Numerous studies have reported the presence of oxidatively modified high-density lipoprotein (OxHDL) within the intima of atheromatous plaques as well as in plasma; however, its role in the pathogenesis of thrombotic disease is not established. We now report that OxHDL, but not native HDL, is a potent inhibitor of platelet activation and aggregation induced by physiologic agonists. This antithrombotic effect was concentration and time dependent and positively correlated with the degree of lipoprotein oxidation. Oxidized lipoproteins are known ligands for scavenger receptors type B, CD36 and scavenger receptor B type I (SR-BI), both of which are expressed on platelets. Studies using murine CD36(-/-) or SR-BI(-/-) platelets demonstrated that the antithrombotic activity of OxHDL depends on platelet SR-BI but not CD36. Binding to SR-BI was required since preincubation of human and murine platelets with anti-SR-BI blocking antibody abrogated the inhibitory effect of OxHDL. Agonist-induced aggregation of platelets from endothelial nitric oxide synthase (eNOS)(-/-), Akt-1(-/-), and Akt-2(-/-) mice was inhibited by OxHDL to the same degree as platelets from wild-type (WT) mice, indicating that the OxHDL effect is mediated by a pathway different from the eNOS/Akt pathway. These novel findings suggest that contrary to the prothrombotic activity of oxidized low-density lipoprotein (OxLDL), HDL upon oxidation acquires antithrombotic activity that depends on platelet SR-BI.

Recent studies have identified a novel family of oxidized phosphatidylcholines (oxPC CD36 ) that serve as highly specific ligands for scavenger receptor CD36. oxPC CD36 accumulate in vivo and mediate macrophage foam cell formation as well as promote platelet hyper-reactivity in hyperlipidemia via CD36. The structural basis of oxPC CD36 binding to CD36 has not been elucidated. We used liquid-phase binding to glutathione S-transferase fusion proteins containing various regions of CD36 to initially identify the region spanning CD36 amino acids 157-171 to contain a major binding site for oxPC CD36 CD36 is a 472-amino-acid, 88-kDa heavily glycosylated transmembrane protein that is expressed in various cell types including macrophages, platelets, microvascular endothelial cells, and adipocytes (1, 2). CD36 has been shown to play a significant role in a number of physiological and pathological processes in vivo including atherogenesis, lipid sensing and metabolism, innate immune responses, angiogenesis, uptake of apoptotic cells, and diabetes (2-4). CD36 involvement in such a variety of processes can be partially explained by its capacity to recognize a number of various distinct ligands. Examples of CD36 ligands include thrombospondin-1 (5), oxidized low density lipoproteins (oxLDL) 2 (6), oxidized phospholipids (7-9), fatty acids (10), microbial diacylglycerides (4), hexarelin (3), collagen (11), and malarial parasite-infected erythrocytes (12).That CD36 can function as a multiligand receptor is conceivable assuming that it has multiple ligand binding domains. Several studies suggest, for example, that the binding sites of thrombospondin-1 and oxLDL on CD36 are different (13,14). Two distinct binding sites are proposed for oxLDL on CD36. Studies using a monoclonal antibody have shown that domain 155-183 of CD36 plays a critical role in the binding of LDL oxidized by copper (15). Solid phase binding assays using recombinant fusion proteins spanning various regions of CD36, however, implicate the domain 28 -93 as the major binding site for oxLDL (16).We have recently identified a novel family of oxidized choline glycerophospholipids (oxPC CD36 ) that mediate CD36-dependent recognition of LDL oxidized by various pathways. The structural aspect of oxPC CD36 essential for high affinity binding to CD36 is an sn-2 acyl group that incorporates a terminal ␥-hydroxy(or oxo)-␣,␤-unsaturated carbonyl. A characteristic feature of oxPC CD36 conformation is a negatively charged distal end of the sn-2 acyl chain residue that partitions into the aqueous phase (17). oxPC CD36 are formed during the oxidation of LDL by multiple distinct pathways, serve as specific high affinity ligands for CD36 (9), and are present in vivo at sites of enhanced oxidative stress (18 -20). OxPC CD36 mediate foam cell formation induced by oxidized LDL via macrophage CD36 and induce a prothrombotic phenotype in hyperlipidemia via platelet CD36 (18,20).In this current study, we investigated the structural basis for the recognition of oxPC CD36 by CD36 using...

