Physiological effects of oxidized phospholipids and their cellular signaling mechanisms in inflammation

Free Radical Biology & Medicine 52 (2012) 266–280

Contents lists available at SciVerse ScienceDirect

Free Radical Biology & Medicine journal homepage: www.elsevier.com/locate/freeradbiomed

Review Article

Physiological effects of oxidized phospholipids and their cellular signaling mechanisms in inflammation Fiona H. Greig a, Simon Kennedy a, Corinne M. Spickett b,⁎ a b

Institute of Cardiovascular and Medical Sciences, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow, G12 8QQ, UK School of Life and Health Sciences, Aston University, Birmingham, B4 7ET, UK

a r t i c l e

i n f o

Article history: Received 1 September 2011 Revised 25 October 2011 Accepted 25 October 2011 Available online 31 October 2011 Keywords: Oxidized phospholipids Chlorinated lipids Apoptosis Leukocyte-endothelial adhesion Cell proliferation Anti-inflammatory effects Free radicals

a b s t r a c t Oxidized phospholipids, such as the products of the oxidation of 1-palmitoyl-2-arachidonoyl-sn-glycero-3phosphocholine by nonenzymatic radical attack, are known to be formed in a number of inflammatory diseases. Interest in the bioactivity and signaling functions of these compounds has increased enormously, with many studies using cultured immortalized and primary cells, tissues, and animals to understand their roles in disease pathology. Initially, oxidized phospholipids were viewed largely as culprits, in line with observations that they have proinflammatory effects, enhancing inflammatory cytokine production, cell adhesion and migration, proliferation, apoptosis, and necrosis, especially in vascular endothelial cells, macrophages, and smooth muscle cells. However, evidence has emerged that these compounds also have protective effects in some situations and cell types; a notable example is their ability to interfere with signaling by certain Toll-like receptors (TLRs) induced by microbial products that normally leads to inflammation. They also have protective effects via the stimulation of small GTPases and induce up-regulation of antioxidant enzymes and cytoskeletal rearrangements that improve endothelial barrier function. Oxidized phospholipids interact with several cellular receptors, including scavenger receptors, platelet-activating factor receptors, peroxisome proliferator-activated receptors, and TLRs. The various and sometimes contradictory effects that have been observed for oxidized phospholipids depend on their concentration, their specific structure, and the cell type investigated. Nevertheless, the underlying molecular mechanisms by which oxidized phospholipids exert their effects in various pathologies are similar. Although our understanding of the actions and mechanisms of these mediators has advanced substantially, many questions do remain about their precise interactions with components of cell signaling pathways. © 2011 Elsevier Inc. All rights reserved.

Contents Introduction . . . . . . . . . . . . . . . . . . . . . . . . . Sources and types of phospholipid oxidation products . . . Occurrence of oxidized phospholipids in disease . . . . . . Formation and occurrence of chlorinated lipids . . . . . . Proinflammatory physiological actions of oxidized and chlorinated Cell proliferation in atherosclerosis . . . . . . . . . . . .

. . . . . . . . . . . . lipids . . .

. . . .

. . . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

. . . . . .

267 267 268 269 269 270

Abbreviations: 2-ClHDA, 2-chlorohexadecanal; BAEC, bovine aortic endothelial cell; CAD, coronary artery disease; CD36, cluster differentiation 36; CL, cardiolipin; COX-2, cyclooxygenase-2, ECM, extracellular matrix; eNOS, endothelial nitric oxide synthase; ER, endoplasmic reticulum; ERK1/2, extracellular-signal-regulated kinase 1/2; GPCR, G-protein-coupled receptor; HAEC, human aortic endothelial cell; HCAEC, human coronary artery endothelial cell; HDL, high-density lipoprotein; HOOA-PC, 1-palmitoyl-2-(5-hydroxy-8-oxo-oct-6-enoyl)-sn-glycero-3-phosphocholine; HUVEC, human umbilical cord endothelial cell; ICAM-1, intracellular adhesion molecule-1; IL, interleukin; LDL, low-density lipoprotein; LOX-1, lectin-like endothelial receptor for oxLDL; LPS, lipopolysaccharide; MCP-1, monocyte chemotactic protein-1; MKP-1, mitogen-activated protein kinase phosphatase-1; mmLDL, minimally modified LDL; MPO, myeloperoxidase; NOX4, NADPH oxidase 4; ONOO−, peroxynitrite; oxLDL, oxidized LDL; oxPAPC, oxidized 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine; PAF, platelet-activating factor; PAMP, pathogen-associated molecular pattern; PAzPC, 1-palmitoyl-2-azelaoyl-sn-glycero-3-phosphocholine; PC, phosphatidylcholine; PE, phosphatidylethanolamine; PECPC, 1-palmitoyl-2-epoxycyclopenteneone-sn-glycero-3-phosphocholine; PEIPC, 1-palmitoyl-2-(5,6-epoxyisoprostane E2)-sn-glycero-3-phosphocholine; PGPC, 1palmitoyl-2-glutaryl-sn-glycero-3-phosphocholine; PI3K, phosphatidylinositol 3-kinase; PKC, protein kinase C; PONPC, 1-palmitoyl-2-(9′-oxo-nonanoyl)-sn-glycero-3-phosphocholine; POVPC, 1-palmitoyl-2-(5′-oxo)valeryl-sn-3-glycerophosphocholine; PPAR, peroxisome proliferator-activated receptor; PS, phosphatidylserine; SAzPC, 1-stearoyl-2-azelaoyl-sn-glycero3-phosphocholine; SGPC, 1-stearoyl-2-glutaroyl-sn-glycero-3-phosphocholine; SM, sphingomyelin; SMase, sphingomyelinase; SOPC, 1-stearoyl-2-oleoyl-sn-glycero-3-phosphocholine; SOVPC, 1-stearoyl-2-(5′-oxo)valeryl-sn-3-glycerophosphocholine; SRA-I/II, scavenger receptor class A types I and II; SREBP, sterol-regulatory element-binding protein; TLR, Toll-like receptor; UVA-1, ultraviolet radiation; VSMC, vascular smooth muscle cell. ⁎ Corresponding author. Fax: + 44 121 2044175. E-mail address: [email protected] (C.M. Spickett). 0891-5849/$ – see front matter © 2011 Elsevier Inc. All rights reserved. doi:10.1016/j.freeradbiomed.2011.10.481

F.H. Greig et al. / Free Radical Biology & Medicine 52 (2012) 266–280

Apoptotic signaling . . . . . . . . . . . . . . . . . . . . Endothelial function . . . . . . . . . . . . . . . . . . . . Leukocyte and monocyte binding . . . . . . . . . . . . . . Receptors for oxidatively modified lipids . . . . . . . . . . . . . Platelet-activating factor receptor . . . . . . . . . . . . . . Scavenger receptors . . . . . . . . . . . . . . . . . . . . Peroxisome proliferator-activated receptors . . . . . . . . . Toll-like receptors . . . . . . . . . . . . . . . . . . . . . Anti-inflammatory and beneficial effects of oxidized phospholipids Summary: advances and limitations . . . . . . . . . . . . . . . Acknowledgments . . . . . . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

Introduction Inflammation is an adaptive multistep response by which the host reacts to a change in homeostasis, usually triggered by a noxious stimulus such as infection or alterations in physiological conditions [1]. The inflammatory response can be divided into two forms of attack to remove the damaged tissue and rectify the homeostatic imbalance: acute and chronic reactions. Acute inflammation usually lasts for hours or days, whereas chronic inflammatory responses persist for a much longer period of time. Many inflammatory diseases involve chronic inflammation, in which there is an ongoing stimulation of immune cells without normal resolution. In these situations oxidative damage to biological molecules often occurs, as activated phagocytes generate a variety of reactive oxygen, nitrogen, and chlorine species, some of which are free radicals. Phospholipids, in particular those containing polyunsaturated fatty acids, are susceptible to peroxidation by free radicals, and monounsaturated fatty acyl groups may also be oxidized by nonradical species such as peroxynitrite (ONOO−) and hypochlorous acid (HOCl). There is now increasing evidence that lipid peroxidation occurs and oxidized phospholipids are formed in inflammatory diseases such as atherosclerosis, neurodegenerative diseases including Alzheimer's disease, Parkinson's disease, and multiple sclerosis, rheumatoid arthritis, diabetes, and systemic lupus erythematosus [2]. Some of the effects of oxidized phospholipids in inflammatory

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

267

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

. . . . . . . . . . . .

271 271 272 273 273 273 274 274 275 276 276 276

diseases are summarized in Fig. 1. As oxidized phospholipids have been found to induce a broad range of proinflammatory effects, there is considerable interest in elucidating their contribution to the pathology of these conditions and determining the molecular signaling mechanisms that underlie the effects [3]. There have been many reviews on the role of oxLDL and oxidized phospholipids in atherosclerosis, but fewer have addressed their involvement in other inflammatory diseases and more generally in inflammatory processes. This review aims to provide a summary of the current knowledge in this broader area, with a focus on the physiological actions and signaling of both oxidized and chlorinated phospholipids. Sources and types of phospholipid oxidation products Phospholipids are major components of cellular membranes and also of circulating plasma lipoproteins; both sites can give rise to oxidized phospholipids after oxidative stress and reactive oxidizing species production. In both cases, phosphatidylcholines (PCs) are the most abundant species, with phosphatidylethanolamines (PEs) and sphingomyelins (SMs) also contributing a substantial proportion; phosphatidylserines (PSs) are less abundant components found normally in the inner leaflet of the plasma membrane. Phosphatidylinositols and cardiolipins (CLs; diphosphatidylglycerols) are cellular rather than circulating lipoprotein components, with cardiolipins located mainly in the

Fig. 1. Summary of the physiological effects of oxidized phospholipids in inflammatory diseases.

268

F.H. Greig et al. / Free Radical Biology & Medicine 52 (2012) 266–280

mitochondrial membrane. It has been shown that apoptotic cells and bodies contain increased levels of phospholipid oxidation products, reflecting the occurrence of oxidative damage during apoptotic events [4]. With regard to lipoproteins, there has been most interest in the formation of oxidized phospholipids in low-density lipoprotein (LDL) in cardiovascular disease and atherosclerosis, although it is now emerging that high-density lipoprotein (HDL) is also susceptible to oxidative damage, which decreases its antiatherogenic properties. Increased levels of oxidized lipoproteins have also been found in other diseases, such as diabetes and Alzheimer disease [2,3]. Although the head groups of some phospholipids, especially PEs, can be oxidized, the term “phospholipid oxidation” usually refers to oxidation of the unsaturated fatty acyl chains. Consequently, analogous oxidation products can arise from several different phospholipid classes. The initial process of lipid peroxidation (a nonenzymatic, radical-catalyzed process) generates phospholipid peroxyl radicals that undergo conversion to phospholipid hydroperoxides and endoperoxides and subsequently further rearrangement to full-chain-length species such as esterified isoprostane-like compounds. Alternatively, during this process the oxidized fatty acyl chain may fragment to yield chain-shortened products with various combinations of hydroxyl-, carbonyl-, or carboxylic acid moieties, often with α,β-double bonds (e.g., hydroxyalkenals) [3,5,6]. The extent of this family of phospholipid oxidation products presents a challenge for understanding the full effects of oxidized phospholipids generated in biological situations. In addition to the nonenzymatic mechanism mentioned above, phospholipid hydroperoxides can be formed by the action of lipoxygenases and cyclooxygenases on both esterified polyunsaturated fatty acids (PUFAs) and free PUFAs released from the cell membrane by the action of phospholipases. In this case, they are usually metabolized further to more stable compounds such as prostaglandins and thromboxanes. Alternatively, they can further form highly reactive lipid peroxyl radicals by nonenzymatic reactions catalyzed by free transition metal ions. Phospholipid hydroperoxides can be reduced and detoxified by a phospholipid-dependent glutathione peroxidase, which converts them to the corresponding alcohol [7]. An extensive discussion of lipid peroxidation reactions is outside the scope of this review but they have been extensively reviewed recently [8]. Although phospholipid hydroperoxides have been found in a number of disease conditions, as discussed in the following section, and may have some important physiological actions, it is very difficult to study their effects owing to their relative instability in cellular or biological situations. In practice, many studies on the effects of oxidized phospholipids (in particular those relating to atherosclerosis) have been carried out using LDL oxidized in vitro, which represents a very complex mixture of lipid oxidation products including oxysterols, oxidized cholesterol esters, oxidized phospholipids, modified plasmalogens, and lysophospholipids. Although it is quite likely that LDL and other lipoproteins may be the source of oxidized phospholipids and contribute to pathology in a variety of diseases, it is important to bear in mind that LDL is a complex molecule containing apolipoprotein B-100 and antioxidants as well as a selection of lipids. Correct interpretation of studies to investigate the effects of oxidized LDL (oxLDL) therefore requires knowledge of the extent to which each of the LDL components is oxidized; otherwise effects may be attributed to oxidized phospholipids that are in reality due to oxysterols or protein modification. Oxidized 1-palmitoyl-2arachidonoyl-sn-glycero-3-phosphocholine (oxPAPC) has also been used as a model system, after the demonstration that it mimics some of the biological effects of oxLDL [9]; this has been characterized as a complex mixture of lipids containing both full-chain and chainshortened products as well as the nonesterified products resulting from oxidized chain cleavage [3,9,10]. A smaller number of studies have investigated the effects of individual oxidized phospholipids, such as the truncated products 1-palmitoyl-2-(5′-oxo)valeryl-sn-3glycerophosphocholine (POVPC), 1-palmitoyl-2-glutaryl-sn-glycero-3-

phosphocholine (PGPC), and 1-palmitoyl-2-(9′-oxo-nonanoyl)-snglycero-3-phosphocholine (PONPC) and the full-length phospholipids 1-palmitoyl-2-(5,6-epoxyisoprostane E2)-sn-glycero-3-phosphocholine (PEIPC) and 1-palmitoyl-2-epoxycyclopenteneone-sn-glycero-3phosphocholine (PECPC). Occurrence of oxidized phospholipids in disease There is steadily increasing evidence that phospholipid oxidation products are formed in vivo and are present in a variety of diseased tissues. There are many studies in which levels of lipid or phospholipid hydroperoxides have been measured using nonspecific methods for generic peroxides and have been found to increase in diseases. For example, in a study of 38 human atherosclerotic plaque samples, a correlation was found between low-molecular-weight iron, lipid hydroperoxides, and lipid peroxidation products, whereas high plaque iron correlated with plaque severity [11]. Recent evidence from mice suggests that the exaggerated inflammatory response in cystic fibrosis may be due to impaired detoxification of phospholipid hydroperoxides [12]. In patients with rheumatoid arthritis, lipid hydroperoxides were also found to be raised, coupled with a reduced activity of several endogenous antioxidants, including paraoxonase 1 [13]. Because oxidized lipids seem to be substrates for the paraoxonase family [14], this may explain why patients with rheumatoid arthritis are predisposed to other diseases such as atherosclerosis [15]. In terms of identifying specific oxidized phospholipids, most attention has been focused on atherosclerosis, as this disease involves both dyslipidemia and chronic inflammation, and the oxidative modification of LDL is well characterized as a primary risk factor in its initiation and progression [16]. Lipoproteins become trapped in the subendothelial layer of the artery wall and after oxidative attack form oxLDL, which is crucial in the formation of foam cells, release of proinflammatory cytokines, and phenotypic changes in the resident cells [17,18]. This is the basis of the response-to-injury hypothesis, in which the inflammation begins as a protective measure and then because of the continued response can become detrimental to the vessel [16]. There is good evidence that various oxidized phospholipids are found in the atherosclerotic plaque. One of the earliest reports was of the presence of PEIPC, POVPC, and PGPC in the aorta of rabbits fed on a high-fat diet [9], whereas hydroxyalkenals such as 5-hydoxy-8-oxo-6-octenoic acid esterified to PC (HOOA-PC) were reported by Hoff et al. [19]. Ravandi and colleagues [20] reported the presence of hydroperoxides, hydroxides, epoxides, isoprostanes, and core aldehydes of the major phospholipids in human atherosclerotic lesions. Fragmented and full-length oxidized phospholipid species have also been observed in human LDL by electrospray mass spectrometry [21]. Lung diseases such as adult respiratory distress syndrome and acute lung injury are now understood to result in oxidized phospholipid formation, often from the surfactant phospholipids present in the lung lining fluid [22,23]. These conditions are related to atherosclerosis in terms of the loss of endothelial cell barrier function, which allows infiltration of inflammatory cells into the interstitial fluid. Early work showed that hydroxyeicosatetraenoic acids and epoxyeicosatetraenoic acids esterified to phospholipids were present in normal murine lung tissue [24], whereas ischemia in rat lungs resulted in the formation of lipid hydroperoxides [25]. More recently, PS and CL oxidation products [26] and isofurans [27] have been characterized in acute lung injury caused by hyperoxia in mice, whereas in humans oxidized PC has been detected using an antibody in lung tissues of patients with idiopathic interstitial pneumonias [28]. In fact, infections of cells or tissues have been found to result in the formation of oxidized phospholipids, such as mycobacteria-infected macrophages showing increased concentrations of PEIPC [29] and alveolar cells infected with influenza A virus generating POVPC, PEIPC, and PGPC [30]. Although oxidized phospholipid formation in other diseases has been less studied, there

