Objectives: Membrane cholesterol is known to modulate a variety of cell signaling pathways and functions. While cholesterol depletion by High-Density-Lipoproteins (HDL) has potent anti-inflammatory... Show moreObjectives: Membrane cholesterol is known to modulate a variety of cell signaling pathways and functions. While cholesterol depletion by High-Density-Lipoproteins (HDL) has potent anti-inflammatory effects in various cell types, its effects on inflammatory responses in macrophages remain ill defined.Methods: Human and murine macrophages were pre-incubated with human reconstituted (apolipoproteinA-I/phosphatidylcholine) or native HDL.Results: HDL pre-incubation significantly decreased LPS-induced anti-inflammatory IL-10 production, while the opposite was observed for the pro-inflammatory mediators IL-12 and TNF. We show that these effects are mediated by passive cholesterol depletion and lipid raft disruption, without involvement of ABCA1, ABCG1, SR-BI or CD36. These pro-inflammatory effects are confirmed in vivoin peritoneal macrophages from ApoA-I transgenic mice, which have high circulating HDL levels. Native and reconstituted HDL enhances Toll-Like-Receptor-induced signaling by activating protein kinase C (PKC), since inhibition of PKC ablated the observed HDL effects. Using macrophages from NF-κB luciferase mice, we observed that HDL induces NF-κB activation. Western blot analyses showed that in particular the p65 subunit was activated. Using specific knock-out mice, we show that the observed HDL effects are independent of IKK, NIK and CKII. Furthermore, we observed that STAT1 is involved in the pro-inflammatory HDL effects on IL-10 and IL-12. On the other hand, we show that HDL enhances ADAM protease activity, thereby mediating TNF-α release.Conclusions: HDL exerts pro-inflammatory effects on macrophages via passive cholesterol depletion by activation of PKC-NF-kB/STAT1. These pro-inflammatory activities on macrophages could at least partly underlie the disappointing therapeutic potential of HDL raising therapy in current cardiovascular clinical trials. Show less
