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Purification and multi-omics characterization of human pre-beta-HDL reveals specific proteomic and lipidomic signature relevant to biological activities
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International audience. AIM: Multiple biological activities of high-density lipoprotein (HDL) particles may reflect their wide diversity. While alpha-migrating spherical HDLs are well characterised in terms of composition and function, small, dense, discoid pre-beta-migrating HDLs which are thought to display potent biological activities, remain difficult to purify and thus poorly characterised. Methods: We developed an original purification protocol yielding up to 2 mg of pre-beta-HDL (d 1.21-1.25 g/ml) from 70ml plasma of healthy volunteers (n=4). Alpha-HDL (d 1.08-1.11 g/ml) were purified concomitantly. Proteomic and lipidomic analyses of pre-beta- and alpha-HDL were performed and major biological activities, including cholesterol efflux and anti-inflammatory capacities, were evaluated in human THP-1 macrophages. Results: Pre-beta-HDL proteomic analysis identified 36 scaffold proteins gathered in three GO-biological processes. Proteins enriched in pre-beta-HDL were associated with inflammatory response, (e.g. PON1, serpin A1, haptoglobin) and immune response (e.g. IGHA1, C4A/B, haptoglobin-related-protein). Most of proteins related to lipid metabolism and transport were common to alpha- and pre-beta-HDL. Importantly, several functional proteins, including LCAT, PLTP, CETP and PON3, were exclusively detected in pre-beta-HDL. Lipidomic analysis revealed that pre-beta- and alpha-HDL contained similar lipid species. Yet, pre-beta-HDL was enriched in negatively-charged biologically-active phosphatidic acid (+35%; p<0.05) and phosphatidylserine (+20%; p<0.05), as compared to alpha-HDL. Cholesterol efflux from human macrophages to pre-beta-HDL was 3.5-fold elevated relative to alpha-HDL, being similar to that to lipid-free apolipoprotein A-I. Incubation with pre-beta-HDL prior to LPS stimulation reduced mRNA levels of IL-6 (-45%, p<0.05), IL-1-beta (- 44%, p<0.05) and TNF-alpha (- 60%, p<0.01) in THP-1 macrophages, while alpha-HDL exerted no effect.Conclusion: The present study, provides the first-ever structure-function characterization of human pre-beta-HDL using multi-omics analysis allowing identification of proteins and lipids potentially accounting for the enhanced biological activities of pre-beta-HDL. Our findings may provide novel clues to understand physiological and physiopathological roles of pre-beta-HDL.
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