F.H. Greig et al. / Free Radical Biology & Medicine 52 (2012) 266–280

have been reports that inflammatory brain diseases are also associated with increased levels of phospholipid oxidation products [31–34], and isolevuglandins (γ-ketoaldehydes) and isofurans have been found in neurodegenerative conditions such as Parkinson's disease and Alzheimer's disease [35,36]. Evidence for the formation of these species in another neurodegenerative disease, glaucoma, comes from the detection of isolevuglandin-modified proteins [37]. Lipid peroxidation has been associated with the pathology of rheumatoid arthritis [38,39], although this may be due to a high incidence of atherosclerosis in these patients [40]. Oxidation products of PAPC have been found in ultraviolet radiation (UVA-1)-treated human keratinocytes and fibroblasts as well as in murine skin subjected to UVA-1 damage [41]. The occurrence of lipid peroxidation and oxidized phospholipids in many inflammatory and immune-based diseases is known [2,42,43], although the precise identity of the products has not been determined in all cases, and often relatively nonspecific methods such as TBARS assays, HPLC of lipid hydroperoxides, or antibody-based methods have been used. The levels of oxidized phospholipids in vivo have proved difficult to quantify, owing to the large number of different structures of oxidatively modified phospholipids that are known to exist and the limited availability of suitable standards. Several studies in humans have been carried out using antibodies raised against oxidized phospholipids, especially one that is reported to be specific for POVPC in lipoproteins, and the levels are usually quoted as a ratio versus lipoprotein (a) (Lp(a)) [44]. Although potentially a good marker, this provides a limited view of the oxidized phospholipids present, so mass spectrometry coupled to HPLC has become the principal method of detection for both human and animal samples because it is able to identify a wide range of oxidized species in a single run. An additional problem encountered when trying to compare concentrations between studies is that the levels of oxidized phospholipids analyzed are quoted in a variety of ways, such as nanograms per milligram of tissue or 10 6 cells, picomoles per nanomole of unmodified or total phospholipid, or in micromolar. In studies of human atherosclerotic plaque material the level of POVPC was 20–40 μg/g tissue, with lower levels (1–8 μg/g tissue) of other aldehydes such as SOVPC, PAzPC, and SAzPC [20]. This is similar to the values reported for rabbit aorta, which range from about 40 to 100 μg/g tissue for POVPC, PGPC, and PEIPC [9,45]. Assuming that 1 g of tissue corresponds to a volume of approximately 1 ml, 1 μg/g tissue would correspond to a concentration of 1.7 μM for POVPC (MW 594) and 1.2 μM for PEIPC (MW 828); thus it can be seen that in atherosclerotic plaque material, concentrations of 1–50 μM or even higher might exist. In contrast, lower values have been reported in plasma samples from LDL receptor knockout (Ldlr−/−) mice on a Western diet and normotriglyceridemic human volunteers [46]. In Ldlr−/− mice the concentrations ranged from 0.5 to 3 μM for a variety of chain-shortened oxidized phospholipids, with the highest values found for the 5-keto-6-octendioic acid ester of 2lysoPC. In human samples the individual oxidized phospholipids could not be quantified, but total oxidized phospholipids were estimated in the range 0.7 to 5 μM. A recent review by Bochkov et al. summarized the levels of oxidized phospholipids in vivo in several species, including human and rodent, under various disease conditions [3]. Thus under pathological conditions in any one tissue, there is evidence that several different species of oxidized phospholipid occur. The variety of fragmentation sites and locations of oxidation, together with the observation of adducts with proteins for several species, argues for their formation in vivo rather than being artifacts of extraction. Although the concentration range is broad and further studies in vivo are needed, it seems clear that oxidized phospholipids are present in the low-micromolar range, and this is the concentration range that has been tested in most cell culture or tissue experiments to investigate their effects in vitro. Many studies have now been carried out using individual isolated or chemically synthesized oxidized phospholipid species, which is important to enable understanding of the precise

269

biological effects of each compound, in contrast to studies of oxidized lipoproteins, in which it is not known which oxidized component is contributing. Formation and occurrence of chlorinated lipids A further level of complexity in the range of oxidized products arises from the fact that the phagocyte enzyme myeloperoxidase (MPO) is able to chlorinate and nitrate biological molecules. MPO produces HOCl and is the only known route for producing chlorinated species under physiological conditions in vivo [47,48]. HOCl is added across the unsaturated bonds of phospholipids by electrophilic attack, forming lipid chlorohydrins [49,50]. Treatment of unsaturated PCs containing one or two double bonds with the MPO/hydrogen peroxide/chloride system predominantly forms chlorohydrins in, for example LDL [51], whereas phospholipids with a higher level of unsaturation (e.g., those with arachidonate chains) were liable to fragment to form lysoPCs [52]. Another type of chlorinated product, formed by the chlorination of plasmalogens by MPO, are α-chloro fatty aldehydes. These are formed when the vinyl ether bonds of PE or PC plasmalogens are targeted by HOCl [53,54]. MPO has been reported to be influential in a number of disease states and in particular, inflammatory disorders. Elevated levels of plasma MPO have been shown in rheumatoid arthritis as well as multiple sclerosis [55,56]. Furthermore, a murine model of inflammatory bladder disease was found to have a 61-fold increase in MPO after 24 h [57] and increased plasma MPO levels were also demonstrated in individuals with liver cirrhosis and chronic hepatitis B [58]. In addition, these effects have been documented in the cardiovascular system, in which patients with unstable coronary artery disease (CAD) were found to have increased levels of MPO in comparison with stable CAD and control patients [59,60]. Furthermore, the active form of the phagocytic enzyme is expressed within human atherosclerotic lesions [61]. This has led to the suggestion of MPO levels being a potentially useful prognostic tool for predicting major cardiac events [60,62]. Thus the probability of some chlorinated products being formed in these conditions is high. 2-Chlorohexadecanal (2-ClHDA) has been reported to be increased more than 1400-fold in atherosclerotic tissues, as analyzed by gas chromatography–mass spectrometry [63]. Increased levels of this α-chloro fatty aldehyde were also demonstrated in the hearts of rats suffering myocardial infarction as a result of coronary artery occlusion and correlated with increased expression of MPO and neutrophil infiltration [64]. More recently, increases in MPO protein level and 2-ClHDA have been observed in mouse brains after administration of lipopolysaccharide (LPS) in vivo [65]. The presence of lysoPC chlorohydrin in human atherosclerotic lesions has been reported by Messner et al., with 69- and 82fold increases in 16:0 and 18:0 lysoPC chlorohydrins, respectively, compared to normal aorta [66]. Although the number of reports on chlorinated lipids and phospholipids in vivo is limited, increasing evidence is emerging that they are formed and play a role in pathology. Proinflammatory physiological actions of oxidized and chlorinated lipids The modification of phospholipids has been shown to be crucial in the progression of some inflammatory diseases, including atherosclerosis [2,3,6], and oxidized phospholipids have been shown to influence the behavior of various cell types, as summarized in Fig. 2. Inflammatory diseases often have common underlying pathophysiology, especially relating to interactions of immune cells with the endothelium of various parts of the vasculature. Vascular smooth muscle cell (VSMC) proliferation, leukocyte migration, apoptosis, and formation of extracellular matrix (ECM) are important features of the remodeling within the vasculature that occurs in atherosclerosis [67]. Endothelial dysfunction and leukocyte migration are also features of acute lung inflammation

270

F.H. Greig et al. / Free Radical Biology & Medicine 52 (2012) 266–280

Fig. 2. Schematic diagram showing the effects of oxidized phospholipids on various cell types. Responses regarded as potentially deleterious are shown in red and potentially beneficial ones are in blue. The flat-ended blocks indicate inhibition of inflammatory responses. DC, dendritic cell; EC, endothelial cell; ϕ, macrophage.

[68]. The biological effects of lipids with regard to proliferation, leukocyte endothelial adhesion, and apoptosis have been broadly researched, whereas there is less evidence of the effects on SMC migration and ECM formation. VSMCs are key producers of ECM within the vessel wall and therefore any effect of the modified lipids on these cells will lead to potential changes in the structure of the artery [69]. Oxidized lipids have been shown to be involved in the initial stages of atherosclerosis in which POVPC and PGPC activate phenotypic switching of VSMCs from the contractile to the synthetic state, both in vitro and in vivo [70]. The oxidized products of PAPC were found to suppress smooth muscle α-actin and induce the nuclear translocation of a known smooth muscle repressor gene, Krüppel-like transcription factor 4 [70]. POVPC and PGPC can activate VSMC migration and type VIII collagen expression, involved in the composition change of ECM in atherosclerotic lesions [38,71]. In the study by Al-Shawaf et al. [38], the migration of VSMCs was found to involve stimulation of the cationic channel TRPC5, resulting in Ca2 + entry, by a mechanism involving Gi/o G-protein-coupled receptors (GPCRs) that had not previously been associated with oxidized phospholipid responses. OxLDL has also been implicated in the switching of M2 macrophages from an anti-inflammatory to a proinflammatory state and the enhanced secretion of monocyte chemotactic protein-1 (MCP-1), leading to the formation of more foam cells [72]. T-cells are also affected by oxLDL and show increased production of receptor activator of NF κB ligand (RANKL) and activation of NF-κB. In addition, RANKL triggers maturation and activation of osteoclasts and therefore contributes to bone resorption, which occurs in osteoporosis, periodontal disease, and rheumatoid arthritis [73]. Cell proliferation in atherosclerosis Proliferation of VSMCs within vessels is essential for the development of atherosclerosis, as it is characterized by a thickening of the artery wall. The effects of modified lipids have previously been found to induce a biphasic effect on VSMCs, whereby at low concentrations, they exert a proliferative action and at higher concentrations, apoptosis predominates [74]. The length of exposure is also an important factor; longer incubations with oxidized lipids induce apoptosis as described below, and therefore a critical balance exists between the effects of modified lipids on VSMCs [75]. This balance is an important aspect of

atherosclerosis and also the process of restenosis, a common side effect seen after treatment for atherosclerosis. Restenosis is characterized by the movement and proliferation of VSMCs causing the luminal area of the artery to narrow and the angina-like symptoms to reappear, and modified phospholipids could play a critical role in this process. The biologically active component of minimally modified LDL (mmLDL) stimulating VSMC proliferation was POVPC, whereas PGPC was shown to have no effect [76]. POVPC triggered the production of lactosylceramide, which led to the generation of superoxide radicals (O2•−) and caused phosphorylation of p44 mitogen-activated protein kinase (MAPK) promoting VSMC proliferation [76]. POVPC and PGPC are thought to work through separate pathways to induce VSMC proliferation. A study that measured 5-ethynyl-2′-deoxyuridine incorporation after POVPC and PGPC were topically applied to mouse carotid arteries in vivo found POVPC-treated arteries to have significant increases in proliferation, but this effect was not witnessed in arteries incubated with PGPC [77]. This could be due to the different structures of the truncated oxidized products; POVPC is an aldehyde and is able to form covalent adducts with proteins in the plasma membrane, whereas PGPC is a carboxylic acid and typically internalized within lysosomes of cells [78]. An inhibitory action of the oxidized lipids has also been reported, whereby VSMC growth was significantly reduced after treatment with PGPC and to a greater extent with POVPC [79]. This suggests that modified phospholipids are not only involved in the production of foam cells but also in the change in the vessel architecture leading to advanced plaque formation in atherosclerosis. In addition to the effects of defined oxidized phospholipids, a number of studies have looked at the effects of oxLDL on cell proliferation within inflammatory diseases. OxLDL stimulates VSMC and fibroblast proliferation through an interaction with the scavenger receptor causing activation of the phosphatidylinositol 3-kinase (PI3K) and phospholipase C pathways, one of its targets being the activation of endothelial nitric oxide synthase (eNOS) [80]. Activation of the PI3K/Akt pathway was also reported in oxLDL-induced VSMC proliferation involving the SM/ceramide pathway leading to the activation of extracellularsignal-regulated kinase 1/2 (ERK1/2) [74]. Furthermore, oxLDL at a low concentration of 5 μg/ml caused an increase in human coronary artery endothelial cell (HCAEC) proliferation [81]. The E-cadherin/βcatenin/T-cell factor pathway is activated after induction of VSMC

F.H. Greig et al. / Free Radical Biology & Medicine 52 (2012) 266–280

proliferation by oxLDL and is influential in cell adhesion and also the inflammatory response [82]. Macrophages are another important cell type in atherosclerosis and their proliferation can also be induced by oxLDL. Treatment with oxLDL has been shown to stimulate macrophage proliferation by the activation of ERK1/2 and MAPKs by measuring the incorporation of [3H]thymidine and cell counts [83]. Oxidized phospholipids, in general, induce proliferation of cells, notably VSMCs and macrophages, at low concentrations and lead to the activation of several pathways involved in the immune response. Apoptotic signaling Cell death plays a critical role in development, homeostasis, and cell turnover by removing damaged or infected cells from tissues or the circulation by an intrinsic mechanism of cell suicide [84]. Apoptosis is characterized by programmed cell death involving cell shrinkage and chromatin condensation with subsequent fragmentation of DNA. Defective or inappropriate apoptosis plays a part in many inflammatory disease pathologies and is known to be an important aspect of remodeling in the vasculature and disease progression in atherosclerosis [85]. Modified lipids have been found to induce apoptotic signaling pathways in several cell types involved in atherosclerotic lesions such as VSMCs and macrophages. PGPC and POVPC, the truncated products from the oxidation of PAPC, have been found to activate apoptosis in VSMCs as their principal method of cell death [79]. Both of the phospholipids induced phosphorylation of MAPKs and activated sphingomyelinases (SMase's), in particular the acid isoform that is known to be involved in the initial stages of apoptosis [86]. Ceramide is a hydrolysis product of SM and mimics the action of mmLDL, inducing apoptosis through the caspase-3 signaling pathway [87]. Evidence of the importance of such effects in vivo comes from the observation that in Ldlr−/− mice crossed with programmed cell death-1 receptor knockout (Pd1−/−) mice, there was a large increase in atherosclerotic lesion size and a change in composition, with the plaques containing more macrophages and T-cells compared with the Ldlr−/− control mice [88]. In rat oligodendrocytes, POVPC was found to activate caspase-3 and -8, but in this cell type neutral SMase, not the acid isoform, was responsible for the increase in ceramide [89]. These results support the concept that oxidized phospholipids would be deleterious in the brain by inducing apoptosis and facilitating neurodegenerative diseases. Apoptosis in VSMCs and macrophages can promote inflammation, accelerate atherosclerosis, and alter the composition of the atherosclerotic lesions [90,91]. Human VSMCs are competent phagocytes and capable of engulfing other VSMCs that are undergoing apoptosis; however, this process is significantly reduced by oxLDL or hyperlipidemia in vivo [92]. The relevance of this observation is emphasized by the fact that high levels of biologically active oxidized lipids are found in apoptotic cells [93]. These VSMCs can then release interleukin-1α (IL-1α), and secondary apoptotic VSMCs releasing both IL-1α and IL-1β promote the inflammatory response by causing surrounding VSMCs to produce proinflammatory cytokines [92]. However, oxLDL is known to contain lysoPC, which has been reported to trigger significant apoptosis in VSMCs, as assessed by DNA fragmentation [94]. Both endothelial cells and macrophages are also sensitive to the apoptotic effects of oxidized phospholipids. Macrophages were shown to activate apoptotic signaling pathways after incubation with LDL oxidized by copper sulfate, measured using a variety of viability assays including TUNEL [95]. In contrast, oxLDL has also been implicated in the survival of macrophages, by a mechanism involving the inhibition of acid SMase activity and a decrease in ceramide [96]. The authors suggested that the survival–apoptosis balance is dose-dependent, and in other studies in which cytotoxicity occurred, higher concentrations were utilized. In addition, IL-10, a key anti-inflammatory cytokine, was found to block macrophage apoptosis induced by oxLDL [97]. Cell death after incubation with modified lipids has also been reported in endothelial cells. OxLDL up-regulated expression of the

