The generation and actions of isoprostanes.
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Animals↗
Biomedical subjects
L J Roberts
Publications and source records attributed to L J Roberts.
Induction of prostaglandin endoperoxide synthase-1 (COX-1) in a human promonocytic cell line by treatment with the differentiating agent TPA.
Prostaglandin endoperoxide synthase (PGH synthase) is responsible for converting arachdonic acid to PGH2, the common precursor of prostaglandins. It has been shown previously that phorbol ester-induced differentiation of human promonocytic leukemia cell lines is accompanied by induction of PGH synthase enzyme and enhanced capacity to produce prostaglandins. However, the identity of the isoform of PGH synthase, i.e., PGH synthase-1 or -2, that is induced under these conditions has not been established. Northern and Western analyses revealed a dramatic increase in levels of PGH synthase-1 mRNA and protein levels within 24 hr after treatment of THP-1 cells with phorbol ester. No significant increase in PGH synthase-2 mRNA or protein was observed. The increases in PGH synthase-1 were accompanied by an enhanced capacity of the cells to produce PGE2. The current findings indicate that expression of PGH synthase-1 is greatly enhanced in a promonocytic cell line by treatment with an agent that induces differentiation.
Calcium↗
A nomenclature system for the isoprostanes.
In 1990, prostaglandin (PG) F2-like compounds were discovered to be produced in abundance in vivo by a free radical mechanism independent of the cyclooxygenase enzyme. Because these compounds are isomeric to cyclooxygenase-derived PGF2 alpha, they were termed F2-isoprostanes (F2-IsoP's). Subsequently, it was also demonstrated that PGD2-like compounds (D2-IsoP's) and PGE2-like compounds (E2-IsoP's) are also produced in vivo as products of this pathway. Four different regioisomers of each of these classes of IsoP's are formed, each of which can be comprised of eight racemic diastereomers. Thus, 64 different F2-IsoP's, E2-IsoP's, and D2-IsoP's can be formed. Interest in these molecules stems not only from the fact that quantification of IsoP's can provide a valuable index of free radical-induced lipid peroxidation in vivo but also from the fact that it has been shown that these compounds are capable of exerting potent biological activity. Because of this potential for exerting biological activity, the chemical syntheses of various IsoP compounds for biological testing has been initiated. As a result, a need for a systematic nomenclature for these compounds has evolved. A facile nomenclature that will allow rational differentiation and designation of each of the isomeric structures comprising the family of IsoP's is presented.
Dinoprost↗
The isoprostanes: unique bioactive products of lipid peroxidation.
The discovery of IsoPs as products of non-enzymatic lipid peroxidation has opened up new areas of investigation regarding the role of free radicals in human physiology and pathophysiology. The quantification of IsoPs as markers of oxidative stress status appears to be an important advance in our ability to explore the role of free radicals in the pathogenesis of human disease. A drawback related to this, however, has been lack of more facile and less expensive methods than mass spectrometry for the measurement of IsoPs. On the other hand, the recent introduction of immunoassay methods for measurement of IsoPs may alleviate this problem, provided they are specific and reliable. If this is the case, immunoassay methodology will most likely lead to an expansion of the use of measurements of IsoPs to assess oxidative stress status in vivo. Another need in the field of free radical medicine is information regarding the clinical pharmacology of antioxidant agents. Because of the evidence implicating free radicals in the pathogenesis of a number of human diseases, large clinical trials are planned or underway to assess whether antioxidants can either prevent the development or ameliorate the pathology of certain human disorders. However, data regarding the most effective doses and combination of antioxidant agents to use in these clinical trials is lacking. As mentioned previously, administration of antioxidants suppresses the formation of IsoPs, even in normal individuals. Thus, measurement of IsoPs may provide a valuable approach to defining the clinical pharmacology of antioxidants. In addition to being markers of oxidative stress, at least two IsoPs possess potent biological activity. The availability of additional IsoPs in synthetic form should broaden our knowledge concerning the role of these molecules as mediators of oxidant stress. Moreover, information regarding the nature of the receptor(s) that mediate the biological actions of IsoPs will be of considerable importance to the development of specific antagonists or agonists of the biological actions of IsoPs. Despite the fact that considerable information has been obtained since the initial report of the discovery of IsoPs, much remains to be understood about these molecules. With continued research in this area, we believe that much new information will emerge that will open up additional important new areas for future investigation.
Biomarkers↗
Identification and relative quantitation of F2-isoprostane regioisomers formed in vivo in the rat.
