Abstract
Human serum amyloid A (SAA) has been demonstrated as a chemoattractant and proinflammatory mediator of lethal systemic inflammatory diseases. In the circulation, it can be sequestered by a high-density lipoprotein, HDL, which carries cholesterol, triglycerides, phospholipids and apolipoproteins (Apo-AI). The capture of SAA by HDL results in the displacement of Apo-AI, and the consequent inhibition of SAA’s chemoattractant activities. It was previously unknown whether HDL similarly inhibits SAA-induced sPLA2 expression, as well as the resultant HMGB1 release, nitric oxide (NO) production and autophagy activation. Here we provided compelling evidence that human SAA effectively upregulated the expression and secretion of both sPLA2-IIE and sPLA2-V in murine macrophages, which were attenuated by HDL in a dose-dependent fashion. Similarly, HDL dose-dependently suppressed SAA-induced HMGB1 release, NO production, and autophagy activation. In both RAW 264.7 cells and primary macrophages, HDL inhibited SAA-induced secretion of several cytokines (e.g., IL-6) and chemokines (e.g., MCP-1 and RANTES) that were likely dependent on functional TLR4 signaling. Collectively, these findings suggest that HDL counter-regulates SAA-induced upregulation and secretion of sPLA2-IIE/V in addition to other TLR4-dependent cytokines and chemokines in macrophage cultures.
Highlights
Harboring various fatty acid side chains and phospholipid head groups [e.g., phosphatidylcholine (PC), phosphatidylserine (PS), or phosphatidyl ethanolamine (PE)], the heterogeneous phospholipids serve as the major components of cytoplasmic membranes and lipoproteinPLOS ONE | DOI:10.1371/journal.pone.0167468 November 29, 2016highdensity lipoproteins (HDL) Counter-Regulates SAA-Induced sPLA2 ExpressionAbbreviations: HDL, high-density lipoproteins; HMGB1, high mobility group box 1; LPS, lipopolysaccharide; NO, nitric oxide; sPLA2, secretory phospholipase A2; SAA, serum amyloid A; RAGE, receptor for advanced glycation end products; RT-PCR, reverse transcription polymerase chain reaction; TLR, toll-like receptors.particles
We recently demonstrated that SAA stimulates macrophages to release HMGB1 (29), it was previously unknown whether SAA upregulates sPLA2 secretion, an essential prerequisite for NO production [6] and HMGB1 release [7]
To assess whether SAA induces sPLA2 in innate immune cells, murine macrophage-like RAW 264.7 cells were stimulated with SAA at various concentrations (0.2, 1.0, 2.0 μg/ml) for 16 h, and the extracellular levels of sPLA2 in the macrophage-conditioned medium were determined by Western blotting using two different antibodies: rabbit polyclonal antibodies against a sPLA2-V peptide (Fig 1A), and rabbit monoclonal antibody against a peptide in the homologous C-terminus of sPLA2s (Fig 1B)
Summary
Harboring various fatty acid side chains and phospholipid head groups [e.g., phosphatidylcholine (PC), phosphatidylserine (PS), or phosphatidyl ethanolamine (PE)], the heterogeneous phospholipids serve as the major components of cytoplasmic membranes and lipoproteinPLOS ONE | DOI:10.1371/journal.pone.0167468 November 29, 2016HDL Counter-Regulates SAA-Induced sPLA2 Expressionparticles. The A2 group of phospholipases (PLA2s) hydrolyzes the fatty acid at the sn-2 position of the glycerol backbone of the phospholipids, releasing lysophospholipid as well as free fatty acids such as arachidonic acid (AA)–a substrate for other signaling lipids including prostaglandin E2 (PGE2), leukotrienes, and eicosanoids. Based on their molecular weight, cellular localization and dependence on calcium, PLA2s are further subdivided into: 1) Ca2+-dependent cytosolic enzymes (cPLA2s); 2) the low-molecular-weight and Ca2+-dependent secretory PLA2s (sPLA2); 3) Ca2+-independent enzymes (iPLA2s); 4) lipoprotein-associated PLA2 (Lp-PLA2 or sPLA2-VII); 5) lysosomal enzymes (LPLA2); and 6) adipose-specific enzymes (AdPLA2s) [1]. The mammalian sPLA2 family contains 10 catalytically active isoforms (IB, IIA, IIC, IID, IIE, IIF, III, V, X, and XIIA) [1], which predominantly hydrolyze phospholipids in the extracellular environment
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