271

lectin-like endothelial receptor for oxLDL (LOX-1), measured using mRNA and protein levels, and incubation of oxLDL with HCAECs induced apoptosis in a concentration- and time-dependent manner [98]. HCAECs treated with oxLDL activated the NF-κB pathway, and an inhibitor of the pathway, caffeic acid phenethyl ester, consequently reduced the oxLDL-mediated apoptosis [98]. Moreover, mmLDL produced by reaction with 15-lipoxygenase was found to increase the expression of LOX-1 receptor in endothelial cells, and in an endothelial cell line in which LOX-1 was overexpressed, intracellular adhesion molecule-1 (ICAM-1) was significantly up-regulated [99]. Similar to VSMCs, oxLDL activates caspase signaling pathways, and in particular caspase-9 and -3 in HCAECs, as well as release of cytochrome c, all leading to apoptosis [100]. The possible mechanisms of action of oxPAPC, POVPC, and PGPC on endothelial cells and VSMCs in atherosclerosis are shown in Fig. 3. Chlorohydrins can be produced from the action of HOCl on phospholipids and lysoPCs. Chlorinated lipids produced by the modification by HOCl on sphingomyelin have been found to activate apoptotic signaling in dopaminergic PC12 neurons by the activation of caspase-3 [101]. Phospholipid chlorohydrins were further found to deplete ATP levels in human myeloid cells and cause a loss of viability [102]. Similarly, phospholipid chlorohydrins depleted ATP levels in U973 cells and activated the caspase-3 signaling cascade [103]. This toxic effect of chlorohydrins demonstrated in myeloid cells was mimicked in endothelial cells in which both chlorohydrins and bromohydrins were cytotoxic in the micromolar range; however, necrosis appeared to be the predominant form of cell death [104]. The modification of phospholipids in general leads to the loss of viability in several cell types in a concentration- and time-dependent manner. Overall, this could contribute to the inflammatory responses in any inflammatory diseases in which neutrophils and MPO are up-regulated. Endothelial function Endothelial dysfunction is a factor in acute inflammatory lung diseases, such as adult respiratory distress syndrome, and is believed to be an early event in the process of atherosclerosis. Clearly, the induction of apoptosis by oxidatively modified phospholipids, as described above, is one mechanism leading to endothelial dysfunction [98,100], but some less severe effects can also contribute to disease. Endothelial cells treated with oxPAPC activated more than 1000 genes and produced a range of inflammatory chemokines including MCP-1, IL-6, and IL-8, involved in both cell adhesion and migration [105]. Oxidatively modified phospholipids have also been found to affect nitric oxide (NO) biosynthesis, which is crucial in the regulation of vascular arterial tone and could occur either through a decrease in NO production or through a decrease in biological activity of NO once produced. In bovine aortic endothelial cells (BAECs), oxLDL rapidly reduced the levels of intracellular NO and induced the production of O2•−; however, oxLDL did not affect the ability of eNOS to produce NO [106]. NO and O2•− can rapidly combine to form ONOO−, which is thought to be the route by which NO has most of its cytotoxic effects [107]. The increase in reactive oxygen species produced by oxPAPC is thought to be controlled by NADPH oxidase 4 (NOX4) indirectly, causing the recruitment of rac1 to the membrane and in turn activating NOX4, enhancing the inflammatory response [108]. Conversely, in human aortic endothelial cells (HAECs), treatment with oxPAPC caused a time-dependent activation of eNOS, which was regulated by the PI3K/Akt pathway [109]. The same group previously found oxPAPC to induce the formation of IL-8, a proinflammatory cytokine, in endothelial cells, and the effect was sustained for up to 24 h [110]. Treatment with N-nitro-L-arginine methyl ester, an NOS inhibitor, reduced the oxPAPC-induced IL-8 transcription [109]. Similarly, incubation with oxLDL caused an increase in intracellular NO in HCAECs, as well as an increase in phosphorylation of PI3K, Akt, and eNOS [81]. OxPAPC activated sterol-regulatory element-binding proteins (SREBP), which are transcription factors involved in cholesterol

272

F.H. Greig et al. / Free Radical Biology & Medicine 52 (2012) 266–280

Fig. 3. Possible mechanisms and effects of oxPAPC-, PGPC-, or POVPC-induced activation of endothelial cells and VSMCs in atherosclerosis. Oxidized phospholipids stimulate the PI3K/Akt pathway leading to the phosphorylation of eNOS and the generation of NO [81] as well as the production of IL-8 by activating SREBP, involved in cholesterol metabolism in endothelial cells [111]. In VSMCs, oxidized phospholipids activate the aSMase/SM/ceramide pathway, inducing either apoptosis or proliferation [74,86].

metabolism that bind to the IL-8 promoter region [111] resulting in increased synthesis of IL-8. Interestingly, although oxidized phospholipids have been shown to contribute to endothelial dysfunction and the action of inflammatory pathways involving NO in relation to atherosclerosis, in lung vascular inflammation they appear to have protective effects, as discussed later. Chlorinated lipids have also been found to have an effect on NO bioavailability in endothelial cells. Treatment with hypochloritemodified LDL of human umbilical cord endothelial cells (HUVECs) caused a decrease in synthesis of cGMP compared with native LDL [112]. This was coupled with the translocation of eNOS away from the plasma membrane, reducing the formation of NO within the cells [112]. Phospholipids modified by the MPO/hydrogen peroxide/ chloride pathway again caused eNOS uncoupling from the plasma membrane and therefore decreased NO production in which 2ClHDA was found to be the active component [113]. Leukocyte and monocyte binding Many diseases with an inflammatory component, such as atherosclerosis, inflammatory bowel disease, and rheumatoid synovitis [114], rely on the adhesion and transmigration of inflammatory cells for their propagation. Although the basic mechanisms of leukocyte adhesion to vascular tissues have been known for some years [115], recent work has focused on pathways that regulate adhesion, the interactions between blood-borne cells, and the importance of modified lipids in modulating leukocyte adhesion. Acute injury or activation of the endothelium recruits neutrophils, the so-called “all-terrain vehicles” of innate immunity [116]. Many of the adhesion molecules that regulate neutrophil adhesion, such as platelet endothelial cell adhesion molecule, ICAM-1, and junction adhesion molecules, are expressed at increased levels at endothelial cell junctions [117]. This may explain why transmigration preferentially occurs at these sites. The endothelium plays a crucial role in sensing the inflammatory stimulus and expressing the relevant adhesion molecules to allow leukocyte adhesion to occur. Thus in diseases such as atherosclerosis, in which lipid-laden and chemokine-producing foam cells are present in the vessel intima, the endothelium will be in a constant state of

activation, which will set up a cycle of inflammation. This section focuses on the effects of oxidized and chlorinated lipids on inflammatory cell adhesion and their influence on disease progression. In an air pouch model of inflammation, oxPAPC induced the selective adhesion of monocytic cells to the air pouch wall, whereas LPS induced accumulation of both monocytes and neutrophils [114]. Immunohistochemical and RT-PCR analysis revealed that, whereas LPS induced adhesion molecule up-regulation, oxPAPC increased the expression of several MCPs, RANTES, and growth-related oncogene α. In CCR2 knockout mice, which lack the receptor for the MCPs, oxPAPC did not induce monocyte adhesion. Similarly, oxPAPC and POVPC induced adhesion of monocytic U937 cells and human peripheral blood monocytes, but not blood neutrophils, to HUVECs in a static adhesion system [118]. In this study oxPAPC induced MAPK pathways in endothelial cells, which involved activation of phospholipase A2 and 12-lipoxygenase. In HAECs, oxPAPC, but not the nonoxidized PAPC, rapidly and transiently induced mitogen-activated protein kinase phosphatase-1 (MKP-1), which mediated monocyte chemotactic activity via production of MCP-1 [119], and inhibition of MKP-1 using a chemical or antisense approach prevented oxPAPC-induced monocyte adhesion. In vivo studies by the Leitinger group also demonstrated a positive effect of oxPAPC on monocyte adhesion in mice [120]. In this study, they observed that application of oxPAPC in pluronic gel to the mouse carotid artery induced atherosclerosis-related gene expression in the artery wall and also monocyte adhesion in isolated, perfused carotid arteries. In atherosclerotic mice, oxPAPC-regulated chemokines were found in the lesions, indicating that oxidized phospholipids, by stimulating chemokine expression, may influence monocyte adhesion, which progresses the disease. Other phospholipids derived from oxidation of PAPC have also been shown to modulate endothelial function and monocyte adhesion [9,121] and HOOA-PC, at a concentration of 10 μg/ml, enhanced monocyte adhesion to HAECs [122]. HOOA-PC also induced MCP-1 and IL-8 expression and inhibited E-selectin on the endothelium in response to LPS, indicating that this modified lipid would selectively enhance monocyte and not neutrophil adhesion. Such a selective effect on monocyte adhesion has also been observed with other, structurally related oxidized phospholipids [123]. One consideration is that many adhesion studies are performed in static

F.H. Greig et al. / Free Radical Biology & Medicine 52 (2012) 266–280

systems, which, although simple, do not mimic the pulsatile flow conditions encountered in vivo. Indeed, a study by Hsiai et al. [124] found that, whereas oxPAPC induced monocyte adhesion to BAECs, high shear stress slew rates significantly reduced the effect of oxPAPC, as did, to a lesser extent, low shear. Oscillating flow enhanced the effects of oxPAPC on monocyte adhesion [124], whereas in all cases, monocyte adhesion was correlated with expression of MCP-1 by the BAECs. At an intracellular level, treatment of HUVECs with aldehydemodified PEs (adducts with 4-hydroxynonenal, 5-oxopentanal, or full-chain isolevuglandins) allowed rapid internalization of the peroxidized phospholipid to the endoplasmic reticulum (ER) and subsequent expression of markers of ER stress such as CHOP and BiP, as well as p38 MAPK activity [125]. Because these ER stress markers are linked to inflammatory chemokine expression, the authors concluded that this may represent one mechanism by which modified lipids activate the endothelium to induce leukocyte adhesion. In addition to their well-characterized effects on monocyte adhesion, oxidized phospholipids can have effects on endothelial cell chemotaxis and angiogenesis. OxPAPC treatment of human endothelial cells was found to reduce gene expression of proangiogenic IL-15 but to up-regulate IL-8 [126]. Presumably the angiogenic potential of oxLDL will be determined by local conditions and the individual components of the oxLDL. Effects on endothelial integrity and intraplaque vascularization would have predictably profound effects on leukocyte recruitment as well as thromboregulation and risk of intraplaque hemorrhage [127]. Because adhesion of monocytes to endothelium is an interactive process, research has also been directed toward understanding how oxidized phospholipids influence monocyte/ macrophage behavior. In human THP-1 macrophages, oxLDL markedly enhanced CCL23 release, a member of the CC chemokine family. The CCL23 stimulated THP-1 chemotaxis, expression of the integrin adhesion molecule CD11c, and release of the matrix metalloproteinase MMP-2 [128]. Furthermore, in human atherosclerotic lesions, CCL23 expression was increased and colocalized with the specific macrophage marker CD68. These data suggest that CCL23 is important in the development of atherosclerosis, and oxidized phospholipids, as components of oxLDL, may well exert similar effects on macrophages. In contrast to oxidized phospholipids, much less has been published on the effects of chlorinated lipids on inflammatory cell adhesion. LysoPC chlorohydrin was identified in human atherosclerotic lesions and led to an increase in the expression of P-selectin on the endothelial cell surface of HCAECs [66]. Interestingly, the chlorinated form of lysoPC differed from the nonchlorinated form in its effect on COX-2, demonstrating that chlorination can have a profound effect on the biological activity of the lipid. In addition to chlorinating susceptible lipids, MPO itself can influence inflammatory cell activity. Klinke et al. [129] demonstrated that neutrophil-derived MPO may influence recruitment of further neutrophils, through an electrostatic interaction between MPO itself and the neutrophil surface after deposition of MPO on the endothelial surface. Dever et al. [130] found that 1-stearoyl-2-oleoyl-snglycero-3-phosphocholine (SOPC) chlorohydrin induced an increase in leukocyte adhesion to atherosclerotic mouse aorta, which was dependent on the expression of P-selectin and ICAM-1 by the vascular endothelium [130]. The effect of the chlorohydrin seemed to be primarily on the artery, as P-selectin expression was up-regulated, whereas leukocyte expression of LFA-1 or PSGL-1 was unchanged. Five-day treatment with the HMG-CoA reductase inhibitor pravastatin or an NO-donating pravastatin derivative abrogated the effects of the SOPC chlorohydrins [131]. Interestingly, a similar effect of a statin has been observed in HCAECs treated with 60 μg/ml oxLDL for 24 h [132]. In this study, a combination of aspirin and pravastatin attenuated the monocyte adhesion induced by oxLDL. In common with the study by Dever, ICAM-1 appeared to be important in mediating the enhanced monocyte adhesion and this, as well as the chemotactic agent MCP-1, was attenuated after aspirin/pravastatin treatment. These two studies show that statins may exert some of their pleiotropic

