Risk factors for atherosclerosis - can they be used to identify the patient with multisystem atherosclerosis?

Risk factors are often used in preventive care programmes to identify the patient at particular risk for developing atherosclerosis. Risk factors for atherosclerosis have also been shown to be linked to the presence of the disease at a given time, a fact that may be helpful when screening for additional atherosclerotic disease in the known arteriopath. Risk factors .were recorded in 471 patients admi"ed to hospital with symptoms of atherosclerosis. In patients admi"ed primarily with peripheral vascular disease, risk factors linked to the presence of additional coronary artery disease were a family history of ischaemic heart disease (odds ratio = 2,6), the presence of carotid artery disease (odds ratio = 1,9) and high fasting serum triglyceride levels (P < 0,04). Grouping these factors together usin.g logistic regression, ischaemic heart disease could be predicted with a sensitivity of 72% and a specificity of 43%. Patients admitted with carotid artery disease were more likely to have ischaemic heart disease in the presence of peripheral vascular disease (odds ratio = 1,9) and a raised serum cholesterol level (P < 0,02), while female gender (odds ratio = 2,9) and an increase in age (P< 0,001) were linked to an increased prevalence of concomitant atherosclerosis in patients admmed with acute myocardial infarction or for elective coronary artery bypass surgery. Using an age cut-off point, additional atherosclerosis could be predicted with a sensitivity of 32% and a specificity of 88% in these patients

Apparent hydroxyl radical generation without transition metal catalysis and tyrosine nitration during oxidation of the anti-tubercular drug, isonicotinic acid hydrazide

Aromatic hydroxylation and formation of thiobarbituric acid-reactive substances occurred in a mixture of isonicotinic acid hydrazide (isoniazid) and catalase. Since these reactions were stimulated by phytic acid (a potent metal chelator), rather than inhibited, transition metal-catalysed hydroxyl radical generation was not implicated. Hydroxylation also occurred with isoniazid and phytic acid in the absence of catalase, albeit to a lesser extent. The independent effects of catalase and phytic acid are related to their abilities to catalyse isoniazid oxidation. In the presence of tyrosine, both the isoniazid/phytic acid system and authentic peroxynitrite generated dityrosine. Authentic peroxynitrite, as well as a phytic acid-mediated isoniazid oxidation product, have absorbance maxima at 302 nm. The yield of this isoniazid-derived product increased with pi-I and in the presence of a superoxide-generating system. A good correlation existed between absorbance at 302 nm and aromatic hydroxylation. Acid-induced decomposition of the 302 nm absorbance in the presence of superoxide dismutase led to the formation of a product absorbing in the same region as peroxynitrite-modified superoxide dismutase (350 nm at acid pH). Catalase catalysed peroxynitrite-mediated, as well as isoniazid/phytic acid-mediated tyrosine nitration, which was accompanied by Compound II formation (ferryl-catalase) in both cases. We postulate that peroxynitrite or a similar species is formed during isoniazid oxidation.Articl

Cloning and expression of the α-L-arabinofuranosidase gene (ABF2) of Aspergillus niger in Saccharomyces cerevisiae

First-strand cDNA was prepared from mRNA of Aspergillus niger MRC11624 induced on oat spelts xylan. Using the cDNA as a template, the α-L-arabinofuranosidase gene (abfB) was amplified with the polymerase chain reaction technique. The abfB DNA fragment was inserted between the yeast phosphoglycerate kinase I gene promoter (PGK1(P)) and terminator (PGK1(T)) sequences on a multicopy episomal plasmid. The resulting construct PGK1(P)abfB-PGK1(T) was designated ABF2. The ABF2 gene was expressed successfully in Saccharomyces cerevisiae and functional α-L-arabinofuranosidase was secreted from the yeast cells. The ABF2 nucleotide sequence was determined and verified to encode a 449-amino-acid protein (Abf2) that is 9.4% identical to the α-L-arabinofuranosidase B of A. niger N400. Maximum α-L-arabinofuranosidase activities of 0.020 U/ml and 1.40 U/ml were obtained with autoselective recombinant S. cerevisiae strains when grown for 48 h in synthetic and complex medium respectively.Articl

Purification and properties of two phospholipase A2 enzymes from berg adder (Bitis atropos) venom

