No abstract

Considerable evidence exists for oxidative damage to extracellular materials during multiple human pathologies. Unlike cells, the extracellular compartment of most biological tissues is less well protected against oxidation than intracellular sites in terms of the presence of both antioxidants (low molecular mass and enzymatic) and repair enzymes. The extracellular compartment may therefore be subject to greater oxidative stress, marked alterations in redox balance and an accumulation of damage due to slow turnover and/or poor repair. The nature and consequences of damage to ECM (extracellular matrix) are poorly understood, despite the growing realization that changes in matrix structure not only have structural consequences, but also play a key role in the regulation of cellular adhesion, proliferation, migration and cell signalling. The ECM also plays a key role in cytokine and growth factor binding, and matrix modifications would therefore be expected to alter these parameters. In the present study, we review mechanisms of oxidative damage to ECM, resulting changes in matrix structure and how this affects cellular behaviour. The role of such damage in the development and progression of inflammatory diseases is also discussed with particular reference to cardiovascular disease.

No abstract

The presence of D D-amino-acid-containing polypeptides, defensin-like peptide (DLP)-2 and Ornithorhyncus venom C-type natriuretic peptide (OvCNP)b, in platypus venom suggested the existence of a mammalian D D-amino-acid-residue isomerase(s) responsible for the modification of the all-L L-amino acid precursors. We show here that this enzyme(s) is present in the venom gland extract and is responsible for the creation of DLP-2 from DLP-4 and OvCNPb from OvCNPa. The isomerisation reaction is freely reversible and under well defined laboratory conditions catalyses the interconversion of the DLPs to full equilibration. The isomerase is $50-60 kDa and is inhibited by methanol and the peptidase inhibitor amastatin. This is the first known L L-to-D D-amino-acid-residue isomerase in a mammal.

The heparan sulfate (HS) proteoglycan perlecan is a major component of basement membranes, plays a key role in extracellular matrix (ECM) structure, interacts with growth factors and adhesion molecules, and regulates the adhesion, differentiation and proliferation of vascular cells. Atherosclerosis is characterized by chronic inflammation and the presence of oxidized materials within lesions, with the majority of protein damage present on ECM, rather than cell, proteins. Weakening of ECM structure plays a key role in lesion rupture, the major cause of heart attacks and strokes. In this study peroxynitrite, a putative lesion oxidant, is shown to damage perlecan structurally and functionally. Exposure of human perlecan to peroxynitrite decreases recognition by antibodies raised against both the core protein and heparan sulfate chains; dose-dependent formation of 3-nitrotyrosine was also detected. These effects were modulated by bicarbonate and reaction pH. Oxidant exposure resulted in aggregate formation, consistent with oxidative protein crosslinking. Peroxynitrite treatment modified functional properties of perlecan that are dependent on both the protein core (decreased binding of human coronary artery endothelial cells), and the HS chains (diminished fibroblast growth factor-2 (FGF-2) receptor-mediated proliferation of Baf-32 cells). The latter is consistent with a decrease in FGF-2 binding to the HS chains of modified perlecan. Immunofluorescence of advanced human atherosclerotic lesions provided evidence for the presence of perlecan and extensive formation of 3-nitrotyrosine epitopes within the intimal region; these materials showing marked co-localization. These data indicate that peroxynitrite induces major structural and functional changes to perlecan and that damage to this material occurs within human atherosclerotic lesions.