273

effects by preventing the action of modified phospholipids on inflammatory cell adhesion. Receptors for oxidatively modified lipids One of the critical questions that has been exercising researchers ever since oxidized phospholipids were first found to have proinflammatory effects is which receptors on or within cells are responsible for recognizing oxidized phospholipids and stimulating the signaling pathways and outcomes described above. Several different types of receptor have been investigated, and interactions with scavenger receptors, peroxisome proliferator-activated receptors (PPARs), the platelet-activating factor (PAF) receptor, and Toll-like receptors (TLRs) are well accepted and discussed below, although there have been some controversies along the path to our current understanding. Other receptors that have been investigated include the vascular endothelial growth factor receptor (VEGFR) and GPCRs. Zimman et al. have reported that VEGFR is involved in the response of HAECs to oxPAPC in a c-Src-dependent mechanism, although it was not established whether oxPAPC interacted directly with VEGFR [133]. Early research on the possible roles of GPCRs focused on the effects of mmLDL and a putative Gs-coupled receptor [134], although it was subsequently suggested that free oxidized fatty acids were in fact responsible for the effects following hydrolytic release from oxidized phospholipids [135]. More recently it has been found that other GPCRs, specifically prostaglandin E2 and prostaglandin D2 receptors, can be activated by oxPAPC and PEIPC, but not POVPC, and these effects do seem to involve direct binding to the receptors, although it was unclear whether oxidized phospholipids or free oxidized fatty acids were responsible [136]. Platelet-activating factor receptor The PAF receptor is also a GPCR [137] and was one of the earliest suggested targets of oxidized phospholipids, owing to the structural similarities between PAF and truncated products from the oxidation of phospholipids [138]. The activation of PAF receptor by specific oxidized lipids, also termed PAF-like lipids, was demonstrated in vascular cells and macrophages [139,140], although the presence of PAF itself in the oxLDL cannot be ruled out [141]. One of the actions of oxidized phospholipids on the PAF receptor is the activation of platelets, inducing platelet aggregation [142]. The importance of the PAF receptor in phospholipid signaling is further seen from the fact that two competitive antagonists at the PAF receptor inhibited VSMC proliferation induced by oxLDL [143]. Similarly, monocyte binding to HAECs induced by oxLDL is also blocked by PAF receptor antagonists; however, PAF alone did not induce monocyte binding [144], which suggests that oxPAPC might be eliciting its effects through a receptor distinct from the PAF receptor. PGPC can activate human neutrophils, and this was also blocked by the action of PAF receptor antagonists [145]. Research has now moved away from the PAF receptor being the primary mechanism of modified phospholipid signaling, as some of the actions of modified phospholipids cannot be mimicked by PAF alone. Even in platelet activation by SAzPC, SOVPC, and SGPC, an increase in intracellular Ca 2 + was not observed, suggesting that the PAF receptor is not responsible for this effect [146]. Scavenger receptors The uncontrolled uptake of oxLDL by macrophages [147] and VSMCs has been implicated in the formation of lipid-laden foam cells and the development of atherosclerotic plaques. Modified lipids have been recognized as possible ligands for scavenger receptor families such as cluster differentiation 36 (CD36), scavenger receptor class A types I and II (SRA-I/II), and the class B scavenger receptor. The CD36 receptor is a platelet-integral membrane glycoprotein and is

274

F.H. Greig et al. / Free Radical Biology & Medicine 52 (2012) 266–280

highly conserved between humans and rodents [148]. CD36 has been demonstrated to be crucial in the progression of atherosclerosis, and mmLDL up-regulates the expression of scavenger receptors in mouse macrophages, including CD36 and SRA [149,150]. The importance of CD36 has been shown by studies in which CD36–ApoE doubleknockout mice were found to have a dramatic reduction in lesion area compared with ApoE−/− mice, both fed on a high-fat diet [151]. Furthermore, SRA-I/II and CD36 have been shown to be the primary receptors involved in the uptake of modified phospholipids by macrophages, as genetic inactivation of both of these receptors caused a significant decrease in atherosclerotic plaques in mice [152]. Some truncated oxidized phospholipids containing hydroxyalkenal moieties have been found to bind with high affinity to the CD36 receptor [153], although other oxidized phospholipids such as POVPC and oxidized PS have also been shown to be ligands at the receptor, which in turn was found to play an important role in macrophage phagocytosis of apoptotic cells [154]. The CD36 receptor has been found to play a critical role in oxLDL-induced adhesion of macrophages to endothelial cells [155]. Scavenger receptors have also been shown to recognize hypochlorite-modified lipids, demonstrated in macrophages showing uptake by CD36 and SRB-I, whereas HOCl–LDL was not recognized by SRA-I [156]. Furthermore, Chinese hamster cells overexpressing SRB-I were incubated with low concentrations of HOCl-modified HDL and found to show impaired efflux of cholesterol from the cells. HOCl–HDL also acted as a competitive inhibitor at SRB-I against the native HDL [157]. This suggests that the antiatherogenic properties of HDL are lost after modification by HOCl, leading to lipid accumulation and foam cell formation. Peroxisome proliferator-activated receptors PPARs are intracellular ligand-activated transcription factors and part of the nuclear receptor superfamily. PPARs are reported to play important roles in the control of the inflammatory response and also in lipid metabolism and are divided into three PPAR subtypes: PPARα, PPARγ, and PPARδ. PPARs can result in either pro- or antiinflammatory outcomes, and both have been observed after treatment of cells with oxidized phospholipids. PPARs regulate gene expression by binding with retinoid-X-receptor as a heterodimeric partner to specific DNA sequence elements termed PPAR-response elements on target genes [158]. The importance of PPARα in inflammatory responses was demonstrated in a study in which Ldlr −/− mice were transfused with reconstituted bone marrow cells from either PPARα−/− or PPARα+/+ mice and then fed on a high-fat diet [159]. The size of the atherosclerotic plaques was significantly larger in the mice containing PPARα −/− cells compared with controls, and macrophages from PPARα −/− mice also demonstrated a significantly increased uptake of oxLDL [159], which suggests an anti-inflammatory action. LDL modified by phospholipase A2 activated PPARδ in macrophages and changed gene expression in relation to lipid metabolism, also suggesting an anti-inflammatory response [160]. In contrast, it has been observed that incubation of mmLDL and the oxidation products of PAPC with transfected HAECs activated PPARα. Furthermore, a synthetic ligand for PPARα caused an increase in IL-8 and MCP-1 production in the HAECs, suggesting a proinflammatory role for the receptor in that cell type [161]. OxLDL has been reported to be a ligand for both PPARα and PPARγ; it activated both subtypes in macrophages, as well as causing the expression of COX-2, which was mediated by the action of ERK1/2 [162]. Oxidatively fragmented alkyl phospholipids, present in oxLDL, have been found to have high affinity for PPARγ [163]. Activation of this subtype by oxidized phospholipids has also been implicated in the switching of chemokine receptors from CCR2 on monocytes to CX3CR1 on activated macrophages involved in foam cell formation and macrophage adhesion [164]. PPARδ had previously been demonstrated to be involved in monocytic differentiation to macrophages, as well as promoting uptake of

oxLDL by CD36 scavenger receptor [165]. In addition, CD36 expression is regulated by PPARγ, and ligands for the receptor have been reported to increase CD36 expression [166]. This led to the idea of a feed-forward loop between CD36 and PPARγ, as their interaction caused a proatherogenic action within the vessel wall [165]. Thus it can be seen that PPAR activation by oxidized phospholipids has shown both pro- and anti-inflammatory actions with importance in inflammatory signaling. Toll-like receptors Another family of cell surface and intracellular receptors that have been enthusiastically investigated for their potential role in signaling proinflammatory effects of oxidized phospholipids are the TLRs. These are innate immune receptors that recognize pathogenassociated molecular patterns (PAMPs) or respond to products of host tissue damage [167]. As there is some structural similarity between oxidized phospholipids and LPS, an agonist of TLR4, it was hypothesized that oxidized phospholipids might activate TLRs to cause proinflammatory responses. TLR4 is responsible for detecting a gram-negative LPS (e.g., from Escherichia coli) as well as other PAMPs such as zymosan, whereas TLR2 heterodimers with TLR1 or TLR6 are able to detect cell wall lipopeptides from gram-positive bacteria. Some early studies provided support for the concept that oxidized phospholipids signaled via TLRs: Miller et al. observed that TLR4 was involved in macrophage responses to mmLDL [168] and in HeLa cells it was found that production of IL-8 in response to oxPAPC was dependent on TLR4 [169]. On the other hand, in these studies it was also noted that NF-κB did not seem to be involved in the oxPAPC-induced signaling and that TNFα expression was not increased; this was surprising as NF-κB activation represents a major pathway for inducing TLR4-dependent proinflammatory gene expression, of which TNFα expression is characteristic [170]. Other studies also supported these findings: Erridge et al. observed that although oxPAPC could induce the production of IL-8 and MCP-1/ MIP-2 and expression of E-selectin in myeloid and endothelial cells, it did not induce production of TNFα [171]. This suggested that if TLRs were able to act as receptors for oxidized phospholipids, the cellular signaling pathway did not follow the classical NF-κB-dependent route. In fact, studies of the dependence of IL-8 promoter expression on TLRs 1, 2, 4, and 6 in response to oxidized phospholipids that were carried out by transfection of a TLR-deficient cell line, HEK-293, suggested that these TLRs were not required for IL-8 up-regulation. The authors proposed that these differences were due to the differential responses of macrophages and HeLa cells to cholesterol depletion and lipid raft disruption induced by oxidized phospholipids: HeLa cells respond by activating NF-κB, whereas in many other cell lines, including macrophages, this prevents TLR dimerization and signaling [172]. An alternative explanation for these results is that oxidized phospholipid interactions with TLR4 cause an activation of the signaling factor TRIF and follow an alternative signaling pathway leading to activation of interferon-regulatory factor 3 and expression of interferon-β and interferon-inducible genes, with some cross talk between transcription factors and gene expression (Fig. 4). Support for this has come from a study that reported that oxPAPC can cause lung injury and IL-6 production by mouse lung macrophages in a pathway involving TLR4/TRIF/ TRAF6 and IKKε activation [173]. It has also been suggested that oxidized phospholipids may not interact directly with TLRs, but may require costimulatory factors to elicit a proinflammatory response. Seimon et al. observed that oxidized phospholipids could cause enhanced macrophage apoptosis under conditions of ER stress and that this required both TLR2/6 and CD36 [174]. It is also worth noting that in murine preosteoblasts, oxPAPC at levels of 40–80 μg/ml did cause increased production of TNFα, though a 72-h time course was required [175]. Thus the proinflammatory effects of oxidized phospholipids via TLRs seem to be cell specific, and it is also important to note

F.H. Greig et al. / Free Radical Biology & Medicine 52 (2012) 266–280

275

Fig. 4. Simplified scheme of interactions between oxidized phospholipids and TLRs, showing potential sites of inhibition (red blocks) and activation (purple arrows). Question marks indicate where the pathways or sites of interaction are not fully understood, but evidence for effects has been obtained. Some or all of these responses have been observed with monocytes/macrophages, dendritic cells, and endothelial cells. CpG, CpG DNA; dsRNA, double-stranded RNA; IRF3, interferon-regulatory factor 3; MD2, myeloid differentiation factor 2; MyD88, myeloid differentiation factor 88; TRIF, TIR-domain-containing adapter-inducing interferon-β; other abbreviations are given in the main list. Note that only the adaptor proteins that define the pathways are shown.

that inhibitory effects of oxidized phospholipids on TLRs also exist, as described below. Anti-inflammatory and beneficial effects of oxidized phospholipids Although many reported effects of oxidized phospholipids are proinflammatory, it is clear that they also have protective and antiinflammatory effects, and in some cases the receptors and signaling components involved are shared with the anti-inflammatory effects. The main mechanisms by which anti-inflammatory effects occur involve the activation of certain subtypes of PPARs as described in the previous section, as well as the activation of small GTPases. There is mounting evidence that oxidized lipids can abrogate PAMP-induced signaling via TLRs. It has been known for some years that oxPAPC, POVPC, and hydroxyalkenal-containing phospholipids prevent LPS-induced neutrophil–endothelial adhesion and E-selectin up-regulation via NFκB activation [122,123,176]. LPS activation of TLR4 is responsible for the septic shock response in endotoxemia, and it was shown that oxPAPC increased the survival of mice injected with lethal doses of LPS and reduced the expression of inflammatory markers [176]. This group demonstrated that the effect was due to competitive binding by oxPAPC blocking the interaction of LPS with its binding protein LBP and CD14, both of which are required for activation of TLR4 (Fig. 4). Subsequently it was shown that TLR2 activation by bacterial lipoprotein was inhibited by the same mechanism and that oxPAPC also binds competitively to the accessory protein MD-2 as an additional mechanism for inhibiting TLR4 signaling [172,177]. Although in macrophages and embryonic kidney cells transfected with TLRs, TLR2 and TLR4 were found to be the only TLRs whose activation was abrogated by oxPAPC, some other studies have reported effects on TLR3 and TLR9. In dendritic cells, it has been found that oxPAPC can inhibit the production of TNFα

and IL-12 p40 induced by the TLR3 ligand poly(I:C), and TLRindependent inhibitory effects have also been reported for sCD40 ligand-induced responses [178]. The inhibition of TLR9 signaling was reported by Ma et al., who observed that oxPAPC inhibited the production of TNFα in response to CpG DNA in cultured macrophages and mice [179]. The effects of a number of different classes and species of oxidized phospholipids were investigated by Oskolkova et al., and all showed similar biphasic concentration profiles in terms of inhibiting LPSinduced IL-8 expression; it was also found that the compounds are stronger antagonists of LPS than direct agonists of inflammation [180]. Although the TLR-inhibitory effects of oxPAPC and its components undoubtedly are beneficial for endotoxemia and some lung pathologies due to infection (as described below), it is also clear that these immunosuppressive effects also have their downside: oxPAPC administration to mice decreased phagocytosis by neutrophils and macrophages and rendered mice more susceptible to E. coli peritonitis [181]. There has been a significant amount of work on protective effects of oxidized phospholipids in lung inflammation, which arises in conditions such as acute lung injury and adult respiratory distress syndrome; these are modeled in animals using mechanical lung injury, sterile immune insult using LPS, or infections. A major factor in lung injury is increased vascular leak, involving loss of endothelial cell barrier function, which allows migration of activated leukocytes to the interstitial fluid resulting in oxidative damage. There are now reports that several oxidized phospholipids, including PECPC, prevent the loss of barrier function, in contrast to unoxidized phospholipids, which had no effect [182]. For example, in rats given aerosolized LPS, concomitant injection of oxPAPC attenuated the production of inflammatory cytokines and barrier disruption [183]. This protective effect seems to depend on oxPAPC activation of three small GTPases, Rho, Rac, and Cdc42, in pulmonary endothelial cells [182], although the receptor involved in the effect is as yet unclear. The outcome of these signaling pathways is a cytoskeletal rearrangement resulting in the formation of microspike-

