Myeloperoxidase and serum amyloid A contribute to impaired in vivo reverse cholesterol transport during the acute phase response but not group IIA secretory phospholipase A(2)

Abstract

Atherosclerosis is linked to inflammation. HDL protects against atherosclerotic cardiovascular disease, mainly by mediating cholesterol efflux and reverse cholesterol transport (RCT). The present study aimed to test the impact of acute inflammation as well as selected acute phase proteins on RCT with a macrophage-to-feces in vivo RCT assay using intraperitoneal administration of [(3)H]cholesterol-labeled macrophage foam cells. In patients with acute sepsis, cholesterol efflux toward plasma and HDL were significantly decreased (P < 0.001). In mice, acute inflammation (75 microg/mouse lipopolysaccharide) decreased [(3)H]cholesterol appearance in plasma (P < 0.05) and tracer excretion into feces both within bile acids (-84%) and neutral sterols (-79%, each P < 0.001). In the absence of systemic inflammation, overexpression of serum amyloid A (SAA, adenovirus) reduced overall RCT (P < 0.05), whereas secretory phospholipase A(2) (sPLA(2), transgenic mice) had no effect. Myeloperoxidase injection reduced tracer appearance in plasma (P < 0.05) as well as RCT (-36%, P < 0.05). Hepatic expression of bile acid synthesis genes (P < 0.01) and transporters mediating biliary sterol excretion (P < 0.01) was decreased by inflammation. In conclusion, our data demonstrate that acute inflammation impairs cholesterol efflux in patients and macrophage-to-feces RCT in vivo in mice. Myeloperoxidase and SAA contribute to a certain extent to reduced RCT during inflammation but not sPLA(2). However, reduced bile acid formation and decreased biliary sterol excretion might represent major contributing factors to decreased RCT in inflammation.

Fig. 1.

Acute inflammation decreases macrophage cholesterol efflux in vitro toward plasma and HDL from sepsis patients. A: Efflux from THP-1 macrophages loaded with [³H]labeled cholesterol and acetylated LDL toward 1% plasma for 24 h. B: Efflux (24 h) from C57BL/6J thioglycollate-elicited peritoneal macrophages loaded with [³H]labeled cholesterol and acetylated LDL toward HDL (50 μg/ml) isolated by sequential ultracentrifugation. Experiments were performed as described under Materials and Methods. Data are given as means ± SEM. n = 10 for patients and n = 8 for controls. * indicates statistically significant differences from healthy controls (at least P < 0.05), and ^# indicates statistically significant differences from patients at day 21 (P < 0.001) as assessed by Student's t-test. Day 0, patients with acute sepsis at the day of admittance to the ICU; day 21, the same patients following full recovery from sepsis 21 days after admittance.

Fig. 2.

Acute inflammation impairs macrophage-to-feces reverse cholesterol transport in vivo in mice. C57BL/6J mice received intraperitoneal injections of either 75μg LPS/mouse or saline and [³H]cholesterol-loaded primary mouse macrophage foam cells as detailed in Materials and Methods and were followed for 48 h. A: FPLC cholesterol profiles of pooled plasma samples after injection of either saline (•) or LPS (▴). The relative elution position of the different lipoprotein subclasses is indicated. B: [³H]cholesterol appearance in plasma obtained 6, 24 and 48 h after macrophage administration. C: [³H]cholesterol tracer within liver 48 h after macrophage administration. D: [³H]tracer appearance in total feces collected continuously from 0 to 48 h after macrophage injection and separated into the bile acid and neutral sterol fractions as indicated. Data in B, C, and D are expressed as percentage of the injected tracer dose and are presented as means ± SEM. n = 5–6 for each group. * indicates statistically significant differences from saline controls (at least P < 0.05) as assessed by Student's t-test.

Fig. 3.

Mouse SAA overexpression in the absence of systemic inflammation decreases macrophage-to-feces reverse cholesterol transport in vivo in mice. On day 3 after injection with either the control adenovirus AdNull or the mouse CE/J SAA expressing adenovirus AdmSAA C57BL/6J mice received [³H]cholesterol-loaded primary mouse macrophage foam cells as detailed in Materials and Methods and were followed for 48 h. A: FPLC cholesterol profiles of pooled plasma samples after injection of either AdNull (•) or AdmSAA (▴). The relative elution position of the different lipoprotein subclasses is indicated. B: [³H]cholesterol appearance in plasma obtained 6, 24, and 48 h after macrophage administration. C: [³H]cholesterol tracer within liver 48 h after macrophage administration. D: [³H]tracer appearance in total feces collected continuously from 0 to 48 h after macrophage injection and separated into the bile acid and neutral sterol fractions as indicated. Data in C, D, and E are expressed as percentage of the injected tracer dose and are presented as means ± SEM. n = 8 for each group. * indicates statistically significant differences from AdNull injected controls (at least P < 0.05) as assessed by Student's t-test.

Fig. 4.

sPLA₂ overexpression in the absence of systemic inflammation does not affect macrophage-to-feces reverse cholesterol transport in vivo in mice. C57BL/6J mice and sPLA₂ transgenic mice received [³H]cholesterol-loaded primary mouse macrophage foam cells as detailed in Materials and Methods and were followed for 48 h. A: FPLC cholesterol profiles of pooled plasma samples in C57BL/6J mice (•) and sPLA₂ transgenic mice (▴). The relative elution position of the different lipoprotein subclasses is indicated. B: [³H]cholesterol appearance in plasma obtained 6, 24, and 48 h after macrophage administration. C: [³H]cholesterol tracer within liver 48 h after macrophage administration. D: [³H]tracer appearance in total feces collected continuously from 0 to 48 h after macrophage injection and separated into the bile acid and neutral sterol fractions as indicated. Data in B, C, and D are expressed as percentage of the injected tracer dose and are presented as means ± SEM. n = 5–6 for each group. * indicates statistically significant differences from wild-type controls (at least P < 0.05) as assessed by Student's t-test.

Fig. 5.

MPO impairs macrophage-to-feces reverse cholesterol transport in vivo in mice. C57BL/6J mice received [³H]cholesterol-loaded primary mouse macrophage foam cells and consecutive injections in 6 h intervals with either 50 µg/mouse purified MPO or vehicle for a total of 24 h as detailed in Materials and Methods. A: FPLC cholesterol profiles of pooled plasma samples after injection of either vehicle (•) or MPO (▴). The relative elution position of the different lipoprotein subclasses is indicated. B: [³H]cholesterol appearance in plasma obtained 6 and 24 h after macrophage administration. C: [³H]cholesterol tracer within liver 24 h after macrophage administration. D: [³H]tracer appearance in total feces collected continuously from 0 to 24 h after macrophage injection and separated into the bile acid and neutral sterol fractions as indicated. Data in B, C, and D are expressed as percentage of the injected tracer dose and are presented as means ± SEM. n = 6–7 for each group. * indicates statistically significant differences from vehicle controls (at least P < 0.05) as assessed by Student's t-test.