Figure 6.

High-density lipoprotein (HDL) and its associated enzyme paraox-onase-1 (PON1) detoxify danger signals on UCMVs. A and B, Unesterified cholesterol–induced microvescicles (UCMVs) were isolated from culture supernatants of human THP-1 monocytes and then incubated for 1 hour at 37°C without or with a subphysiological concentration of human HDL (100 μg HDL protein/mL, equivalent to 4 mg HDL cholesterol per 100 mL). Microvesicles were repurified from vehicle or HDL before our analyses. A, Representative histograms from flow cytometry of UCMVs stained with an IgMk isotype control antibody (blue curves), UCMVs stained with the EO14 antibody (filled purple curves), and HDL-treated UCMVs stained with the EO14 antibody (green curves). B, Wild–type murine aortic explants were incubated for 5 hours ex vivo with 0 (Control) or 3×10⁴ per μL of UCMVs or HDL-treated UCMVs. Explants were rinsed and then endothelial activation was assessed by the adherence of labeled monocytes. C and D, UCMVs were isolated from culture supernatants of human THP-1 monocytes and then incubated for 1 hour at 37°C without or with recombinant evolved PON1 (100 μg/mL). Microvesicles were repurified from vehicle or PON1 before our analyses. C, Representative histograms from flow cytometry of UCMVs stained with an IgMk isotype control antibody (blue curves), UCMVs stained with the EO14 antibody (filled purple curves), and PON1-treated UCMVs stained with the EO14 antibody (green curves). D, Wild–type murine aortic explants were incubated for 5 hours ex vivo with 0 (Control) or 3×10⁴ per μL of UCMVs or PON1-treated UCMVs. Explants were rinsed and then endothelial activation was assessed by the adherence of labeled monocytes. B and D, Mean±SEM, n=3–4. In each of these panels, P<0.001 by ANOVA, displayed are significance levels for individual pairwise comparisons by the Student-Newman-Keuls test.