276

F.H. Greig et al. / Free Radical Biology & Medicine 52 (2012) 266–280

like actin structures at endothelial cell–cell interfaces and enhanced expression of proteins required for tight junctions and adherens junctions. It is known that protein kinase A, protein kinase C (PKC), and Src are involved upstream of the GTPases and that the guanine nucleotide exchange factors Tiam1 and bPIX are required [184–186], although it is not known whether the phospholipids activate the GTPases by direct engagement or by binding to a cell surface receptor such as VEGFR2. Interestingly, TLR4 can signal to c-Src and cAMP, whereas CD36 signals to the latter only, and PAF receptor and VEGFR2 are both upstream of PKC [6]. Other possible mechanisms may include the phosphorylation and activation of the focal adhesion proteins paxillin and focal adhesion kinase, as well as actin-binding proteins and cofillin [68]. In addition to these well-established anti-inflammatory mechanisms, other effects have been reported for oxidized phospholipids that can be considered beneficial. It has been known for some time that oxLDL and oxPAPC can increase glutathione levels in endothelial cells and protect against oxidant-induced toxicity [187], but more recently it has been found that this effect involves activation of Nrf2 and its translocation to the nucleus to up-regulate expression of genes containing an antioxidant-response element, such as glutamate–cysteine ligase, heme oxygenase-1, and NAD(P)H quinone oxidoreductase-1 [188]. The effect was observed for phospholipids containing an electrophilic epoxyisoprostane ring and are in line with current understanding on the ability of free hydroxyalkenals to induce Nrf2. Another potentially protective effect reported was the activation of phospholipase A2 by PONPC, which similarly was found to involve the formation of a Schiff's base adduct between the protein and the phospholipid [189]. As specific hydrolysis of oxidized chains at the sn-2 position is thought to contribute to detoxification, this could represent a feedback mechanism for limiting proinflammatory effects. Summary: advances and limitations Accumulating data from the past decade clearly demonstrate that oxidized and chlorinated lipids and phospholipids are formed in vivo, in a variety of pathological situations. In parallel, there are a steadily increasing number of studies reporting biological effects of oxLDL and individual oxidized and chlorinated lipid species on cells and tissues, and our understanding of the responses plus underlying mechanisms has advanced enormously. One emerging paradigm is that the response to individual species can differ substantially between cell or tissue types, to the extent that both beneficial and deleterious effects are possible. An example is the barrier-protective effect and inhibition of leukocyte migration in pulmonary endothelial cells contrasted with increased leukocyte–endothelial adhesion and inflammation in coronary or aortic endothelial cells, or the role of acid SMase in vascular cells versus neutral SMase in neuron apoptosis. Another finding is that individual species of oxidized phospholipid can cause different effects on the same cell type: in particular, truncated products often show different effects compared to full-chain-length oxidized products, and reactive, electrophilic compounds such as hydroxyalkenals have different effects compared to species containing carboxylic acid groups. The former may relate to the extent to which the oxidized chain protrudes from the lipid bilayer and the latter to the ability to form covalent adducts with proteins or DNA, thus altering structure and activity. One important issue, which may limit the interpretation of studies using oxidized lipoproteins, is the question of whether it is better to investigate the effects of a biologically relevant mixture, such as oxidized lipoproteins, or individual oxidized phospholipids. As mentioned in the introduction, lipoproteins are composed of many different oxidized and nonoxidized compounds, so it is difficult to determine which are responsible for the bioactivity observed. Even oxPAPC may contain some free aldehydes formed from fragmentation of the oxidized chains, which also have cytotoxic and bioactive effects. Also, the varied responses sometimes observed by different researchers may reflect different molecular compositions of the oxPAPC or

lipoproteins or the fact that many effects are dose-dependent. Therefore, studies with pure preparations of oxidized phospholipids are ultimately needed to confirm the causative agents, but should be combined with studies using more physiologically relevant mixtures to provide information on the likely balance of response in vivo. Given the large number of different oxidized phospholipid species that have been identified in several analyses of human and animal tissues, it seems likely that the situation in vivo is highly complex. The concentrations of oxidized phospholipids reported in human tissue samples on the whole match adequately with the concentrations used in studies of the effects of individual oxidized species in vitro, and therefore it can be concluded that the effects observed in experiments with cultured cells and tissues are of relevance to human pathology. From such research, the anti-inflammatory and arguably beneficial effects of the family of oxidized phospholipids are now becoming established, as discussed in several reviews [3,6,68,190]. It seems that the formation of oxidized phospholipids may sometimes act as a feedback loop, limiting the extent to which inflammation can progress. This has led to interest in the potential therapeutic value of these compounds and the synthesis of analogues such as lecinoxoids with enhanced biological stability [190]. Acknowledgments The authors gratefully acknowledge support from the British Heart Foundation in the form of Ph.D. Studentship Grant FS/08/071/ 26212 to F.H.G. C.M.S. thanks members of the 08-EuroMEMBRANESFP-021 Consortium for stimulating discussions in the field. References [1] Medzhitov, R. Origin and physiological roles of inflammation. Nature 454: 428–435; 2008. [2] Leitinger, N. The role of phospholipid oxidation products in inflammatory and autoimmune diseases: evidence from animal models and in humans. Subcell. Biochem. 49:325–350; 2008. [3] Bochkov, V. N.; Oskolkova, O. V.; Birukov, K. G.; Levonen, A. L.; Binder, C. J.; Stockl, J. Generation and biological activities of oxidized phospholipids. Antioxid. Redox Signal. 12:1009–1059; 2010. [4] Huber, J.; Vales, A.; Mitulovic, G.; Blumer, M.; Schmid, R.; Witztum, J. L.; Binder, B. R.; Leitinger, N. Oxidized membrane vesicles and blebs from apoptotic cells contain biologically active oxidized phospholipids that induce monocyte– endothelial interactions. Arterioscler. Thromb. Vasc. Biol. 22:101–107; 2002. [5] Spickett, C. M.; Wiswedel, I.; Siems, W.; Zarkovic, K.; Zarkovic, N. Advances in methods for the determination of biologically relevant lipid peroxidation products. Free Radic. Res. 44:1172–1202; 2010. [6] Fruhwirth, G. O.; Loidl, A.; Hermetter, A. Oxidized phospholipids: from molecular properties to disease. Biochim. Biophys. Acta 1772:718–736; 2007. [7] Conrad, M.; Schneider, M.; Seiler, A.; Bornkamm, G. W. Physiological role of phospholipid hydroperoxide glutathione peroxidase in mammals. Biol. Chem. 388:1019–1025; 2007. [8] Spiteller, G. Peroxyl radicals: inductors of neurodegenerative and other inflammatory diseases. Their origin and how they transform cholesterol, phospholipids, plasmalogens, polyunsaturated fatty acids, sugars, and proteins into deleterious products. Free Radic. Biol. Med. 41:362–387; 2006. [9] Watson, A. D.; Leitinger, N.; Navab, M.; Faull, K. F.; Horkko, S.; Witztum, J. L.; Palinski, W.; Schwenke, D.; Salomon, R. G.; Sha, W.; Subbanagounder, G.; Fogelman, A. M.; Berliner, J. A. Structural identification by mass spectrometry of oxidized phospholipids in minimally oxidized low density lipoprotein that induce monocyte/endothelial interactions and evidence for their presence in vivo. J. Biol. Chem. 272:13597–13607; 1997. [10] Spickett, C. M.; Dever, G. Studies of phospholipid oxidation by electrospray mass spectrometry: from analysis in cells to biological effects. Biofactors 24:17–31; 2005. [11] Lapenna, D.; Pierdomenico, S. D.; Ciofani, G.; Ucchino, S.; Neri, M.; Giamberardino, M. A.; Cuccurullo, F. Association of body iron stores with low molecular weight iron and oxidant damage of human atherosclerotic plaques. Free Radic. Biol. Med. 42: 492–498; 2007. [12] Trudel, S.; Kelly, M.; Fritsch, J.; Nguyen-Khoa, T.; Therond, P.; Couturier, M.; Dadlez, M.; Debski, J.; Touqui, L.; Vallee, B.; Ollero, M.; Edelman, A.; Brouillard, F. Peroxiredoxin 6 fails to limit phospholipid peroxidation in lung from Cftrknockout mice subjected to oxidative challenge. PLoS One 4:e6075; 2009. [13] Isik, A.; Koca, S. S.; Ustundag, B.; Celik, H.; Yildirim, A. Paraoxonase and arylesterase levels in rheumatoid arthritis. Clin. Rheumatol. 26:342–348; 2007. [14] Precourt, L. P.; Amre, D.; Denis, M. C.; Lavoie, J. C.; Delvin, E.; Seidman, E.; Levy, E. The three-gene paraoxonase family: physiologic roles, actions and regulation. Atherosclerosis 214:20–36; 2011.

F.H. Greig et al. / Free Radical Biology & Medicine 52 (2012) 266–280 [15] Pereira, I. A.; Borba, E. F. Multiple factors determine the increased prevalence of atherosclerosis in rheumatoid arthritis. Acta Reumatol. Port. 33:47–55; 2008. [16] Ross, R. Atherosclerosis—an inflammatory disease. N. Engl. J. Med. 340:115–126; 1999. [17] Witztum, J. L.; Steinberg, D. Role of oxidized low density lipoprotein in atherogenesis. J. Clin. Invest. 88:1785–1792; 1991. [18] Chisolm, G. M.; Steinberg, D. The oxidative modification hypothesis of atherogenesis: an overview. Free Radic. Biol. Med. 28:1815–1826; 2000. [19] Hoff, H. F.; O'Neil, J.; Wu, Z.; Hoppe, G.; Salomon, R. L. Phospholipid hydroxyalkenals: biological and chemical properties of specific oxidized lipids present in atherosclerotic lesions. Arterioscler. Thromb. Vasc. Biol. 23:275–282; 2003. [20] Ravandi, A.; Babaei, S.; Leung, R.; Monge, J. C.; Hoppe, G.; Hoff, H.; Kamido, H.; Kuksis, A. Phospholipids and oxophospholipids in atherosclerotic plaques at different stages of plaque development. Lipids 39:97–109; 2004. [21] Davis, B.; Koster, G.; Douet, L. J.; Scigelova, M.; Woffendin, G.; Ward, J. M.; Smith, A.; Humphries, J.; Burnand, K. G.; Macphee, C. H.; Postle, A. D. Electrospray ionization mass spectrometry identifies substrates and products of lipoproteinassociated phospholipase A2 in oxidized human low density lipoprotein. J. Biol. Chem. 283:6428–6437; 2008. [22] Chabot, F.; Mitchell, J. A.; Gutteridge, J. M.; Evans, T. W. Reactive oxygen species in acute lung injury. Eur. Respir. J. 11:745–757; 1998. [23] Lang, J. D.; McArdle, P. J.; O'Reilly, P. J.; Matalon, S. Oxidant–antioxidant balance in acute lung injury. Chest 122:314S–320S; 2002. [24] Nakamura, T.; Henson, P. M.; Murphy, R. C. Occurrence of oxidized metabolites of arachidonic acid esterified to phospholipids in murine lung tissue. Anal. Biochem. 262:23–32; 1998. [25] Matot, I.; Manevich, Y.; Al-Mehdi, A. B.; Song, C.; Fisher, A. B. Fluorescence imaging of lipid peroxidation in isolated rat lungs during nonhypoxic lung ischemia. Free Radic. Biol. Med. 34:785–790; 2003. [26] Tyurin, V. A.; Tyurina, Y. Y.; Jung, M. Y.; Tungekar, M. A.; Wasserloos, K. J.; Bayir, H.; Greenberger, J. S.; Kochanek, P. M.; Shvedova, A. A.; Pitt, B.; Kagan, V. E. Mass-spectrometric analysis of hydroperoxy- and hydroxy-derivatives of cardiolipin and phosphatidylserine in cells and tissues induced by pro-apoptotic and pro-inflammatory stimuli. J Chromatogr B Analyt Technol Biomed Life Sci 877: 2863–2872; 2009. [27] Fessel, J. P.; Porter, N. A.; Moore, K. P.; Sheller, J. R.; Roberts II, L. J. Discovery of lipid peroxidation products formed in vivo with a substituted tetrahydrofuran ring (isofurans) that are favored by increased oxygen tension. Proc. Natl. Acad. Sci. U. S. A. 99:16713–16718; 2002. [28] Yoshimi, N.; Ikura, Y.; Sugama, Y.; Kayo, S.; Ohsawa, M.; Yamamoto, S.; Inoue, Y.; Hirata, K.; Itabe, H.; Yoshikawa, J.; Ueda, M. Oxidized phosphatidylcholine in alveolar macrophages in idiopathic interstitial pneumonias. Lung 183:109–121; 2005. [29] Cruz, D.; Watson, A. D.; Miller, C. S.; Montoya, D.; Ochoa, M. T.; Sieling, P. A.; Gutierrez, M. A.; Navab, M.; Reddy, S. T.; Witztum, J. L.; Fogelman, A. M.; Rea, T. H.; Eisenberg, D.; Berliner, J.; Modlin, R. L. Host-derived oxidized phospholipids and HDL regulate innate immunity in human leprosy. J. Clin. Invest. 118:2917–2928; 2008. [30] Van Lenten, B. J.; Wagner, A. C.; Navab, M.; Anantharamaiah, G. M.; Hui, E. K.; Nayak, D. P.; Fogelman, A. M. D-4F, an apolipoprotein A-I mimetic peptide, inhibits the inflammatory response induced by influenza A infection of human type II pneumocytes. Circulation 110:3252–3258; 2004. [31] Farooqui, A. A.; Horrocks, L. A. Lipid peroxides in the free radical pathophysiology of brain diseases. Cell. Mol. Neurobiol. 18:599–608; 1998. [32] Jolivalt, C.; Leininger-Muller, B.; Bertrand, P.; Herber, R.; Christen, Y.; Siest, G. Differential oxidation of apolipoprotein E isoforms and interaction with phospholipids. Free Radic. Biol. Med. 28:129–140; 2000. [33] Ross, B. M.; Moszczynska, A.; Erlich, J.; Kish, S. J. Low activity of key phospholipid catabolic and anabolic enzymes in human substantia nigra: possible implications for Parkinson's disease. Neuroscience 83:791–798; 1998. [34] Haider, L.; Fischer, M. T.; Frischer, J. M.; Bauer, J.; Hoftberger, R.; Botond, G.; Esterbauer, H.; Binder, C. J.; Witztum, J. L.; Lassmann, H. Oxidative damage in multiple sclerosis lesions. Brain 134:1914–1924; 2011. [35] Zagol-Ikapitte, I.; Masterson, T. S.; Amarnath, V.; Montine, T. J.; Andreasson, K. I.; Boutaud, O.; Oates, J. A. Prostaglandin H2-derived adducts of proteins correlate with Alzheimer's disease severity. J. Neurochem. 94:1140–1145; 2005. [36] Fessel, J. P.; Hulette, C.; Powell, S.; Roberts, L. J.; Zhang II, J. Isofurans, but not F2isoprostanes, are increased in the substantia nigra of patients with Parkinson's disease and with dementia with Lewy body disease. J. Neurochem. 85:645–650; 2003. [37] Govindarajan, B.; Laird, J.; Salomon, R. G.; Bhattacharya, S. K. Isolevuglandinmodified proteins, including elevated levels of inactive calpain-1, accumulate in glaucomatous trabecular meshwork. Biochemistry 47:817–825; 2008. [38] Al-Shawaf, E.; Naylor, J.; Taylor, H.; Riches, K.; Milligan, C. J.; O'Regan, D.; Porter, K. E.; Li, J.; Beech, D. J. Short-term stimulation of calcium-permeable transient receptor potential canonical 5-containing channels by oxidized phospholipids. Arterioscler. Thromb. Vasc. Biol. 30:1453–1459; 2010. [39] Wang, J.; Hu, B.; Kong, L.; Cai, H.; Zhang, C. Native, oxidized lipoprotein(a) and lipoprotein(a) immune complex in patients with active and inactive rheumatoid arthritis: plasma concentrations and relationship to inflammation. Clin. Chim. Acta 390:67–71; 2008. [40] Cvetkovic, J. T.; Wallberg-Jonsson, S.; Ahmed, E.; Rantapaa-Dahlqvist, S.; Lefvert, A. K. Increased levels of autoantibodies against copper-oxidized low density lipoprotein, malondialdehyde-modified low density lipoprotein and cardiolipin in patients with rheumatoid arthritis. Rheumatology (Oxford) 41:988–995; 2002.