Two phospholipases A2 (PLA2-1 and PLA2-2) of berg adder (Bitis atropos) venom were purified by successive chromatography on Cellex-CM, Sephadex G-75 and Bio-Gel P-100 columns. PLA2-1 is an acidic PLA2 (pl 6.6) with a molecular weight of 29 393.12 as determined by electrospray ionization mass-spectrometry (ESI-MS). When treated with a reducing agent, it split into two chains of about 15.7 kDa and 13 kDa, as revealed by sodium dodecyl sulphate-polycrylamide gel electrophoresis. Although PLA2-1 displayed cytotoxic activity in vitro on Be-11 melanoma cells (EC50=76.6 μg ml-1), no toxic effects were noted in mice. The molecular weight of PLA2-2 as determined with ESI-MS is 14 034.89. This enzyme, a basic monomeric PLA2 (pl 9.3), induced neurotoxic symptoms in mice. An intra-peritoneal LD50 of 1.7 mg kg-1 was calculated. Its N-terminal 19 amino acid sequence was Asn-Leu-Tyr-Gln-Phe-Gly-Lys-Met-Ile-Ser-His-Lys-Thr-Asn-Asn-Gly-Pro-Leu-Ala. This is the first report of the isolation of a dimeric as well as monomeric PLA2 enzyme from the venom of Bitis atropos.Articl

Aromatic hydroxylation during the myeloperoxidase-oxidase oxidation of hydrazines

Benzoic acid was found to be hydroxylated by a mixture of myeloperoxidase (MPO) and the mycobactericidal drug, isoniazid. Aromatic hydroxylation and formation of compound III (oxyperoxidase) were coincident during the MPO-oxidase oxidation of isoniazid which proceeded without augmentation from the reagent hydrogen peroxide. An intermediate of isoniazid reduced ferric MPO to ferrous MPO which associated with dioxygen to form compound III. Aromatic hydroxylation also occurred in a mixture of isoniazid (or phenylhydrazine) and a ferric salt. Hydroxylations in both the enzymatic and nonenzymatic reaction systems were inhibited by the iron chelator, desferal, as well as by the specific hydroxyl radical scavenger, mannitol. To distinguish between the hydroxylating intermediates in the different reaction systems, the unique properties of the natural antioxidant, phytic acid, were exploited. Phytic acid inhibited aromatic hydroxylation in the Fe3+-INH system, which is in accordance with its known properties as a powerful inhibitor of iron-driven reactions (·OH formation). By contrast, phytic acid stimulated hydroxylation in the enzymatic system which was accompanied by a concomitant stimulation in the rate of compound III formation. These events were, however, not directly related to each other. Phytic acid had a direct effect on the redox transformation of isoniazid by stimulating superoxide generation during auto-oxidation of the drug. In addition, phytic acid also facilitated compound III decay in the absence of isoniazid, suggesting that it may also regulate the oxygen affinity of MPO, similar to its effect on the oxygenation of haemoglobin. The data on aromatic hydroxylation in the MPO-isoniazid system do not support a role for ·OH in the reaction and may fit the model for the P450 mixed oxidase system.Articl

Anti-oxidant properties of H2-receptor antagonists. Effects on myeloperoxidase-catalysed reactions and hydroxyl radical generation in a ferrous-hydrogen peroxide system

Ulcerogenesis of the gastroduodenal mucosa is caused by the digestive action of gastric juice and initially involves an inflammatory reaction with infiltration of phagocytes. The anti-inflammatory activity of many drugs have been attributed to the inhibition of the leukocyte enzyme, myeloperoxidase (MPO). In this study, the H2-antagonists in clinical use were found to be potent inhibitors of MPO-catalysed reactions (IC50 < 3 μM) under conditions resembling those in experiments with intact neutrophils. Since peak plasma concentrations of cimetidine, ranitidine and nizatidine are well within the micromolar range, after oral therapeutic dosing, our results may be of clinical relevance. The inhibitory actions of cimetidine and nizatidine were largely due to scavenging of hypochlorous acid (HOCl), a powerful chlorinating oxidant produced in the MPO-H2O2-Cl- system. In contrast to famotidine, ranitidine was also a potent scavenger of HOCl, while both drugs inhibited MPO reversibly by converting it to compound II, which is inactive in the oxidation of Cl-. The HOCl scavenging potencies of ranitidine and nizatidine were about three times higher than that of the anti-rheumatic drug, penicillamine, which had a potency similar to that of cimetidine. The rapid HOCl scavenging ability of penicillamine is thought to contribute to its anti-inflammatory effects. Using riboflavin as a probe, the H2-antagonists were found to be inhibitors of hydroxyl radical (·OH) generated in a Fe2+-H2O2 reaction mixture. Spectral analyses of the interaction of iron ions with the drugs and studies with chelators, suggest that the drugs were efficient chelators of Fe2+, in addition to their ·OH scavenging abilities. Since the gastrointestinal tract can contain potentially reactive iron, the simultaneous presence of H2-antagonists may help to suppress iron-driven steps in tissue damage.Articl