston 77030, USA.
Cavanaugh
P G
PG
Nicolson
G L
GL
D000595:Amino Acid Sequence; D000818:Animals; D018932:Chemokine CCL2; D002630:Chemotactic Factors / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology; D004730:Endothelium, Vascular / Q000737:chemistry*; D008168:Lung / Q000098:blood supply / Q000737:chemistry*; D008175:Lung Neoplasms / Q000556:secondary*; D016403:Lymphoma, Large B-Cell, Diffuse / Q000473:pathology*; D051379:Mice; D008833:Microcirculation / Q000737:chemistry; D008969:Molecular Sequence Data
D018932:Chemokine CCL2; D002630:Chemotactic Factors