F2-isoprostanes are a complex mixture of isomers formed in four regioisomeric family types by free radical-initiated oxidation of arachidonic acid present in membrane phospholipids. F2-isoprostanes isolated from the livers of rats treated with carbon tetrachloride were separated by initial reverse phase HPLC and detected using electrospray ionization mass spectrometry with the characteristic loss of 44 u (C2H4O) from the common 1,3-diol cyclopropane ring found in these eicosanoids. Collision induced decomposition of the carboxylate anions from the separated F2-isoprostanes formed abundant ions characteristic for regioisomers of Type I (m/z 115), Type III (m/z 127), and Type IV (m/z 193), which made possible characterization of these three family subtypes by LC/MS/MS. Capillary GC/MS was employed to further identify the F2-isoprostane regioisomers using electron ionization mass spectrometry and to obtain characteristic mass spectra of the pentafluorobenzyl ester trimethylsilyl ether derivatives. Quantitation of the F2-isoprostanes separated by both reverse-phase HPLC and capillary GC/MS was carried out using negative ion chemical ionization mass spectrometry. The most abundant isomers identified were Type I and IV regioisomers constituting 33 and 25% of the total products, respectively. As expected, the Type II and III regioisomer products were of less abundance. Over 45 F2-isoprostanes could be separated in this complex mixture, suggesting random production of each regioisomeric subtype in this in vivo model.
Animals↗
Vascular superoxide dismutase deficiency impairs endothelial vasodilator function through direct inactivation of nitric oxide and increased lipid peroxidation.
Nitric oxide (NO) and superoxide are both constitutive products of the endothelium. Because NO is readily inactivated by superoxide, the bioactivity of endothelium-derived NO (EDNO) is dependent on local activity of superoxide dismutase (SOD). We examined the effects of chronic inhibition of copper-zinc SOD (CuZnSOD) using a rat model of dietary copper restriction. Male weanling Sprague-Dawley rats were fed a Cu-deficient diet and received either no Cu replacement (Cu-deficient) or Cu in the drinking water (Cu-sufficient). Compared with Cu-sufficient animals, Cu-deficiency was associated with a 68% reduction in CuZnSOD activity and a 58% increase in vascular superoxide as estimated by lucigenin chemiluminescence (both P < .05). Compared with Cu-sufficient animals, arterial relaxation in the thoracic aorta from Cu-deficient animals was 10-fold less sensitive to acetylcholine, a receptor-dependent EDNO agonist, but only 1.5-fold less sensitive to A23187, a receptor-independent EDNO agonist, and only 1.25-fold less sensitive to authentic NO (all P < .05). In contrast, acute inhibition of CuZnSOD with 10 mM diethyldithiocarbamate produced a more uniform reduction in sensitivity to acetylcholine (8-fold), A23187 (10-fold), and NO (4-fold; all P < .001). Cu-deficient animals demonstrated a 2.5-fold increase in plasma-esterified F2-isoprostanes, a stable marker of lipid peroxidation, that correlated inversely with arterial relaxation to acetylcholine (R = -.83; P < .0009) but not A23187 or authentic NO. From these findings, we conclude that chronic inhibition of CuZnSOD inhibits EDNO-mediated arterial relaxation through two mechanisms, one being direct inactivation of NO and the other being lipid peroxidation that preferentially interrupts receptor-mediated stimulation of EDNO.
Acetylcholine↗
Prostaglandin F2-like compounds, F2-isoprostanes, are present in increased amounts in human atherosclerotic lesions.
Oxidative modification of LDL is believed to play a major role in atherogenesis. As major lipid peroxidation products oxygenated linoleic acid derivatives and oxysterols have been described in human atherosclerotic lesions. Here we report that human lesions contain isoprostanes as peroxidation products of arachidonic acid at a level of 27.1 +/- 21.2 pg/mg wet weight (n = 10), which corresponds to 75.9 +/- 59.3 pg/mg dry weight, n contrast, human umbilical veins (n = 10), which were used as nonatherosclerotic control vessels, contain much smaller amounts of isoprostanes (1.4 +/- 0.7 pg/mg wet weight, which corresponds to 11.7 +/- 6.2 pg/mg dry weight), and there are significant differences between the two types of vessels. As major products of linoleic acid oxidation, racemic hydroxy linoleate isomers were detected in the lesional ester lipids. In human lesions, the hydroxy linoleic acid/linoleic acid ratio was about 0.5%, a result indicating that 5 out of 1000 linoleate residues are present as hydroxylated derivatives. In umbilical veins, no hydroxy linoleic acid could be detected. These data show that human atherosclerotic lesions contain increased amounts of hydroxy linoleic acid isomers and isoprostanes when compared with nonatherosclerotic vessel wall and suggest a link between local lipid peroxidation and progression of atherosclerosis. For evaluation of the degree of lipid peroxidation, the determination of the hydroxy linoleic acid/linoleic acid ratio appears to be more suitable than the isoprostane content.
