Secretory PLA2 inhibitor indoxam suppresses LDL modification and associated inflammatory responses in TNFα-stimulated human endothelial cells

Background and purpose: Secretory phospholipase A2 (sPLA2) is implicated in atherosclerosis, although the effects of specific sPLA2 inhibitors have not been studied. We investigated the effects of the indole analogue indoxam on low-density lipoprotein (LDL) modification by sPLA2 enzymes of different types and on the associated inflammatory responses in human umbilical vein endothelial cells (HUVEC). Experimental approach: LDL modification was assessed by measuring the contents of two major molecular species of lysophosphatidylcholine (LPC) using electrospray ionization-liquid chromatography/mass spectrometry. The proinflammatory activity of the modified LDL was evaluated by determining monocyte chemoattractant protein-1 (MCP-1) mRNA expression and transcriptional factor nuclear factor-kappaB (NF-κB) activity in HUVEC. Key results: Indoxam dose-dependently inhibited palmitoyl- and stearoyl-LPC production in LDL incubated with snake venom sPLA2 (IC50 1.2 μM for palmitoyl-LPC, 0.8 μM for stearoyl-LPC). MCP-1 mRNA expression and NF-κB activity were enhanced by venom sPLA2-treated LDL, which was completely suppressed by indoxam but not by thioetheramide-PC, a competitive sPLA2 inhibitor. Indoxam also suppressed LPC production in LDL treated with human synovial type IIA sPLA2. Tumour necrosis factor α (TNFα) increased type V sPLA2 expression in HUVEC. Indoxam dose-dependently suppressed LPC production in native and glycoxidized LDL treated with TNFα-stimulated HUVEC. Indoxam suppressed MCP-1 mRNA expression and NF-κB activity in TNFα-stimulated HUVEC incubated with native or glycoxidized LDL. Conclusions and implications: Indoxam prevented sPLA2-induced LPC production in native and glycoxidized LDL as well as LDL-induced inflammatory activity in HUVEC. Our results suggest that indoxam may be a potentially useful anti-atherogenic agent. British Journal of Pharmacology (2008) 153, 1399–1408; doi:10.1038/bjp.2008.12; published online 11 February 2008