277

[41] Gruber, F.; Oskolkova, O.; Leitner, A.; Mildner, M.; Mlitz, V.; Lengauer, B.; Kadl, A.; Mrass, P.; Kronke, G.; Binder, B. R.; Bochkov, V. N.; Leitinger, N.; Tschachler, E. Photooxidation generates biologically active phospholipids that induce heme oxygenase-1 in skin cells. J. Biol. Chem. 282:16934–16941; 2007. [42] Hahn, B. H.; McMahon, M. Atherosclerosis and systemic lupus erythematosus: the role of altered lipids and of autoantibodies. Lupus 17:368–370; 2008. [43] Hotamisligil, G. S. Inflammation and metabolic disorders. Nature 444:860–867; 2006. [44] Tsimikas, S.; Kiechl, S.; Willeit, J.; Mayr, M.; Miller, E. R.; Kronenberg, F.; Xu, Q.; Bergmark, C.; Weger, S.; Oberhollenzer, F.; Witztum, J. L. Oxidized phospholipids predict the presence and progression of carotid and femoral atherosclerosis and symptomatic cardiovascular disease: five-year prospective results from the Bruneck study. J. Am. Coll. Cardiol. 47:2219–2228; 2006. [45] Subbanagounder, G.; Watson, A. D.; Berliner, J. A. Bioactive products of phospholipid oxidation: isolation, identification, measurement and activities. Free Radic. Biol. Med. 28:1751–1761; 2000. [46] Podrez, E. A.; Byzova, T. V.; Febbraio, M.; Salomon, R. G.; Ma, Y.; Valiyaveettil, M.; Poliakov, E.; Sun, M.; Finton, P. J.; Curtis, B. R.; Chen, J.; Zhang, R.; Silverstein, R. L.; Hazen, S. L. Platelet CD36 links hyperlipidemia, oxidant stress and a prothrombotic phenotype. Nat. Med. 13:1086–1095; 2007. [47] Nicholls, S. J.; Hazen, S. L. Myeloperoxidase and cardiovascular disease. Arterioscler. Thromb. Vasc. Biol. 25:1102–1111; 2005. [48] Podrez, E. A.; Abu-Soud, H. M.; Hazen, S. L. Myeloperoxidase-generated oxidants and atherosclerosis. Free Radic. Biol. Med. 28:1717–1725; 2000. [49] Winterbourn, C. C.; van den Berg, J. J.; Roitman, E.; Kuypers, F. A. Chlorohydrin formation from unsaturated fatty acids reacted with hypochlorous acid. Arch. Biochem. Biophys. 296:547–555; 1992. [50] Spickett, C. M. Chlorinated lipids and fatty acids: an emerging role in pathology. Pharmacol. Ther. 115:400–409; 2007. [51] Jerlich, A.; Pitt, A. R.; Schaur, R. J.; Spickett, C. M. Pathways of phospholipid oxidation by HOCl in human LDL detected by LC-MS. Free Radic. Biol. Med. 28: 673–682; 2000. [52] Panasenko, O. M.; Spalteholz, H.; Schiller, J.; Arnhold, J. Myeloperoxidase-induced formation of chlorohydrins and lysophospholipids from unsaturated phosphatidylcholines. Free Radic. Biol. Med. 34:553–562; 2003. [53] Wildsmith, K. R.; Albert, C. J.; Anbukumar, D. S.; Ford, D. A. Metabolism of myeloperoxidase-derived 2-chlorohexadecanal. J. Biol. Chem. 281:16849–16860; 2006. [54] Albert, C. J.; Crowley, J. R.; Hsu, F. F.; Thukkani, A. K.; Ford, D. A. Reactive chlorinating species produced by myeloperoxidase target the vinyl ether bond of plasmalogens— identification of 8-chlorohexadecanal. J. Biol. Chem. 276:23733–23741; 2001. [55] Fernandes, R. M.; Silva, N. P.; Sato, E. I. Increased myeloperoxidase plasma levels in rheumatoid arthritis. Rheumatol. Int. (in press). doi:10.1007/s00296-011-1810-5. [56] Minohara, M.; Matsuoka, T.; Li, W.; Osoegawa, M.; Ishizu, T.; Ohyagi, Y.; Kira, J. Upregulation of myeloperoxidase in patients with opticospinal multiple sclerosis: positive correlation with disease severity. J. Neuroimmunol. 178:156–160; 2006. [57] Oottamasathien, S.; Jia, W.; McCoard, L.; Slack, S.; Zhang, J.; Skardal, A.; Job, K.; Kennedy, T. P.; Dull, R. O.; Prestwich, G. D. A murine model of inflammatory bladder disease: cathelicidin peptide induced bladder inflammation and treatment with sulfated polysaccharides. J. Urol. 186 (4 Suppl.):1684–1692; 2011. [58] Mohamadkhani, A.; Rastgar Jazii, F.; Sayehmiri, K.; Jafari-Nejad, S.; MontaserKouhsari, L.; Poustchi, H.; Montazeri, G. Plasma myeloperoxidase activity and apolipoprotein a-1 expression in chronic hepatitis B patients. Arch. Iran. Med. 14:254–258; 2011. [59] Samsamshariat, S. Z.; Basati, G.; Movahedian, A.; Pourfarzam, M.; Sarrafzadegan, N. Elevated plasma myeloperoxidase levels in relation to circulating inflammatory markers in coronary artery disease. Biomark. Med. 5:377–385; 2011. [60] Baldus, S.; Heeschen, C.; Meinertz, T.; Zeiher, A. M.; Eiserich, J. P.; Munzel, T.; Simoons, M. L.; Hamm, C. W. Myeloperoxidase serum levels predict risk in patients with acute coronary syndromes. Circulation 108:1440–1445; 2003. [61] Daugherty, A.; Dunn, J. L.; Rateri, D. L.; Heinecke, J. W. Myeloperoxidase, a catalyst for lipoprotein oxidation, is expressed in human atherosclerotic lesions. J. Clin. Invest. 94:437–444; 1994. [62] Brennan, M. L.; Penn, M. S.; Van Lente, F.; Nambi, V.; Shishehbor, M. H.; Aviles, R. J.; Goormastic, M.; Pepoy, M. L.; McErlean, E. S.; Topol, E. J.; Nissen, S. E.; Hazen, S. L. Prognostic value of myeloperoxidase in patients with chest pain. N. Engl. J. Med. 349:1595–1604; 2003. [63] Thukkani, A. K.; McHowat, J.; Hsu, F. -F.; Brennan, M. -L.; Hazen, S. L.; Ford, D. A. Identification of α-chloro fatty aldehydes and unsaturated lysophosphatidylcholine molecular species in human atherosclerotic lesions. Circulation 108:3128–3133; 2003. [64] Thukkani, A. K.; Martinson, B. D.; Albert, C. J.; Vogler, G. A.; Ford, D. A. Neutrophilmediated accumulation of 2-ClHDA during myocardial infarction: 2-ClHDAmediated myocardial injury. Am. J. Physiol. Heart Circ. Physiol. 288:H2955–H2964; 2005. [65] Ullen, A.; Fauler, G.; Kofeler, H.; Waltl, S.; Nusshold, C.; Bernhart, E.; Reicher, H.; Leis, H. J.; Wintersperger, A.; Malle, E.; Sattler, W. Mouse brain plasmalogens are targets for hypochlorous acid-mediated modification in vitro and in vivo. Free Radic. Biol. Med. 49:1655–1665; 2010. [66] Messner, M. C.; Albert, C. J.; McHowat, J.; Ford, D. A. Identification of lysophosphatidylcholine-chlorohydrin in human atherosclerotic lesions. Lipids 43:243–249; 2008. [67] Gibbons, G. H.; Dzau, V. J. The emerging concept of vascular remodeling. N. Engl. J. Med. 330:1431–1438; 1994. [68] Fu, P.; Birukov, K. G. Oxidized phospholipids in control of inflammation and endothelial barrier. Transl. Res. 153:166–176; 2009.

278

F.H. Greig et al. / Free Radical Biology & Medicine 52 (2012) 266–280

[69] Doran, A. C.; Meller, N.; McNamara, C. A. Role of smooth muscle cells in the initiation and early progression of atherosclerosis. Arterioscler. Thromb. Vasc. Biol. 28:812–819; 2008. [70] Pidkovka, N. A.; Cherepanova, O. A.; Yoshida, T.; Alexander, M. R.; Deaton, R. A.; Thomas, J. A.; Leitinger, N.; Owens, G. K. Oxidized phospholipids induce phenotypic switching of vascular smooth muscle cells in vivo and in vitro. Circ. Res. 101:792–801; 2007. [71] Cherepanova, O. A.; Pidkovka, N. A.; Sarmento, O. F.; Yoshida, T.; Gan, Q.; Adiguzel, E.; Bendeck, M. P.; Berliner, J.; Leitinger, N.; Owens, G. K. Oxidized phospholipids induce type VIII collagen expression and vascular smooth muscle cell migration. Circ. Res. 104:609–618; 2009. [72] van Tits, L. J.; Stienstra, R.; van Lent, P. L.; Netea, M. G.; Joosten, L. A.; Stalenhoef, A. F. Oxidized LDL enhances pro-inflammatory responses of alternatively activated M2 macrophages: a crucial role for Kruppel-like factor 2. Atherosclerosis 214:345–349; 2011. [73] Graham, L. S.; Parhami, F.; Tintut, Y.; Kitchen, C. M.; Demer, L. L.; Effros, R. B. Oxidized lipids enhance RANKL production by T lymphocytes: implications for lipid-induced bone loss. Clin. Immunol. 133:265–275; 2009. [74] Auge, N.; Garcia, V.; Maupas-Schwalm, F.; Levade, T.; Salvayre, R.; Negre-Salvayre, A. Oxidized LDL-induced smooth muscle cell proliferation involves the EGF receptor/ PI-3 kinase/Akt and the sphingolipid signaling pathways. Arterioscler. Thromb. Vasc. Biol. 22:1990–1995; 2002. [75] Chahine, M. N.; Blackwood, D. P.; Dibrov, E.; Richard, M. N.; Pierce, G. N. Oxidized LDL affects smooth muscle cell growth through MAPK-mediated actions on nuclear protein import. J. Mol. Cell. Cardiol. 46:431–441; 2009. [76] Chatterjee, S.; Berliner, J. A.; Subbanagounder, G. G.; Bhunia, A. K.; Koh, S. Identification of a biologically active component in minimally oxidized low density lipoprotein (MM-LDL) responsible for aortic smooth muscle cell proliferation. Glycoconj. J. 20:331–338; 2004. [77] Johnstone, S. R.; Ross, J.; Rizzo, M. J.; Straub, A. C.; Lampe, P. D.; Leitinger, N.; Isakson, B. E. Oxidized phospholipid species promote in vivo differential cx43 phosphorylation and vascular smooth muscle cell proliferation. Am. J. Pathol. 175:916–924; 2009. [78] Moumtzi, A.; Trenker, M.; Flicker, K.; Zenzmaier, E.; Saf, R.; Hermetter, A. Import and fate of fluorescent analogs of oxidized phospholipids in vascular smooth muscle cells. J. Lipid Res. 48:565–582; 2007. [79] Fruhwirth, G. O.; Moumtzi, A.; Loidl, A.; Ingolic, E.; Hermetter, A. The oxidized phospholipids POVPC and PGPC inhibit growth and induce apoptosis in vascular smooth muscle cells. Biochim. Biophys. Acta 1761:1060–1069; 2006. [80] Zettler, M. E.; Prociuk, M. A.; Austria, J. A.; Massaeli, H.; Zhong, G.; Pierce, G. N. OxLDL stimulates cell proliferation through a general induction of cell cycle proteins. Am. J. Physiol. Heart Circ. Physiol. 284:H644–H653; 2003. [81] Yu, S.; Wong, S. L.; Lau, C. W.; Huang, Y.; Yu, C. M. Oxidized LDL at low concentration promotes in-vitro angiogenesis and activates nitric oxide synthase through PI3K/Akt/eNOS pathway in human coronary artery endothelial cells. Biochem. Biophys. Res. Commun. 407:44–48; 2011. [82] Bedel, A.; Negre-Salvayre, A.; Heeneman, S.; Grazide, M. H.; Thiers, J. C.; Salvayre, R.; Maupas-Schwalm, F. E-cadherin/beta-catenin/T-cell factor pathway is involved in smooth muscle cell proliferation elicited by oxidized low-density lipoprotein. Circ. Res. 103:694–701; 2008. [83] Senokuchi, T.; Matsumura, T.; Sakai, M.; Matsuo, T.; Yano, M.; Kiritoshi, S.; Sonoda, K.; Kukidome, D.; Nishikawa, T.; Araki, E. Extracellular signalregulated kinase and p38 mitogen-activated protein kinase mediate macrophage proliferation induced by oxidized low-density lipoprotein. Atherosclerosis 176:233–245; 2004. [84] Mallat, Z.; Tedgui, A. Apoptosis in the vasculature: mechanisms and functional importance. Br. J. Pharmacol. 130:947–962; 2000. [85] Bennett, M. R.; Boyle, J. J. Apoptosis of vascular smooth muscle cells in atherosclerosis. Atherosclerosis 138:3–9; 1998. [86] Loidl, A.; Sevcsik, E.; Riesenhuber, G.; Deigner, H. P.; Hermetter, A. Oxidized phospholipids in minimally modified low density lipoprotein induce apoptotic signaling via activation of acid sphingomyelinase in arterial smooth muscle cells. J. Biol. Chem. 278:32921–32928; 2003. [87] Loidl, A.; Claus, R.; Ingolic, E.; Deigner, H. P.; Hermetter, A. Role of ceramide in activation of stress-associated MAP kinases by minimally modified LDL in vascular smooth muscle cells. Biochim. Biophys. Acta 1690:150–158; 2004. [88] Bu, D. X.; Tarrio, M.; Maganto-Garcia, E.; Stavrakis, G.; Tajima, G.; Lederer, J.; Jarolim, P.; Freeman, G. J.; Sharpe, A. H.; Lichtman, A. H. Impairment of the programmed cell death-1 pathway increases atherosclerotic lesion development and inflammation. Arterioscler. Thromb. Vasc. Biol. 31:1100–1107; 2011. [89] Qin, J.; Testai, F. D.; Dawson, S.; Kilkus, J.; Dawson, G. Oxidized phosphatidylcholine formation and action in oligodendrocytes. J. Neurochem. 110:1388–1399; 2009. [90] Clarke, M. C.; Littlewood, T. D.; Figg, N.; Maguire, J. J.; Davenport, A. P.; Goddard, M.; Bennett, M. R. Chronic apoptosis of vascular smooth muscle cells accelerates atherosclerosis and promotes calcification and medial degeneration. Circ. Res. 102:1529–1538; 2008. [91] Ait-Oufella, H.; Kinugawa, K.; Zoll, J.; Simon, T.; Boddaert, J.; Heeneman, S.; Blanc-Brude, O.; Barateau, V.; Potteaux, S.; Merval, R.; Esposito, B.; Teissier, E.; Daemen, M. J.; Leseche, G.; Boulanger, C.; Tedgui, A.; Mallat, Z. Lactadherin deficiency leads to apoptotic cell accumulation and accelerated atherosclerosis in mice. Circulation 115:2168–2177; 2007. [92] Clarke, M. C.; Talib, S.; Figg, N. L.; Bennett, M. R. Vascular smooth muscle cell apoptosis induces interleukin-1-directed inflammation: effects of hyperlipidemiamediated inhibition of phagocytosis. Circ. Res. 106:363–372; 2010.

