Peroxynitrite activates the cyclooxygenase activities of constitutive and inducible prostaglandin endoperoxide synthases by serving as a substrate for the enzymes' peroxidase activities. Activation of purified enzyme is induced by direct addition of peroxynitrite or by in situ generation of peroxynitrite from NO coupling to superoxide anion. Cu,Znsuperoxide dismutase completely inhibits cyclooxygenase activation in systems where peroxynitrite is generated in situ from superoxide. In the murine macrophage cell line RAW264.7, the lipophilic superoxide dismutase-mimetic agents, Cu(II) (3,5-diisopropylsalicylic acid) 2 , and Mn(III) tetrakis(1-methyl-4-pyridyl)porphyrin dose-dependently decrease the synthesis of prostaglandins without affecting the levels of NO synthase or prostaglandin endoperoxide synthase or by inhibiting the release of arachidonic acid. These findings support the hypothesis that peroxynitrite is an important modulator of cyclooxygenase activity in inf lammatory cells and establish that superoxide anion serves as a biochemical link between NO and prostaglandin biosynthesis.Prostaglandins and thromboxanes are important mediators of inflammation, hyperalgesia, cell growth, and hemostasis inter alia. The committed step in prostaglandin biosynthesis is the oxygenation of arachidonic acid (AA) by prostaglandin endoperoxide (PGH) synthase, a bifunctional, membrane-bound hemeprotein (1-4). Inhibition of the cyclooxygenase activity of PGH synthase is the basis for the pharmacological action of nonsteroidal antiinflammatory drugs (5, 6). Two different PGH synthases exist in vertebrates-PGH synthase-1, which is expressed constitutively and occurs in many tissues, and PGH synthase-2, which is inducible and expressed transiently (7-10). PGH synthase-2 is present at high levels in monocytes͞ macrophages, where it appears to play a major role in the production of inflammatory prostaglandins (7,11).Several recent reports demonstrate that NO stimulates prostaglandin biosynthesis in vivo, in perfused organs, and in macrophages (12-17). The stimulatory effect of NO is rapid and appears to be the result of direct activation of cyclooxygenase activity (16). However, conflicting reports exist regarding the ability of NO to stimulate purified PGH synthase, and it is possible that a derivative of NO is responsible for the activation in inflammatory cells (16,(18)(19)(20).The principal mechanism described for direct activation of the cyclooxygenase activity of PGH synthase is reaction of fatty acid hydroperoxides with the heme prosthetic group to generate a protein radical. This protein radical (probably Tyr-385) serves as the catalytic oxidant of AA (21-23). The identity of the hydroperoxides that activate PGH synthase in different cell types is uncertain because fatty acid hydroperoxides are excellent substrates for glutathione peroxidase (GSH-Px)-catalyzed reduction by glutathione (GSH) (24). The potential for control of prostaglandin biosynthesis by GSH-Px͞GSH reduction of hydroperoxide activators exhibits s...

Studies of oxidative damage during the progression of Alzheimer’s disease (AD) suggest its central role in disease pathogenesis. To investigate levels of nucleic acid oxidation in both early and late stages of AD, levels of multiple base adducts were quantified in nuclear and mitochondrial DNA from the superior and middle temporal gyri (SMTG), inferior parietal lobule (IPL), and cerebellum (CER) of age-matched normal control subjects, subjects with mild cognitive impairment, preclinical AD, late-stage AD, and non-AD neurological disorders (diseased control; DC) using gas chromatography/mass spectrometry. Median levels of multiple DNA adducts in nuclear and mitochondrial DNA were significantly (P ≤ 0.05) elevated in the SMTG, IPL, and CER in multiple stages of AD and in DC subjects. Elevated levels of fapyguanine and fapyadenine in mitochondrial DNA suggest a hypoxic environment early in the progression of AD and in DC subjects. Overall, these data suggest that oxidative damage is an early event not only in the pathogenesis of AD, but is also present in neurodegenerative diseases in general.

Clostridium thermocellum is a candidate organism for consolidated bioprocessing of lignocellulosic biomass into ethanol. However, commercial use is limited due to growth inhibition at modest ethanol concentrations. Recently, an ethanol-adapted strain of C. thermocellum was produced. Since ethanol adaptation in microorganisms has been linked to modification of membrane lipids, we tested the hypothesis that ethanol adaptation in C. thermocellum involves lipid modification by comparing the fatty acid composition and membrane anisotropy of wild-type and ethanol-adapted strains. Derivatization to fatty acid methyl esters provided quantitative lipid analysis. Compared to wild-type, the ethanol-adapted strain had a larger percentage of fatty acids with chain lengths >16:0 and showed a significant increase in the percentage of 16:0 plasmalogens. Structural identification of fatty acids was confirmed through mass spectral fragmentation patterns of picolinyl esters. Ethanol adaptation did not involve modification at sites of methyl branching or the unsaturation index. Comparison of steady-state fluorescence anisotropy experiments, in the absence and presence of ethanol, provided evidence for the effects of ethanol on membrane fluidity. In the presence of ethanol, both strains displayed increased fluidity by approximately 12%. These data support the model that ethanol adaptation was the result of fatty acid changes that increased membrane rigidity that counter-acted the fluidizing effect of ethanol.

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