Determinants of endothelial function in a cohort of patients with peripheral artery disease.

OBJECTIVES: Endothelial dysfunction assessed by brachial artery flow-mediated dilatation (FMD) has been associated with cardiovascular events. There have been relatively few studies examining FMD or other measures of endothelial function in patients with peripheral artery disease (PAD). The aim of this study was to examine determinants of FMD in a homogenous cohort of patients with PAD. METHODS: We prospectively assessed patients presenting with life style-limiting intermittent claudication to establish the presence of cardiovascular risk factors, obesity and metabolic syndrome. Fasting serum was assayed for lipids, C-reactive protein, adiponectin, leptin, resistin and osteoprotegerin (OPG). FMD was measured by high-resolution ultrasound. RESULTS: Serum concentrations of OPG were elevated in patients with obesity and metabolic syndrome. FMD was impaired in patients with obesity and metabolic syndrome and negatively correlated with serum concentrations of OPG. By multiple regression analysis, metabolic syndrome was independently associated with impaired FMD after adjustment for age, smoking, ischaemic heart disease, cerebrovascular disease and severity of PAD. CONCLUSIONS: Our findings suggest that metabolic syndrome is an important determinant of endothelial function in patients with PAD, and OPG may be a useful biomarker of this effect.

Inhibition of human glutathione S-transferases by basic triphenylmethane dyes.

Human glutathione S-transferases (GSTs) of the Alpha-, Mu- and Pi- classes were expressed in E. coli and isolated by affinity chromatography. They were tested for their susceptibility to inhibition by basic triphenylmethane dyes. hGSTA 1-1 was inhibited by Malachite Green with a Ki value of the order of 10 microM. The inhibitory species appeared to be the dye-GSH adduct. This isoenzyme was not inhibited by either Crystal Violet or Ethyl Violet at concentrations up to 50 microM. hGSTM 2-2 was weakly inhibited by all three dyes tested with Ki values being in the range 40-80 microM. For all dyes the inhibition was best characterised as non-competitive. hGSTP 1-1 was not inhibited by Crystal Violet or by Ethyl Violet but was strongly inhibited by Malachite Green (Ki = 0.3 microM). The mode of inhibition appeared to be non-competitive but it seems probable that the mechanism is complex. There is at present no evidence to show clearly whether the dominant inhibitory species is the free dye or the adduct.

Inhibition of glutathione S-transferases from rat liver by basic triphenylmethane dyes.

(1) Basic triphenylmethane dyes related to pararosanilin inhibit class alpha glutathione S-transferases (GSTs) from rat liver. The inhibitory potency of each dye correlates with its octanol-water partition coefficient. Values of Ki determined at pH 6.5 ranged from about 1 x 10(-7) M for Ethyl violet to 7 x 10(-5) M for Methyl green. GST 3-3, a class mu isoenzyme, was an order of magnitude less sensitive to inhibition by Ethyl violet. (2) All of the dyes tested were bleached to varying degrees by glutathione. The bleaching appears to result from the formation of an adduct between the dye and glutathione. At pH 6.5, adduct formation is significant only for Malachite green and Methyl green. There is kinetic evidence that for these dyes the adduct contributes significantly to the overall inhibition. It is probable that at physiological pH, all of the dyes would exist to a significant extent in the adduct form. (3) The dyes are excreted extensively in the bile, at least partly as the glutathione adduct. The free dye is regenerated on standing, it is assumed as a result of removal of glutathione by oxidation.