Aged↗
Monetary reinforcement of abstinence from cocaine among mentally ill patients with cocaine dependence.
OBJECTIVE: The study investigated whether contingency management could reduce cocaine use by patients with schizophrenia. METHODS: An A-B-A research design, with two-month baseline, intervention, and follow-up phases, was used to study two homeless, treatment-resistant male outpatients with DSM-III-R diagnoses of schizophrenia and cocaine dependence. During the intervention phase, subjects provided daily urine specimens for testing for the cocaine metabolite benzoylecgonine (BE) and received $25 for each negative test. Concentrations of BE and metabolites of other illicit drugs were assayed twice a week to determine the amount of drug use in addition to frequency. Analysis of variance was used to compare drug use during the three study phases. RESULTS: During the intervention, the proportion of tests positive for cocaine was lower for both subjects. Mean urinary concentrations of BE were significantly lower during the intervention than during the baseline. CONCLUSIONS: These results suggest that modest monetary reinforcement of abstinence may decrease cocaine use among cocaine-dependent patients with schizophrenia.
Adult↗
Progressive myoclonus epilepsy in young adults with neuropathologic features of Alzheimer's disease.
Progressive myoclonus epilepsy (PME) may develop in adult life. We present two patients with PME appearing around the age of 30 years in whom the disorder represented a manifestation of Alzheimer's disease. This diagnosis must be considered in addition to possible Kufs' disease or myoclonic epilepsy with ragged red fibers (MERRF) when PME develops in young adults.
Adult↗
Nonenzymatic free radical-catalyzed generation of thromboxane-like compounds (isothromboxanes) in vivo.
The isoprostanes (IsoPs) are novel bioactive prostaglandin-like compounds produced in vivo by free radical-catalyzed peroxidation of arachidonyl-containing lipids. Previously, we have identified IsoPs containing F-type and D- and E-type prostane rings that are formed by reduction and rearrangement of IsoP endoperoxide intermediates, respectively. We now explore whether thromboxane B2 (TxB2)-like compounds, termed B2-isothromboxanes (B2-IsoTxs), are formed by rearrangement of IsoP endoperoxides. Detection of these compounds was carried out using a stable isotope dilution mass spectrometric assay originally developed for quantification of cyclooxygenase-derived TxB2. Incubations of arachidonic acid with Fe/ADP/ascorbate for 30 min in vitro generated a series of peaks representing putative B2-IsoTx at levels of 62.4 +/- 21.0 ng/mg arachidonate. Using various chemical modification and derivatization approaches, it was determined that these compounds contained hemiacetal ring structures and two double bonds, as would be expected for B2-IsoTx. Analysis of the compounds by electron ionization mass spectrometry yielded multiple mass spectra similar to those of TxB2. B2-IsoTxs are also formed esterified to phospholipids; oxidation of arachidonyl-containing phosphatidylcholine in vitro followed by hydrolysis resulted in the release of large amounts of these compounds. To explore whether B2-IsoTxs are also formed in vivo, a well characterized animal model of lipid peroxidation consisting of orogastric administration of CCl4 to rats was used. Levels of B2-IsoTx esterified in lipids in the liver increased 41-fold from 2.5 +/- 0.5 to 102 +/- 30 ng/g of liver. In addition, circulating levels of free compounds increased from undetectable (<5 pg/ml) to 185 +/- 30 pg/ml after CCl4, a 37-fold increase. Thus, we have provided evidence that IsoTxs constitute another novel class of eicosanoids produced in vivo nonenzymatically by free radical-catalyzed lipid peroxidation. These studies thus expand our understanding of products of lipid peroxidation formed in vivo from the free radical-catalyzed peroxidation of arachidonic acid.
Animals↗
Identification of the major urinary metabolite of the F2-isoprostane 8-iso-prostaglandin F2alpha in humans.
