Abstract
OBJECTIVE: Monocyte chemoattractant protein-1 (MCP-1), encoded by the CCL2 gene, plays an important role in the initiation and progression of atherosclerosis. Ea.hy 926 endothelial cells grown under low folate conditions (LO cells) synthesize more MCP-1 mRNA and secrete more MCP-1 protein than folate-replete control cells (HI cells). We investigated the mechanisms underlying the modulation of MCP-1 expression by long-term "folate stress". METHODS AND RESULTS: CCL2 transcription, assessed using promoter-reporter assays, is up-regulated in LO cells relative to HI cells, whereas MCP-1 mRNA stability is unchanged. This quantitative transcriptional bias under chronic low folate conditions is not attributable to differences in active NF-kappaB, but is associated with elevated levels of both total p38 and phospho-p38 that are detectable by Western immunoblotting. Transient, acute methotrexate-mediated folate depletion or exposure to high concentrations of homocysteine (Hcy) had no effect on MCP-1 synthesis by Ea.hy 926 cells. The p38 inhibitor SB-203580 abolished the excess MCP-1 production by LO cells. The quantitative transcriptional bias of CCL2 in LO cells was retained following massive induction by TNF-alpha. CONCLUSION: During long-term folate stress, p38 is the primary determinant of CCL2 transcription. Long-term folate insufficiency "primes" Ea.hy 926 endothelial cells to have a quantitatively more vigorous response to cytokine-mediated inflammatory stress.