Increased platelet responsiveness following coronary stenting. Heparin as a possible aetiological factor in stent thrombosis.

AIMS: Platelet activation may be a determinant of thrombotic and restenotic complications following intracoronary stenting. In order to measure the effect of stenting on platelet activation antigen expression we used whole blood flow cytometry in 18 patients undergoing Palmaz-Schatz stenting (treated with full anticoagulation) and compared these with a group of 18 patients undergoing elective angioplasty. The effects of low molecular weight heparin and unfractionated heparin on platelet behaviour were also studied, both in vitro and in vivo to determine the contribution of prolonged heparin therapy to platelet activation following stenting. METHODS AND RESULTS: Fibrinogen binding to activated GPIIb-IIIa, and surface expression of P-selectin, GPIb and GPIIb-IIIa antigens were measured in unstimulated peripheral blood samples (rest) and on stimulation with adenosine diphosphate (0.1-10 micromol x 1(-1)) and thrombin (0.02-0.16 U x ml(-1)). No changes were seen in resting samples following angioplasty or stenting. Agonist responsiveness was unaltered after angioplasty, but in stented patients antigen expression in response to thrombin was significantly reduced (P< or =0.04), whilst the adenosine diphosphate response was significantly increased (P=0.01). Similar effects were observed in patients with unstable angina treated with either low molecular weight heparin or unfractionated heparin in vivo. In vitro, both unfractionated and low molecular weight heparin inhibited thrombin-induced platelet activation, but stimulation of adenosine diphosphate responses was more marked with unfractionated than low molecular weight heparin. CONCLUSIONS: There was a significant increase in platelet responsiveness to adenosine diphosphate following intracoronary stenting in patients treated with conventional anticoagulants. This was probably a consequence of treatment with heparin. Activation of platelets by heparin may explain the increased rate of stent thrombosis in patients treated with anticoagulant therapy. Low molecular weight heparins stimulate platelets less than unfractionated heparin.

Altered platelet function detected by flow cytometry. Effects of coronary artery disease and age.

Platelet activation state and responsiveness to physiological agonists were measured in 65 patients with documented coronary artery disease (54 male and 11 female; mean age, 58 years). Twelve patients (mean age, 52 years), selected at random from the male cohort, were compared with 12 age-matched male control subjects (mean age, 52 years) and with 10 normal, young male subjects (mean age, 25 years). Whole-blood flow cytometry was used to measure platelet activation status ex vivo and platelet responsiveness to physiological agonists in vitro. Peripheral blood samples were analyzed for bound fibrinogen and expression of P-selectin, GPIb, and GPIIb-IIIa at rest and in response to ADP (0.1 to 10 mumol/L) and thrombin (0.02 to 0.32 mu/mL). No significant differences were seen in the basal levels of fibrinogen binding between any of the groups, but P-selectin expression was significantly lower in patients compared with age-matched control subjects (P = .0005). When stimulated with agonists, patients' platelets had significantly decreased fibrinogen binding (P < .03) but no difference in P-selectin expression compared with the age-matched group. Both agonist-induced fibrinogen binding and P-selectin expression were, however, higher in the young subjects compared with either the older control group or the patients (P < .05). GPIb and GPIIb-IIIa expression were lowest in the patients with angina and highest in the young control subjects, with levels in the age-matched control subjects falling between these values. Data from the total patient cohort (n = 65) were identical to those in the smaller cohort (n = 12). In conclusion, atherosclerosis impairs platelet aggregatory responses (fibrinogen binding) over and above the decreased response seen with age. Platelet degranulation (P-selectin expression) is also impaired in patients with coronary artery disease, but only in comparison with younger subjects, not age-matched controls.

Contribution of phenols, quinones and reactive oxygen species to the mutagenicity of white grape juice in the Ames test.

The purpose of this study was to evaluate the role of phenols, quinones and reactive oxygen species in the mutagenicity of white grape juice in the Ames mutagenicity test. Mutagenicity was markedly suppressed by reduced glutathione but was not influenced by superoxide dismutase or catalase. In the presence of grape polyphenol oxidase, the mutagenicity of grape juice was markedly increased. When hepatic cytosol from Aroclor 1254-induced rats, supplemented with a reduced nicotinamide adenine dinucleotide phosphate-generating system. served as an activation system, an increase in the mutagenicity of grape juice was observed. The cytosol-induced mutagenicity of grape juice was attenuated in the presence of superoxide dismutase, catalase and glutathione. It is concluded that polyphenol oxidase-catalysed oxidation of phenolic compounds generates genotoxic species that are, at least partly, responsible for the mutagenicity of grape juice. In the presence of hepatic cytosol, one-electron reduction of grape juice quinones leads to the production of reactive oxygen species resulting in an increase in the mutagenic response.

Mutagenicity of white grape juice in the Ames test.

The mutagenicity of commercially available white grape juice was evaluated in the Ames mutagenicity test. Grape juice elicited a positive mutagenic, response in Salmonella typhimurium strain TA104 and a weaker response in strains TA97, TA98, TA100 and TA1530. The mutagenic response was evident in the absence of an activation system and inclusion of such a system did not influence mutagenicity. The grape juice-mediated mutagenic response was not due to histidine residues in the juice or likely treatment with sulfite. Moreover, freshly prepared grape juice displayed a similar mutagenic response. Three different brands of commercially available white grape juice were investigated in the Ames test; they all provoked a clear positive mutagenic response, but the degree of mutagenicity differed and could not be attributed to differences in the content of solids. It is concluded that grapes contain direct-acting genotoxic component(s).

Antimutagenicity of three isomers of aminobenzoic acid in Salmonella typhimurium.

The m-, o- and p-isomers of aminobenzoic acid (ABA) repressed the mutagenicity of N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) in Salmonella typhimurium TA100. Their antimutagenic potency was in the order of o-ABA > m-ABA > p-ABA. The mechanism of this antimutagenicity is ascribed mainly to the decomposition of MNNG induced by the aminobenzoic acid isomers outside or within the bacterial cells. The inhibition of plant cell peroxidases and bacterial acetyltransferases that are required for the plant activation of 2-aminofluorene (2-AF) to mutagenic product(s) may participate in the repression of 2-AF mutagenesis by the aminobenzoic acids in S. typhimurium strain YG1024. The aminobenzoic acid isomers exhibited no inhibitory effects towards the direct-acting agent 2-acetoxy-2-acetylaminofluorene, the stable diacetylated metabolic product of 2-AF.