[93] Chang, M. K.; Binder, C. J.; Miller, Y. I.; Subbanagounder, G.; Silverman, G. J.; Berliner, J. A.; Witztum, J. L. Apoptotic cells with oxidation-specific epitopes are immunogenic and proinflammatory. J. Exp. Med. 200:1359–1370; 2004. [94] Hsieh, C. C.; Yen, M. H.; Liu, H. W.; Lau, Y. T. Lysophosphatidylcholine induces apoptotic and non-apoptotic death in vascular smooth muscle cells: in comparison with oxidized LDL. Atherosclerosis 151:481–491; 2000. [95] Hardwick, S. J.; Hegyi, L.; Clare, K.; Law, N. S.; Carpenter, K. L.; Mitchinson, M. J.; Skepper, J. N. Apoptosis in human monocyte–macrophages exposed to oxidized low density lipoprotein. J. Pathol. 179:294–302; 1996. [96] Hundal, R. S.; Gomez-Munoz, A.; Kong, J. Y.; Salh, B. S.; Marotta, A.; Duronio, V.; Steinbrecher, U. P. Oxidized low density lipoprotein inhibits macrophage apoptosis by blocking ceramide generation, thereby maintaining protein kinase B activation and Bcl-XL levels. J. Biol. Chem. 278:24399–24408; 2003. [97] Yang, H.; Chen, S. C. The effect of interleukin-10 on apoptosis in macrophages stimulated by oxLDL. Eur. J. Pharmacol. 657:126–130; 2011. [98] Li, D.; Mehta, J. L. Upregulation of endothelial receptor for oxidized LDL (LOX-1) by oxidized LDL and implications in apoptosis of human coronary artery endothelial cells: evidence from use of antisense LOX-1 mRNA and chemical inhibitors. Arterioscler. Thromb. Vasc. Biol. 20:1116–1122; 2000. [99] Pirillo, A.; Reduzzi, A.; Ferri, N.; Kuhn, H.; Corsini, A.; Catapano, A. L. Upregulation of lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) by 15lipoxygenase-modified LDL in endothelial cells. Atherosclerosis 214:331–337; 2011. [100] Chen, J.; Mehta, J. L.; Haider, N.; Zhang, X.; Narula, J.; Li, D. Role of caspases in ox-LDL-induced apoptotic cascade in human coronary artery endothelial cells. Circ. Res. 94:370–376; 2004. [101] Nusshold, C.; Kollroser, M.; Kofeler, H.; Rechberger, G.; Reicher, H.; Ullen, A.; Bernhart, E.; Waltl, S.; Kratzer, I.; Hermetter, A.; Hackl, H.; Trajanoski, Z.; Hrzenjak, A.; Malle, E.; Sattler, W. Hypochlorite modification of sphingomyelin generates chlorinated lipid species that induce apoptosis and proteome alterations in dopaminergic PC12 neurons in vitro. Free Radic. Biol. Med. 48:1588–1600; 2010. [102] Dever, G.; Stewart, L. J.; Pitt, A. R.; Spickett, C. M. Phospholipid chlorohydrins cause ATP depletion and toxicity in human myeloid cells. FEBS Lett. 540: 245–250; 2003. [103] Dever, G.; Wainwright, C. L.; Kennedy, S.; Spickett, C. M. Fatty acid and phospholipid chlorohydrins cause cell stress and endothelial adhesion. Acta Biochim. Pol. 53:761–768; 2006. [104] Vissers, M. C.; Carr, A. C.; Winterbourn, C. C. Fatty acid chlorohydrins and bromohydrins are cytotoxic to human endothelial cells. Redox Rep. 6:49–55; 2001. [105] Gargalovic, P. S.; Imura, M.; Zhang, B.; Gharavi, N. M.; Clark, M. J.; Pagnon, J.; Yang, W. P.; He, A.; Truong, A.; Patel, S.; Nelson, S. F.; Horvath, S.; Berliner, J. A.; Kirchgessner, T. G.; Lusis, A. J. Identification of inflammatory gene modules based on variations of human endothelial cell responses to oxidized lipids. Proc. Natl. Acad. Sci. U. S. A. 103:12741–12746; 2006. [106] Cominacini, L.; Rigoni, A.; Pasini, A. F.; Garbin, U.; Davoli, A.; Campagnola, M.; Pastorino, A. M.; Lo Cascio, V.; Sawamura, T. The binding of oxidized low density lipoprotein (ox-LDL) to ox-LDL receptor-1 reduces the intracellular concentration of nitric oxide in endothelial cells through an increased production of superoxide. J. Biol. Chem. 276:13750–13755; 2001. [107] Beckman, J. S.; Koppenol, W. H. Nitric oxide, superoxide, and peroxynitrite: the good, the bad, and ugly. Am. J. Physiol. 271:C1424–C1437; 1996. [108] Lee, S.; Gharavi, N. M.; Honda, H.; Chang, I.; Kim, B.; Jen, N.; Li, R.; Zimman, A.; Berliner, J. A. A role for NADPH oxidase 4 in the activation of vascular endothelial cells by oxidized phospholipids. Free Radic. Biol. Med. 47:145–151; 2009. [109] Gharavi, N. M.; Baker, N. A.; Mouillesseaux, K. P.; Yeung, W.; Honda, H. M.; Hsieh, X.; Yeh, M.; Smart, E. J.; Berliner, J. A. Role of endothelial nitric oxide synthase in the regulation of SREBP activation by oxidized phospholipids. Circ. Res. 98:768–776; 2006. [110] Yeh, M.; Leitinger, N.; de Martin, R.; Onai, N.; Matsushima, K.; Vora, D. K.; Berliner, J. A.; Reddy, S. T. Increased transcription of IL-8 in endothelial cells is differentially regulated by TNF-alpha and oxidized phospholipids. Arterioscler. Thromb. Vasc. Biol. 21:1585–1591; 2001. [111] Yeh, M.; Cole, A. L.; Choi, J.; Liu, Y.; Tulchinsky, D.; Qiao, J. H.; Fishbein, M. C.; Dooley, A. N.; Hovnanian, T.; Mouilleseaux, K.; Vora, D. K.; Yang, W. P.; Gargalovic, P.; Kirchgessner, T.; Shyy, J. Y.; Berliner, J. A. Role for sterol regulatory elementbinding protein in activation of endothelial cells by phospholipid oxidation products. Circ. Res. 95:780–788; 2004. [112] Nuszkowski, A.; Grabner, R.; Marsche, G.; Unbehaun, A.; Malle, E.; Heller, R. Hypochlorite-modified low density lipoprotein inhibits nitric oxide synthesis in endothelial cells via an intracellular dislocalization of endothelial nitric-oxide synthase. J. Biol. Chem. 276:14212–14221; 2001. [113] Marsche, G.; Heller, R.; Fauler, G.; Kovacevic, A.; Nuszkowski, A.; Graier, W.; Sattler, W.; Malle, E. 2-Chlorohexadecanal derived from hypochlorite-modified highdensity lipoprotein-associated plasmalogen is a natural inhibitor of endothelial nitric oxide biosynthesis. Arterioscler. Thromb. Vasc. Biol. 24:2302–2306; 2004. [114] Kadl, A.; Galkina, E.; Leitinger, N. Induction of CCR2-dependent macrophage accumulation by oxidized phospholipids in the air-pouch model of inflammation. Arthritis Rheum. 60:1362–1371; 2009. [115] Springer, T. A. Adhesion receptors of the immune system. Nature 346:425–434; 1990. [116] Williams, M. R.; Azcutia, V.; Newton, G.; Alcaide, P.; Luscinskas, F. W. Emerging mechanisms of neutrophil recruitment across endothelium. Trends Immunol. 32: 461–469; 2011. [117] Alcaide, P.; Auerbach, S.; Luscinskas, F. W. Neutrophil recruitment under shear flow: it's all about endothelial cell rings and gaps. Microcirculation 16:43–57; 2009.

F.H. Greig et al. / Free Radical Biology & Medicine 52 (2012) 266–280 [118] Huber, J.; Furnkranz, A.; Bochkov, V. N.; Patricia, M. K.; Lee, H.; Hedrick, C. C.; Berliner, J. A.; Binder, B. R.; Leitinger, N. Specific monocyte adhesion to endothelial cells induced by oxidized phospholipids involves activation of cPLA2 and lipoxygenase. J. Lipid Res. 47:1054–1062; 2006. [119] Reddy, S.; Hama, S.; Grijalva, V.; Hassan, K.; Mottahedeh, R.; Hough, G.; Wadleigh, D. J.; Navab, M.; Fogelman, A. M. Mitogen-activated protein kinase phosphatase 1 activity is necessary for oxidized phospholipids to induce monocyte chemotactic activity in human aortic endothelial cells. J. Biol. Chem. 276:17030–17035; 2001. [120] Furnkranz, A.; Schober, A.; Bochkov, V. N.; Bashtrykov, P.; Kronke, G.; Kadl, A.; Binder, B. R.; Weber, C.; Leitinger, N. Oxidized phospholipids trigger atherogenic inflammation in murine arteries. Arterioscler. Thromb. Vasc. Biol. 25:633–638; 2005. [121] Watson, A. D.; Subbanagounder, G.; Welsbie, D. S.; Faull, K. F.; Navab, M.; Jung, M. E.; Fogelman, A. M.; Berliner, J. A. Structural identification of a novel proinflammatory epoxyisoprostane phospholipid in mildly oxidized low density lipoprotein. J. Biol. Chem. 274:24787–24798; 1999. [122] Subbanagounder, G.; Deng, Y.; Borromeo, C.; Dooley, A. N.; Berliner, J. A.; Salomon, R. G. Hydroxy alkenal phospholipids regulate inflammatory functions of endothelial cells. Vascul. Pharmacol. 38:201–209; 2002. [123] Leitinger, N.; Tyner, T. R.; Oslund, L.; Rizza, C.; Subbanagounder, G.; Lee, H.; Shih, P. T.; Mackman, N.; Tigyi, G.; Territo, M. C.; Berliner, J. A.; Vora, D. K. Structurally similar oxidized phospholipids differentially regulate endothelial binding of monocytes and neutrophils. Proc. Natl. Acad. Sci. U. S. A. 96:12010–12015; 1999. [124] Hsiai, T. K.; Cho, S. K.; Reddy, S.; Hama, S.; Navab, M.; Demer, L. L.; Honda, H. M.; Ho, C. M. Pulsatile flow regulates monocyte adhesion to oxidized lipid-induced endothelial cells. Arterioscler. Thromb. Vasc. Biol. 21:1770–1776; 2001. [125] Guo, L.; Chen, Z.; Cox, B. E.; Amarnath, V.; Epand, R. F.; Epand, R. M.; Davies, S. S. Phosphatidylethanolamines modified by gamma-ketoaldehyde (gammaKA) induce endoplasmic reticulum stress and endothelial activation. J. Biol. Chem. 286: 18170–18180; 2011. [126] Kiec-Wilk, B.; Polus, A.; Razny, U.; Cialowicz, U.; Dembinska-Kiec, A. Modulation of endothelial cell proliferation and capillary network formation by the ox-LDL component: 1-palmitoyl-2-archidonoyl-sn-glycero-3-phosphocholine (ox-PAPC). Genes Nutr. 6:347–351; 2011. [127] Pamukcu, B.; Lip, G. Y.; Devitt, A.; Griffiths, H.; Shantsila, E. The role of monocytes in atherosclerotic coronary artery disease. Ann. Med. 42:394–403; 2010. [128] Kim, C. S.; Kang, J. H.; Cho, H. R.; Blankenship, T. N.; Erickson, K. L.; Kawada, T.; Yu, R. Potential involvement of CCL23 in atherosclerotic lesion formation/progression by the enhancement of chemotaxis, adhesion molecule expression, and MMP-2 release from monocytes. Inflamm. Res. 60:889–895; 2011. [129] Klinke, A.; Nussbaum, C.; Kubala, L.; Friedrichs, K.; Rudolph, T. K.; Rudolph, V.; Paust, H. J.; Schroder, C.; Benten, D.; Lau, D.; Szocs, K.; Furtmuller, P. G.; Heeringa, P.; Sydow, K.; Duchstein, H. J.; Ehmke, H.; Schumacher, U.; Meinertz, T.; Sperandio, M.; Baldus, S. Myeloperoxidase attracts neutrophils by physical forces. Blood 117: 1350–1358; 2011. [130] Dever, G. J.; Benson, R.; Wainwright, C. L.; Kennedy, S.; Spickett, C. M. Phospholipid chlorohydrin induces leukocyte adhesion to ApoE−/− mouse arteries via upregulation of P-selectin. Free Radic. Biol. Med. 44:452–463; 2008. [131] Dever, G.; Spickett, C. M.; Kennedy, S.; Rush, C.; Tennant, G.; Monopoli, A.; Wainwright, C. L. The nitric oxide-donating pravastatin derivative, NCX 6550 [(1S-[1alpha(betaS*, deltaS*), 2alpha, 6alpha, 8beta-(R*), 8a alpha]]1,2,6,7,8,8a-hexahydro-beta, delta, 6-trihydroxy-2-methyl-8-(2-methyl-1oxobutoxy)-1-naphtalene-heptanoic acid 4-(nitrooxy)butyl ester)], reduces splenocyte adhesion and reactive oxygen species generation in normal and atherosclerotic mice. J. Pharmacol. Exp. Ther. 320:419–426; 2007. [132] Chen, J. W.; Zhou, S. B.; Tan, Z. M. Aspirin and pravastatin reduce lectin-like oxidized low density lipoprotein receptor-1 expression, adhesion molecules and oxidative stress in human coronary artery endothelial cells. Chin. Med. J. (English) 123:1553–1556; 2010. [133] Zimman, A.; Mouillesseaux, K. P.; Le, T.; Gharavi, N. M.; Ryvkin, A.; Graeber, T. G.; Chen, T. T.; Watson, A. D.; Berliner, J. A. Vascular endothelial growth factor receptor 2 plays a role in the activation of aortic endothelial cells by oxidized phospholipids. Arterioscler. Thromb. Vasc. Biol. 27:332–338; 2007. [134] Parhami, F.; Fang, Z. T.; Yang, B.; Fogelman, A. M.; Berliner, J. A. Stimulation of Gs and inhibition of Gi protein functions by minimally oxidized LDL. Arterioscler. Thromb. Vasc. Biol. 15:2019–2024; 1995. [135] Obinata, H.; Hattori, T.; Nakane, S.; Tatei, K.; Izumi, T. Identification of 9hydroxyoctadecadienoic acid and other oxidized free fatty acids as ligands of the G protein-coupled receptor G2A. J. Biol. Chem. 280:40676–40683; 2005. [136] Li, R.; Mouillesseaux, K. P.; Montoya, D.; Cruz, D.; Gharavi, N.; Dun, M.; Koroniak, L.; Berliner, J. A. Identification of prostaglandin E2 receptor subtype 2 as a receptor activated by OxPAPC. Circ. Res. 98:642–650; 2006. [137] Honda, Z.; Ishii, S.; Shimizu, T. Platelet-activating factor receptor. J. Biochem. 131: 773–779; 2002. [138] Marathe, G. K.; Prescott, S. M.; Zimmerman, G. A.; McIntyre, T. M. Oxidized LDL contains inflammatory PAF-like phospholipids. Trends Cardiovasc. Med. 11: 139–142; 2001. [139] Marathe, G. K.; Zimmerman, G. A.; Prescott, S. M.; McIntyre, T. M. Activation of vascular cells by PAF-like lipids in oxidized LDL. Vascul. Pharmacol. 38: 193–200; 2002. [140] Pegorier, S.; Stengel, D.; Durand, H.; Croset, M.; Ninio, E. Oxidized phospholipid: POVPC binds to platelet-activating-factor receptor on human macrophages: implications in atherosclerosis. Atherosclerosis 188:433–443; 2006. [141] Stafforini, D. M.; McIntyre, T. M.; Zimmerman, G. A.; Prescott, S. M. Platelet-activating factor, a pleiotrophic mediator of physiological and pathological processes. Crit. Rev. Clin. Lab. Sci. 40:643–672; 2003.