NF-B is known to exert its antiviral innate immune response via the IFN-␤-induced Janus kinase͞signal transducers and activators of transcription pathway. However, our current studies have demonstrated that activated NF-B is capable of directly establishing an antiviral state independent of IFN or secreted soluble factor(s) against two highly pathogenic respiratory RNA viruses. Human parainfluenza virus type 3, a mildly cytopathic virus that induced NF-B very early during infection was converted to a virulent virus after NF-B inhibition. In contrast, a highly cytopathic virus, human respiratory syncytial virus that induced NF-B late during infection, was converted to a mildly cytopathic virus after NF-B induction before virus replication. This interconversion of cytopathic phenotypes of viruses after NF-B modulation was further shown to be independent of IFN and soluble secreted factors(s). Moreover, tumor necrosis factor ␣ (TNF-␣) and IL-1␤ elicited an antiviral response, which was NF-B-dependent. Thus, NF-B induction directly confers an essential innate antiviral response against human parainfluenza virus type 3 and respiratory syncytial virus, which is independent of IFN-inducible factor(s).

Background: Infection of intestinal epithelial cells by pathogenic Salmonella leads to activation of signaling cascades that ultimately initiate the proinflammatory gene program. The transcription factor NF-κB is a key regulator/activator of this gene program and is potently activated. We explored the mechanism by which Salmonella activates NF-κB during infection of cultured intestinal epithelial cells and found that flagellin produced by the bacteria and contained on them leads to NF-κB activation in all the cells; invasion of cells by the bacteria is not required to activate NF-κB.

Specific oxidized phospholipids (oxPC CD36 ) accumulate in vivo at sites of oxidative stress and serve as high affinity ligands for scavenger receptors class B (CD36 and SR-BI). Recognition of oxPC CD36 by scavenger receptors plays a role in several pathophysiological processes. The structural basis for the recognition of oxPC CD36 by CD36 and SR-BI is poorly understood. A characteristic feature of oxPC CD36 is an sn-2 acyl group that incorporates a terminal ␥-hydroxy (or oxo)-␣,␤-unsaturated carbonyl. In the present study, a series of model oxidized phospholipids were designed, synthesized, and tested for their ability to serve as ligands for CD36 and SR-BI. We demonstrated that intact the sn-1 hydrophobic chain, the sn-3 hydrophilic phosphocholine or phosphatidic acid group, and the polar sn-2 tail are absolutely essential for high affinity binding. We further found that a terminal negatively charged carboxylate at the sn-2 position suffices to generate high binding affinity to class B scavenger receptors. In addition, factors such as polarity, rigidity, optimal chain length of sn-2, and sn-3 positions and negative charge at the sn-3 position of phospholipids further modulate the binding affinity. We conclude that all three positions of oxidized phospholipids are essential for the effective recognition by scavenger receptors class B. Furthermore, the structure of residues in these positions controls the affinity of the binding. The present studies suggest that, in addition to oxPC CD36 , other oxidized phospholipids observed in vivo may represent novel ligands for scavenger receptors class B. Specific oxidized phospholipids (oxPC CD36 )3 accumulate at sites of oxidative stress in vivo such as within atherosclerotic lesions and plasma in dyslipidemia (1, 2). They serve as high affinity ligands for scavenger receptors class B: CD36 and SR-BI (3, 4). Recognition of oxPC CD36 on the surface of cell membranes and lipoprotein particles by scavenger receptors class B plays an important role in several pathophysiological processes, including atherosclerosis and thrombosis. oxPC CD36 phospholipids mediate uptake of oxidized low density lipoprotein (oxLDL) by macrophages via CD36 and promote a pro-thrombotic state via platelet scavenger receptor CD36 (1, 2). oxPC CD36 phospholipids also prevent binding of high density lipoprotein by SR-BI because of the close proximity of the binding sites for these two ligands on SR-BI. Furthermore, oxPC CD36 interfere with SR-BI-mediated selective uptake of cholesteryl esters in hepatocytes (4). These data demonstrate that oxidative stress and accumulation of specific oxidized phospholipids may have a detrimental effect due to specific interaction with scavenger receptors class B. However, the exact molecular mechanism of the recognition of oxPC CD36 by scavenger receptors class B is poorly understood.Initial studies have demonstrated that the sn-2 acyl group of oxPC CD36 that incorporates a terminal ␥-hydroxy (or oxo)-␣,␤-unsaturated carbonyl is essential for high affinity binding to...