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The extracellular matrix (ECM) is critical to biological architecture and determines cellular properties, function and activity. In many situations it is highly abundant, with collagens and elastin being some of the most abundant proteins in mammals. The ECM comprises of multiple different protein species and sugar polymers, with both different isoforms and post-translational modifications (PTMs) providing a large variety of microenvironments that play a key role in determining tissue structure and health. A number of the PTMs (e.g. cross-links) present in the ECM are critical to integrity and function, whereas others are deleterious to both ECM structure and associated cells. Modifications induced by reactive oxidants and electrophiles have been reported to accumulate in some ECM with increasing age. This accumulation can be exacerbated by disease, and in particular those associated with acute or chronic inflammation, obesity and diabetes. This is likely to be due to higher fluxes of modifying agents in these conditions. In this focused review, the role and effects of oxidants and other electrophiles on ECM are discussed, with a particular focus on the artery wall and atherosclerotic cardiovascular disease. Modifications generated on ECM components are reviewed, together with the effects of these species on cellular properties including adhesion, proliferation, migration, viability, metabolic activity, gene expression and phenotype. Increasing data indicates that ECM modifications are both prevalent in human and mammalian tissues and play an important role in disease development and progression. [ABSTRACT FROM AUTHOR]

The pentose phosphate pathway (PPP) is a key metabolic pathway. The oxidative phase of this process involves three reactions catalyzed by glucose-6-phosphate dehydrogenase (G6PDH), 6-phosphogluconolactonase (6PGL) and 6-phosphogluconate dehydrogenase (6PGDH) enzymes. The first and third steps (catalyzed by G6PDH and 6PGDH, respectively) are responsible for generating reduced nicotinamide adenine dinucleotide phosphate (NAPDH), a key cofactor for maintaining the reducing power of cells and detoxification of both endogenous and exogenous oxidants and electrophiles. Despite the importance of these enzymes, little attention has been paid to the fact that these proteins are targets of oxidants. In response to oxidative stimuli metabolic pathways are modulated, with the PPP often up-regulated in order to enhance or maintain the reductive capacity of cells. Under such circumstances, oxidation and inactivation of the PPP enzymes could be detrimental. Damage to the PPP enzymes may result in a downward spiral, as depending on the extent and sites of modification, these alterations may result in a loss of enzymatic activity and therefore increased oxidative damage due to NADPH depletion. In recent years, it has become evident that the three enzymes of the oxidative phase of the PPP have different susceptibilities to inactivation on exposure to different oxidants. In this review, we discuss existing knowledge on the role that these enzymes play in the metabolism of cells, and their susceptibility to oxidation and inactivation with special emphasis on NADPH production. Perspectives on achieving a better understanding of the molecular basis of the oxidation these enzymes within cellular environments are given. [ABSTRACT FROM AUTHOR]

Smokers have an elevated risk of cardiovascular disease but the origin(s) of this increased risk are incompletely defined. Considerable evidence supports an accumulation of the oxidantgenerating enzyme MPO (myeloperoxidase) in the inflamed artery wall, and smokers have high levels of SCN-, a preferred MPO substrate, with this resulting in HOSCN (hypothiocyanous acid) formation. We hypothesized that this thiol-specific oxidant may target the Zn2+ -thiol cluster of eNOS (endothelial nitric oxide synthase), resulting in enzyme dysfunction and reduced formation of the critical signallingmoleculeNO·.DecreasedNO· bioavailability is an early and critical event in atherogenesis, and HOSCN-mediated damage to eNOS may contribute to smokingassociated disease. In the present study it is shown that exposure of isolated eNOS to HOSCN or MPO/H2O2/SCN- decreased active dimeric eNOS levels, and increased inactive monomer and Zn2+ release, compared with controls, HOCl (hypochlorous acid)- or MPO/H2O2/Cl- -treated samples. eNOS activity was increasingly compromised byMPO/H2O2/Cl- with increasing SCN- concentrations. Exposure ofHCAEC (human coronary artery endothelial cell) lysates to pre-formed HOSCN, or MPO/H2O2/Cl- with increasing SCN-, increased eNOS monomerization and Zn2+ release, and decreased activity. Intact HCAECs exposed to HOCl and HOSCN had decreased eNOS activity and NO2- /NO3- formation (products of NO· decomposition), and increased free Zn2+ . Exposure of isolated rat aortic rings to HOSCN resulted in thiol loss, and decreased eNOS activity and cGMP levels. Overall these data indicate that high SCN- levels, as seen in smokers, can increase HOSCN formation and enhance eNOS dysfunction in human endothelial cells, with this potentially contributing to increased atherogenesis in smokers. [ABSTRACT FROM AUTHOR]

Oxidative modification of HDLs (high-density lipoproteins) by MPO (myeloperoxidase) compromises its anti-atherogenic properties, which may contribute to the development of atherosclerosis. Although it has been established that HOCl (hypochlorous acid) produced by MPO targets apoA-I (apolipoprotein A-I), the major apolipoprotein of HDLs, the role of the other major oxidant generated by MPO, HOSCN (hypothiocyanous acid), in the generation of dysfunctional HDLs has not been examined. In the present study, we characterize the structural and functional modifications of lipid-free apoA-I and rHDL (reconstituted discoidal HDL) containing apoA-I complexed with phospholipid, induced by HOSCN and its decomposition product, OCN- (cyanate). Treatment of apoA-I with HOSCN resulted in the oxidation of tryptophan residues, whereas OCN- induced carbamylation of lysine residues to yield homocitrulline. Tryptophan residues were more readily oxidized on apoA-I contained in rHDLs. Exposure of lipid-free apoA-I to HOSCNandOCN- significantly reduced the extent of cholesterol efflux from cholesterol-loaded macrophages when compared with unmodified apoA-I. In contrast, HOSCN did not affect the antiinflammatory properties of rHDL. The ability of HOSCN to impair apoA-I-mediated cholesterol efflux may contribute to the development of atherosclerosis, particularly in smokers who have high plasma levels of SCN- (thiocyanate). [ABSTRACT FROM AUTHOR]