F2-isoprostanes are prostaglandin-like products of nonenzymatic lipid peroxidation. Measurement of levels of endogenous unmetabolized F2-isoprostanes has proven to be a valuable approach to assess oxidative stress in vivo. However, measurement of levels of urinary metabolites of F2-isoprostanes in timed urine collections offers an advantage over measuring unmetabolized F2-isoprostanes, e. g. in a plasma sample, in that it can provide an integrated index of isoprostane production over time. Therefore, we sought to identify the major urinary metabolite in humans of one of the more abundant F2-isoprostanes produced, 8-iso-prostaglandin F2alpha (8-iso-PGF2alpha). 20 microCi of tritiated 8-iso-PGF2alpha was infused over 1 h into a male volunteer. 75% of the infused radioactivity was excreted into the urine during the following 4.5 h and was combined with urine collected for 4 h from a rhesus monkey following infusion of 500 microg of unlabeled 8-iso-PGF2alpha. Urinary metabolites were isolated and purified by adsorption chromatography and high pressure liquid chromatography. The major urinary metabolite, representing 29% of the total extractable recovered radioactivity in the urine, was structurally identified by gas chromatography and mass spectrometry as 2,3-dinor-5, 6-dihydro-8-iso-prostaglandin F2alpha. The identification of 2, 3-dinor-5,6-dihydro-prostaglandin F2alpha as the major urinary metabolite of 8-iso-prostaglandin F2alpha provides the basis for the development of methods of assay for its quantification as a means to obtain an integrated assessment of oxidative stress status in humans.
Animals↗
Diisopropylphosphorofluoridate-induced muscle hyperactivity associated with enhanced lipid peroxidation in vivo.
Acute exposure to acetylcholinesterase (AChE) inhibitors such as organophosphates and carbamates induces functional changes at the neuromuscular junctions, leading to fasciculations that ultimately cause muscle fiber necrosis. There is recent evidence that oxygen free radical formation may be a factor in the toxicity of these insecticides. One of the targets of free radical-induced injury is lipid peroxidation. The role of lipid peroxidation in diisopropylphosphorofluoridate (DFP)-induced muscle necrosis was investigated by quantifying two products resulting from the oxidation of lipids in muscle tissue-the thiobarbituric acid-malondialdehyde complex (TBA-MDA) and F2-isoprostanes, the latter being a novel and extremely accurate marker of lipid peroxidation in vivo. When compared with control animals, significant increases in MDA of 96% and in F2-isoprostanes of 56% were found in the diaphragms of rats treated with 2.0 mg/kg DFP after 60 min (P < 0.01). In rats pretreated with the neuromuscular blocking agent d-tubocurarine or the lazaroid U-78517F, an antioxidant, no DFP-induced increases in either MDA or F2-isoprostanes were observed. It is suggested that the AChE inhibitor-induced cholinergic hyperactivity initiates that accumulation of free radicals leading to lipid peroxidation, which may be the initiator of the AChE inhibitor-induced cell injury.
Animals↗
Plasma prostaglandin E1 concentrations and hemodynamics during intravenous infusions of prostaglandin E1 in humans and swine.
Prostaglandin (PG) E1 administered intravenously has been used for the treatment of primary nonfunction of hepatic allografts and fulminant hepatic failure. It has been proposed that this therapy may improve hepatic blood flow via the vasodilating properties of PGE1. However, PGE1 undergoes extensive metabolic inactivation by the lung and the concentration of PGE1 reaching the liver during intravenous administration has not been determined. Thus, we measured plasma PGE1 concentrations in patients with hepatic dysfunction being treated with PGE1 and in a swine model of PGE1 infusion. We also determined the hemodynamic effects of PGE1 infusion in swine. Blood was sampled from the pulmonary artery, carotid artery, portal vein, and hepatic vein in swine infused with PGE1 (range, 0.67-4.9 microg/kg/hr) demonstrating: (1) a pulmonary extraction ratio of PGE1 of 0.78 +/- 0.12, (2) a splanchnic extraction ratio of PGE1 of 0.54 +/- 0.23, and (3) levels of PGE1 in the systemic circulation of </= 78 pg/ml, even at the highest infusion rates. Despite significant increases in body temperature and pulse rate, hepatic hemodynamics were not affected by the PGE1 infusions in healthy swine. Seven patients receiving intravenous PGE1 for hepatic dysfunction (0.11-1.30 microg/kg/hr) had a pulmonary extraction ratio of 0.69 +/- 0.17. Systemic arterial concentrations of PGE1 were </= 62 pg/ml. These results suggest that due to clearance of PGE2 in the pulmonary and splanchnic circulations, current clinical protocols for intravenous administration of PGE1 are not likely to affect perihepatic hemodynamics.
Adult↗
The isoprostanes. Current knowledge and directions for future research.
