Antisense oligomers for selective suppression of MCP-1 synthesis in human pulmonary endothelial cells

Endothelial synthesis of the C-C chemokine monocyte chemotactic protein-1 ( MCP-1) has been implicated in the regulation of monocyte recruitment for ex travascular pools under both physiologic and inflammatory conditions. We de signed and characterized five antisense phosphorothioate oligodeoxynucleoti des (PS-ODN) targeting MCP-1 secretion by human pulmonary artery endothelia l cells (HPAEC) and pulmonary microvascular endothelial cells (HMVEC-L). Th e most effective PS-ODN (MCP-1 AS 2) dose-dependently suppressed the secret ion of MCP-1 but not the secretion of the C-X-C chemokine interleukin-8 (IL -8) in both HPAEC and HMVEC-L in the nanomolar concentration range. Mismatc h controls bearing 2 or 4 bp substitutions showed markedly reduced inhibito ry capacity. MCP-1 mRNA levels were not affected even at the highest PS-ODN doses employed (ribonuclease protection assay), suggesting a translational arrest of MCP-1 production. Accordingly, PS-ODN exhibited no nonspecific s ide effects on immediate-early gene regulation of the transcription factor nuclear factor-kappa B (NF-kappa B), as analyzed by gel shift assays. Antis ense pretreatment of HPAEC reduced the monocyte chemotactic bioactivity lib erated from tumor necrosis factor-alpha (TNF-alpha)-activated endothelial c ells (EC) and reduced the TNF-alpha-induced transendothelial monocyte migra tion. We conclude that nanomolar concentrations of specific antisense oligo deoxynucleotides effectively inhibit human endothelial MCP-1 synthesis and may thus provide a rational approach to modulate monocyte recruitment under inflammatory conditions.