F2-Isoprostanes are prostaglandin (PG) isomers formed in situ in cell membranes by peroxidation of arachidonic acid. 8-epi PGF2alpha and IPF2alpha-I are F2-isoprostanes produced in humans which circulate in plasma and are excreted in urine. Measurement of F2-isoprostanes may offer a sensitive, specific, and noninvasive method for measuring oxidant stress in clinical settings where reactive oxygen species are putatively involved. We determined whether isoprostanes were present in human atherosclerotic lesions, where lipid peroxidation is thought to occur in vivo. 8-epi PGF2alpha ranged from 1.310-3.450 pmol/micromol phospholipid in atherectomy specimens compared with 0.045-0.115 pmol/micromol phospholipid (P < 0.001) in vascular tissue devoid of atherosclerosis. Corresponding values of IPF2alpha-I were 5.6-13.8 vs. 0.16-0.44 pmol/micromol phospholipid (P < 0.001). Levels of the two isoprostanes in vascular tissue were highly correlated (r = 0.80, P < 0.0001). Immunohistochemical studies confirmed that foam cells adjacent to the lipid necrotic core of the plaque were markedly positive for 8-epi PGF2alpha. These cells were also reactive with anti-CD68, an epitope specific for human monocyte/macrophages. 8-epi PGF2alpha immunoreactivity was also detected in cells positive for anti-alpha-smooth muscle actin antibody, which specifically recognizes vascular smooth muscle cells. Our results indicate that 8-epi PGF2alpha and IPF2alpha-I, two distinct F2-isoprostanes and markers of oxidative stress in vivo, are present in human atherosclerotic plaque. Quantitation of these chemically stable products of lipid peroxidation in target tissues, as well as in biological fluids, may aid in the rational development of antioxidant drugs in humans.

Isoprostanes are prostaglandin isomers produced from arachidonic acid by a free radical-catalyzed mechanism. Urinary excretion of 8- iso -prostaglandin F _2α , an isomer of the PGG/H synthase (cyclooxygenase or COX) enzyme product, prostaglandin F _2α (PGF _2α ), has exhibited promise as an index of oxidant stress in vivo. We have developed a quantitative method to measure isoprostane F _2α -I, (IPF _2α -I) a class I isomer (8- iso -PGF _2α is class IV), using gas chromatography/mass spectrometry. IPF _2α -I is severalfold as abundant in human urine as 8- iso- PGF _2α , with mean values of 737 ± 20.6 pg/mg creatinine. Both isoprostanes are formed in a free radical-dependent manner in low density lipoprotein oxidized by copper in vitro . However, IPF _2α -I, unlike 8- iso -PGF _2α , is not formed in a COX-dependent manner by platelets activated by thrombin or collagen in vitro . Similarly, COX inhibition in vivo has no effect on IPF _2α -I. Neither serum IPF _2α -I, an index of cellular capacity to generate the isoprostane, nor urinary excretion of IPF _2α -I, an index of actual generation in vivo , is depressed by aspirin or indomethacin. In contrast, both serum thromboxane B ₂ and urinary excretion of its 11-dehydro metabolite are depressed by the COX inhibitors. Although serum 8- iso- PGF _2α formation is substantially depressed by COX inhibitors, urinary excretion of the compound is unaffected. Urinary IPF _2α -I is elevated in cigarette smokers compared with controls (1525 ± 180 versus 740 ± 40 pg/mg creatinine; P < 0.01) and is highly correlated with urinary 8- iso -PGF _2α ( r = 0.9; P < 0.001). Urinary IPF _2α -I is a novel index of lipid peroxidation in vivo , which can be measured with precision and sensitivity. It is an abundant F ₂ -isoprostane formed in a free radical- but not COX-dependent manner. Although 8- iso -PGF _2α may be formed as a minor product of COX, this pathway contributes trivially, if at all, to levels in urine. Urinary excretion of both isoprostanes is elevated in cigarette smokers.

This article reviews our current understanding of the role of oxygen free radicals in platelet activation. Several studies have indicated that platelets, in analogy to other circulating blood cells, are able to produce oxygen free radicals, which are likely to play an important role in the mechanism of platelet activation and aggregation. Platelet activation has been obtained with very low, physiologically relevant concentrations of radicals generated chemically, by leukocytes, and by hemoglobin derived from membrane leakage of erythrocytes. Knowledge of the role of reactive species in platelet physiology is relevant because platelets are brought into close contact with other cells capable of producing free radicals, such as neutrophils, macrophages, and endothelial cells, during the formation of thrombus. The physiopatological importance of these findings is high because it is now emerging that free radicals may have a role in the mechanism of atherosclerosis and its thrombotic complications, where the causative role of platelets is well documented. This background suggests therapeutic interventions with antioxidants as antiplatelet agents to improve the pharmacological effect of classical antiplatelet drug such as aspirin.