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

Abstract

Endothelial synthesis of the C-C chemokine monocyte chemotactic protein-1 (MCP-1) has been implicated in the regulation of monocyte recruitment for extravascular pools under both physiologic and inflammatory conditions. We designed and characterized five antisense phosphorothioate oligodeoxynucleotides (PS-ODN) targeting MCP-1 secretion by human pulmonary artery endothelial cells (HPAEC) and pulmonary microvascular endothelial cells (HMVEC-L). The most effective PS-ODN (MCP-1 AS 2) dose-dependently suppressed the secretion 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. Mismatch controls bearing 2 or 4 bp substitutions showed markedly reduced inhibitory 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 side effects on immediate-early gene regulation of the transcription factor nuclear factor-kappaB (NF-kappaB), as analyzed by gel shift assays. Antisense pretreatment of HPAEC reduced the monocyte chemotactic bioactivity liberated from tumor necrosis factor-alpha (TNF-alpha)-activated endothelial cells (EC) and reduced the TNF-alpha-induced transendothelial monocyte migration. We conclude that nanomolar concentrations of specific antisense oligodeoxynucleotides effectively inhibit human endothelial MCP-1 synthesis and may thus provide a rational approach to modulate monocyte recruitment under inflammatory conditions.