Induction of Monocyte Chemoattractant Protein-1 Synthesis in Human Monocytes During Transendothelial Migration In Vitro

Abstract

Abstract Monocyte chemoattractant protein-1 (MCP-1, or monocyte chemotactic and activating factor) plays important roles in the recruitment of monocytes and thus in the development of atherosclerosis. In this study, we determined whether MCP-1 synthesis was induced by the cellular interaction between monocytes and endothelial cells during the process of transendothelial migration. We found that when human peripheral blood monocytes (2.5×10 6 cells) and umbilical vein endothelial cells (HUVECs; 5.0×10 5 cells) were cocultured for 5 hours, 7.9 ng/mL MCP-1 was secreted into the medium, whereas when the two were cultured separately, MCP-1 levels were 1.0 and 0.9 ng/mL, respectively. Furthermore, the use of interleukin-1β (IL-1β)–pretreated HUVECs in cocultures induced twice the levels of MCP-1 as in unstimulated HUVEC culture. Conditioned medium had transendothelial chemotactic activity for monocytes, and this activity was completely abolished by addition of anti–MCP-1 antibody. Although MCP-1 mRNA levels were very low or undetectable in HUVECs or monocytes alone, message could be detected after 2 hours of coculture in total mRNA preparations from both monocytes and HUVECs. mRNA levels increased by 4 hours and had declined slightly by 24 hours. The rapid induction of message suggests that cell contact between monocytes and HUVECs induces the de novo synthesis of MCP-1 protein. Anti–interleukin (IL)-1α/β and anti–tumor necrosis factor-α antibodies, or anti–lymphocyte function–associated antigen-1 and very late antigen-4 antibodies, had little or no inhibitory effects on MCP-1 secretion by cocultures. Immunohistochemistry revealed that monocytes adherent to or having migrated across unstimulated HUVEC monolayers as well as the HUVECs themselves expressed MCP-1 protein. However, nonadherent monocytes failed to express it. This finding suggests that the monocyte–endothelial cell adhesive interaction results in an MCP-1–inductive signal to each cell type. MCP-1 expression by migrated monocytes may indicate that monocytes are primed to produce MCP-1 during transmigration and can secrete it in normal tissue in which inflammatory cytokines that induce MCP-1 are otherwise absent.