279

[142] Chen, R.; Chen, X.; Salomon, R. G.; McIntyre, T. M. Platelet activation by low concentrations of intact oxidized LDL particles involves the PAF receptor. Arterioscler. Thromb. Vasc. Biol. 29:363–371; 2009. [143] Heery, J. M.; Kozak, M.; Stafforini, D. M.; Jones, D. A.; Zimmerman, G. A.; McIntyre, T. M.; Prescott, S. M. Oxidatively modified LDL contains phospholipids with platelet-activating factor-like activity and stimulates the growth of smooth muscle cells. J. Clin. Invest. 96:2322–2330; 1995. [144] Leitinger, N.; Watson, A. D.; Faull, K. F.; Fogelman, A. M.; Berliner, J. A. Monocyte binding to endothelial cells induced by oxidized phospholipids present in minimally oxidized low density lipoprotein is inhibited by a platelet activating factor receptor antagonist. Adv. Exp. Med. Biol. 433:379–382; 1997. [145] Smiley, P. L.; Stremler, K. E.; Prescott, S. M.; Zimmerman, G. A.; McIntyre, T. M. Oxidatively fragmented phosphatidylcholines activate human neutrophils through the receptor for platelet-activating factor. J. Biol. Chem. 266:11104–11110; 1991. [146] Gopfert, M. S.; Siedler, F.; Siess, W.; Sellmayer, A. Structural identification of oxidized acyl-phosphatidylcholines that induce platelet activation. J. Vasc. Res. 42:120–132; 2005. [147] Suzuki, H.; Kurihara, Y.; Takeya, M.; Kamada, N.; Kataoka, M.; Jishage, K.; Ueda, O.; Sakaguchi, H.; Higashi, T.; Suzuki, T.; Takashima, Y.; Kawabe, Y.; Cynshi, O.; Wada, Y.; Honda, M.; Kurihara, H.; Aburatani, H.; Doi, T.; Matsumoto, A.; Azuma, S.; Noda, T.; Toyoda, Y.; Itakura, H.; Yazaki, Y.; Kodama, T., et al. A role for macrophage scavenger receptors in atherosclerosis and susceptibility to infection. Nature 386:292–296; 1997. [148] Febbraio, M.; Hajjar, D. P.; Silverstein, R. L. CD36: a class B scavenger receptor involved in angiogenesis, atherosclerosis, inflammation, and lipid metabolism. J. Clin. Invest. 108:785–791; 2001. [149] Endemann, G.; Stanton, L. W.; Madden, K. S.; Bryant, C. M.; White, R. T.; Protter, A. A. CD36 is a receptor for oxidized low density lipoprotein. J. Biol. Chem. 268: 11811–11816; 1993. [150] Yoshida, H.; Quehenberger, O.; Kondratenko, N.; Green, S.; Steinberg, D. Minimally oxidized low-density lipoprotein increases expression of scavenger receptor A, CD36, and macrosialin in resident mouse peritoneal macrophages. Arterioscler. Thromb. Vasc. Biol. 18:794–802; 1998. [151] Febbraio, M.; Podrez, E. A.; Smith, J. D.; Hajjar, D. P.; Hazen, S. L.; Hoff, H. F.; Sharma, K.; Silverstein, R. L. Targeted disruption of the class B scavenger receptor CD36 protects against atherosclerotic lesion development in mice. J. Clin. Invest. 105:1049–1056; 2000. [152] Kunjathoor, V. V.; Febbraio, M.; Podrez, E. A.; Moore, K. J.; Andersson, L.; Koehn, S.; Rhee, J. S.; Silverstein, R.; Hoff, H. F.; Freeman, M. W. Scavenger receptors class A-I/II and CD36 are the principal receptors responsible for the uptake of modified low density lipoprotein leading to lipid loading in macrophages. J. Biol. Chem. 277:49982–49988; 2002. [153] Podrez, E. A.; Poliakov, E.; Shen, Z.; Zhang, R.; Deng, Y.; Sun, M.; Finton, P. J.; Shan, L.; Febbraio, M.; Hajjar, D. P.; Silverstein, R. L.; Hoff, H. F.; Salomon, R. G.; Hazen, S. L. A novel family of atherogenic oxidized phospholipids promotes macrophage foam cell formation via the scavenger receptor CD36 and is enriched in atherosclerotic lesions. J. Biol. Chem. 277:38517–38523; 2002. [154] Greenberg, M. E.; Sun, M.; Zhang, R.; Febbraio, M.; Silverstein, R.; Hazen, S. L. Oxidized phosphatidylserine–CD36 interactions play an essential role in macrophage-dependent phagocytosis of apoptotic cells. J. Exp. Med. 203: 2613–2625; 2006. [155] Kopprasch, S.; Pietzsch, J.; Westendorf, T.; Kruse, H. J.; Grassler, J. The pivotal role of scavenger receptor CD36 and phagocyte-derived oxidants in oxidized low density lipoprotein-induced adhesion to endothelial cells. Int. J. Biochem. Cell Biol. 36:460–471; 2004. [156] Marsche, G.; Zimmermann, R.; Horiuchi, S.; Tandon, N. N.; Sattler, W.; Malle, E. Class B scavenger receptors CD36 and SR-BI are receptors for hypochloritemodified low density lipoprotein. J. Biol. Chem. 278:47562–47570; 2003. [157] Marsche, G.; Hammer, A.; Oskolkova, O.; Kozarsky, K. F.; Sattler, W.; Malle, E. Hypochlorite-modified high density lipoprotein, a high affinity ligand to scavenger receptor class B, type I, impairs high density lipoprotein-dependent selective lipid uptake and reverse cholesterol transport. J. Biol. Chem. 277:32172–32179; 2002. [158] Tugwood, J. D.; Issemann, I.; Anderson, R. G.; Bundell, K. R.; McPheat, W. L.; Green, S. The mouse peroxisome proliferator activated receptor recognizes a response element in the 5′ flanking sequence of the rat acyl CoA oxidase gene. EMBO J. 11:433–439; 1992. [159] Babaev, V. R.; Ishiguro, H.; Ding, L.; Yancey, P. G.; Dove, D. E.; Kovacs, W. J.; Semenkovich, C. F.; Fazio, S.; Linton, M. F. Macrophage expression of peroxisome proliferator-activated receptor-alpha reduces atherosclerosis in low-density lipoprotein receptor-deficient mice. Circulation 116:1404–1412; 2007. [160] Namgaladze, D.; Morbitzer, D.; von Knethen, A.; Brune, B. Phospholipase A2modified low-density lipoprotein activates macrophage peroxisome proliferatoractivated receptors. Arterioscler. Thromb. Vasc. Biol. 30:313–320; 2010. [161] Lee, H.; Shi, W.; Tontonoz, P.; Wang, S.; Subbanagounder, G.; Hedrick, C. C.; Hama, S.; Borromeo, C.; Evans, R. M.; Berliner, J. A.; Nagy, L. Role for peroxisome proliferator-activated receptor alpha in oxidized phospholipid-induced synthesis of monocyte chemotactic protein-1 and interleukin-8 by endothelial cells. Circ. Res. 87:516–521; 2000. [162] Taketa, K.; Matsumura, T.; Yano, M.; Ishii, N.; Senokuchi, T.; Motoshima, H.; Murata, Y.; Kim-Mitsuyama, S.; Kawada, T.; Itabe, H.; Takeya, M.; Nishikawa, T.; Tsuruzoe, K.; Araki, E. Oxidized low density lipoprotein activates peroxisome proliferator-activated receptor-alpha (PPARalpha) and PPARgamma through MAPK-dependent COX-2 expression in macrophages. J. Biol. Chem. 283: 9852–9862; 2008.

280

F.H. Greig et al. / Free Radical Biology & Medicine 52 (2012) 266–280

[163] Davies, S. S.; Pontsler, A. V.; Marathe, G. K.; Harrison, K. A.; Murphy, R. C.; Hinshaw, J. C.; Prestwich, G. D.; Hilaire, A. S.; Prescott, S. M.; Zimmerman, G. A.; McIntyre, T. M. Oxidized alkyl phospholipids are specific, high affinity peroxisome proliferatoractivated receptor gamma ligands and agonists. J. Biol. Chem. 276:16015–16023; 2001. [164] Barlic, J.; Zhang, Y.; Foley, J. F.; Murphy, P. M. Oxidized lipid-driven chemokine receptor switch, CCR2 to CX3CR1, mediates adhesion of human macrophages to coronary artery smooth muscle cells through a peroxisome proliferatoractivated receptor gamma-dependent pathway. Circulation 114:807–819; 2006. [165] Tontonoz, P.; Nagy, L.; Alvarez, J. G.; Thomazy, V. A.; Evans, R. M. PPARgamma promotes monocyte/macrophage differentiation and uptake of oxidized LDL. Cell 93:241–252; 1998. [166] Nagy, L.; Tontonoz, P.; Alvarez, J. G.; Chen, H.; Evans, R. M. Oxidized LDL regulates macrophage gene expression through ligand activation of PPARgamma. Cell 93:229–240; 1998. [167] Tobias, P.; Curtiss, L. K. The immune system and atherogenesis. Paying the price for pathogen protection: toll receptors in atherogenesis. J. Lipid Res. 46:404–411; 2005. [168] Miller, Y. I.; Viriyakosol, S.; Worrall, D. S.; Boullier, A.; Butler, S.; Witztum, J. L. Toll-like receptor 4-dependent and -independent cytokine secretion induced by minimally oxidized low-density lipoprotein in macrophages. Arterioscler. Thromb. Vasc. Biol. 25:1213–1219; 2005. [169] Walton, K. A.; Hsieh, X.; Gharavi, N.; Wang, S.; Wang, G.; Yeh, M.; Cole, A. L.; Berliner, J. A. Receptors involved in the oxidized 1-palmitoyl-2-arachidonoylsn-glycero-3-phosphorylcholine-mediated synthesis of interleukin-8: a role for Toll-like receptor 4 and a glycosylphosphatidylinositol-anchored protein. J. Biol. Chem. 278:29661–29666; 2003. [170] Verstrepen, L.; Bekaert, T.; Chau, T. L.; Tavernier, J.; Chariot, A.; Beyaert, R. TLR-4, IL-1R and TNF-R signaling to NF-kappaB: variations on a common theme. Cell. Mol. Life Sci. 65:2964–2978; 2008. [171] Erridge, C.; Webb, D. J.; Spickett, C. M. Toll-like receptor 4 signalling is neither sufficient nor required for oxidised phospholipid mediated induction of interleukin-8 expression. Atherosclerosis 193:77–85; 2007. [172] Erridge, C.; Kennedy, S.; Spickett, C. M.; Webb, D. J. Oxidized phospholipid inhibition of toll-like receptor (TLR) signaling is restricted to TLR2 and TLR4: roles for CD14, LPS-binding protein, and MD2 as targets for specificity of inhibition. J. Biol. Chem. 283:24748–24759; 2008. [173] Imai, Y.; Kuba, K.; Neely, G. G.; Yaghubian-Malhami, R.; Perkmann, T.; van Loo, G.; Ermolaeva, M.; Veldhuizen, R.; Leung, Y. H.; Wang, H.; Liu, H.; Sun, Y.; Pasparakis, M.; Kopf, M.; Mech, C.; Bavari, S.; Peiris, J. S.; Slutsky, A. S.; Akira, S.; Hultqvist, M.; Holmdahl, R.; Nicholls, J.; Jiang, C.; Binder, C. J.; Penninger, J. M. Identification of oxidative stress and Toll-like receptor 4 signaling as a key pathway of acute lung injury. Cell 133:235–249; 2008. [174] Seimon, T. A.; Nadolski, M. J.; Liao, X.; Magallon, J.; Nguyen, M.; Feric, N. T.; Koschinsky, M. L.; Harkewicz, R.; Witztum, J. L.; Tsimikas, S.; Golenbock, D.; Moore, K. J.; Tabas, I. Atherogenic lipids and lipoproteins trigger CD36-TLR2dependent apoptosis in macrophages undergoing endoplasmic reticulum stress. Cell Metab. 12:467–482; 2010. [175] Tseng, W.; Lu, J.; Bishop, G. A.; Watson, A. D.; Sage, A. P.; Demer, L.; Tintut, Y. Regulation of interleukin-6 expression in osteoblasts by oxidized phospholipids. J. Lipid Res. 51:1010–1016; 2010. [176] Bochkov, V. N.; Kadl, A.; Huber, J.; Gruber, F.; Binder, B. R.; Leitinger, N. Protective role of phospholipid oxidation products in endotoxin-induced tissue damage. Nature 419:77–81; 2002.

[177] von Schlieffen, E.; Oskolkova, O. V.; Schabbauer, G.; Gruber, F.; Bluml, S.; Genest, M.; Kadl, A.; Marsik, C.; Knapp, S.; Chow, J.; Leitinger, N.; Binder, B. R.; Bochkov, V. N. Multi-hit inhibition of circulating and cell-associated components of the toll-like receptor 4 pathway by oxidized phospholipids. Arterioscler. Thromb. Vasc. Biol. 29:356–362; 2009. [178] Bluml, S.; Kirchberger, S.; Bochkov, V. N.; Kronke, G.; Stuhlmeier, K.; Majdic, O.; Zlabinger, G. J.; Knapp, W.; Binder, B. R.; Stockl, J.; Leitinger, N. Oxidized phospholipids negatively regulate dendritic cell maturation induced by TLRs and CD40. J. Immunol. 175:501–508; 2005. [179] Ma, Z.; Li, J.; Yang, L.; Mu, Y.; Xie, W.; Pitt, B.; Li, S. Inhibition of LPS- and CpG DNA-induced TNF-alpha response by oxidized phospholipids. Am. J. Physiol. Lung Cell. Mol. Physiol. 286:L808–L816; 2004. [180] Oskolkova, O. V.; Afonyushkin, T.; Preinerstorfer, B.; Bicker, W.; von Schlieffen, E.; Hainzl, E.; Demyanets, S.; Schabbauer, G.; Lindner, W.; Tselepis, A. D.; Wojta, J.; Binder, B. R.; Bochkov, V. N. Oxidized phospholipids are more potent antagonists of lipopolysaccharide than inducers of inflammation. J. Immunol. 185:7706–7712; 2010. [181] Knapp, S.; Matt, U.; Leitinger, N.; van der Poll, T. Oxidized phospholipids inhibit phagocytosis and impair outcome in gram-negative sepsis in vivo. J. Immunol. 178:993–1001; 2007. [182] Birukov, K. G.; Bochkov, V. N.; Birukova, A. A.; Kawkitinarong, K.; Rios, A.; Leitner, A.; Verin, A. D.; Bokoch, G. M.; Leitinger, N.; Garcia, J. G. Epoxycyclopentenone-containing oxidized phospholipids restore endothelial barrier function via Cdc42 and Rac. Circ. Res. 95:892–901; 2004. [183] Nonas, S.; Miller, I.; Kawkitinarong, K.; Chatchavalvanich, S.; Gorshkova, I.; Bochkov, V. N.; Leitinger, N.; Natarajan, V.; Garcia, J. G.; Birukov, K. G. Oxidized phospholipids reduce vascular leak and inflammation in rat model of acute lung injury. Am. J. Respir. Crit. Care Med. 173:1130–1138; 2006. [184] Birukova, A. A.; Chatchavalvanich, S.; Oskolkova, O.; Bochkov, V. N.; Birukov, K. G. Signaling pathways involved in OxPAPC-induced pulmonary endothelial barrier protection. Microvasc. Res. 73:173–181; 2007. [185] Birukova, A. A.; Malyukova, I.; Mikaelyan, A.; Fu, P.; Birukov, K. G. Tiam1 and betaPIX mediate Rac-dependent endothelial barrier protective response to oxidized phospholipids. J. Cell. Physiol. 211:608–617; 2007. [186] Birukova, A. A.; Zebda, N.; Fu, P.; Poroyko, V.; Cokic, I.; Birukov, K. G. Association between adherens junctions and tight junctions via Rap1 promotes barrier protective effects of oxidized phospholipids. J. Cell. Physiol. 226:2052–2062; 2011. [187] Moellering, D. R.; Levonen, A. L.; Go, Y. M.; Patel, R. P.; Dickinson, D. A.; Forman, H. J.; Darley-Usmar, V. M. Induction of glutathione synthesis by oxidized low-density lipoprotein and 1-palmitoyl-2-arachidonyl phosphatidylcholine: protection against quinone-mediated oxidative stress. Biochem. J. 362:51–59; 2002. [188] Jyrkkanen, H. K.; Kansanen, E.; Inkala, M.; Kivela, A. M.; Hurttila, H.; Heinonen, S. E.; Goldsteins, G.; Jauhiainen, S.; Tiainen, S.; Makkonen, H.; Oskolkova, O.; Afonyushkin, T.; Koistinaho, J.; Yamamoto, M.; Bochkov, V. N.; Yla-Herttuala, S.; Levonen, A. L. Nrf2 regulates antioxidant gene expression evoked by oxidized phospholipids in endothelial cells and murine arteries in vivo. Circ. Res. 103:e1–e9; 2008. [189] Code, C.; Mahalka, A. K.; Bry, K.; Kinnunen, P. K. Activation of phospholipase A2 by 1-palmitoyl-2-(9′-oxo-nonanoyl)-sn-glycero-3-phosphocholine in vitro. Biochim. Biophys. Acta 1798:1593–1600; 2010. [190] Feige, E.; Mendel, I.; George, J.; Yacov, N.; Harats, D. Modified phospholipids as anti-inflammatory compounds. Curr. Opin. Lipidol. 21:525–529; 2010.