The isoprostanes are a unique series of prostaglandin-like compounds formed in vivo from the free radical-catalyzed peroxidation of arachidonic acid independent of the cyclooxygenase enzyme. The purpose of this commentary is to summarize the status of our current knowledge regarding the isoprostanes and discuss what we consider to be avenues for further research. Novel aspects related to the biochemistry of isoprostane formation and methods by which these compounds are analyzed, including potential pitfalls that may occur during analysis, are discussed first. The isoprostanes possess potent biological activity, and their potential role in mediating certain aspects of the detrimental effects of oxidant stress is then examined. In addition, evidence is presented that these biological effects may be mediated through interaction with a unique receptor. A considerable portion of this commentary deals with the utility of measuring isoprostanes as markers of oxidant injury both in vitro and in vivo. A number of studies have shown these compounds to be extremely accurate markers of lipid peroxidation in animal models of oxidative stress and have illuminated the role of oxidant injury in association with several human diseases. Finally, based upon our current knowledge of the isoprostanes, directions for future research are proposed.
Arachidonic Acid↗
Evidence of free radical-mediated injury (isoprostane overproduction) in scleroderma.
OBJECTIVE: Free radical-induced oxidative stress with consequent lipid peroxidation and resultant tissue damage has been suggested as a potential mechanism of the pathogenesis of scleroderma. However, because reliable measurement of lipid peroxidation in vivo is difficult, it has not been possible to adequately examine this hypothesis. We have previously described a series of bioactive prostaglandin F2-like compounds, termed F2-isoprostanes, produced in vivo in humans by the non-cyclooxygenase, free radical-catalyzed, peroxidation of arachidonic acid and have shown them to be a reliable measure of lipid peroxidation in vivo. In the present study, we determined whether scleroderma is associated with enhanced oxidative stress. METHODS: As a measure of oxidative stress, we determined urinary concentrations of a tetranor-dicarboxylic acid metabolite of F2-isoprostanes (F2IP-M) by mass spectrometry in 8 patients with scleroderma (representing a wide spectrum of disease, including limited disease with refractory digital ulceration or pulmonary hypertension, and diffuse disease) and in 10 healthy control subjects. RESULTS: F2IP-M concentrations were significantly higher in patients with scleroderma (mean +/- SEM 3.41 +/- 0.64 ng/mg of creatinine) than in healthy controls (1.22 +/- 0.14 ng/mg of creatinine) (P = 0.002). These elevations occurred in patients with limited disease and in those with diffuse disease. CONCLUSION: The increased level of urinary F2IP-M supports the hypothesis that free radical-induced oxidative injury occurs in scleroderma and provides a biologic marker whose relationship to disease activity and disease therapy may be important. These findings may also provide a rationale for exploring whether antioxidant therapy may influence the natural course of the disease.
Adult↗
Isoprostanes--prostaglandin-like compounds formed in vivo independently of cyclooxygenase: use as clinical indicators of oxidant damage.
F2-isoprostanes are prostanoids produced independently of cyclooxygenase by free radical-catalyzed peroxidation of arachidonic acid-containing lipids. Quantification of F2-isoprostanes from biologic fluids and tissues represents an important advance in the detection and measurement of lipid peroxidation in vivo. In addition, efforts to understand both the biophysical effects of isoprostane containing lipids and the biologic effects of free isoprostanes should lead to a better understanding of the mechanisms responsible for oxidant stress-related alterations in homeostasis. Continued application of F2-isoprostane measurement in experimental models of free radical-induced injury and human disease may allow better design and evaluation of antioxidant therapeutic strategies.
Animals↗
Demonstration of halothane-induced hepatic lipid peroxidation in rats by quantification of F2-isoprostanes.
BACKGROUND: Halothane can be reductively metabolized to free radical intermediates that may initiate lipid peroxidation. Hypoxia and phenobarbital pretreatment in Sprague-Dawley rats increases reductive metabolism of halothane. F(2)-isoprostanes, a novel measure of lipid peroxidation in vivo, were used to quantify halothane-induced lipid peroxidation in rats. METHODS: Rats were exposed to 1% halothane or 14% O(2) for 2 h. Pretreatments included phenobarbital, isoniazid, or vehicle. Rats also were exposed to halothane, enflurane, and desflurane at 21% O(2). Lipid peroxidation was assessed by mass spectrometric quantification of F(2)-isoprostanes. RESULTS: Exposure of phenobarbital-pretreated rats to 1% halothane at 21% O(2) for 2 h caused liver and plasma F(2)-isoprostane concentrations to increase fivefold compared to nonhalothane control rats. This halothane-induced increase was enhanced by 14% O(2), but hypoxia alone had no significant effect. Alanine aminotransferase activity at 24 h was significantly increased only in the 1% halothane/14% O(2) group. The effect of cytochrome P450 enzyme induction on halothane-induced F(2)-isoprostane production and liver injury was determined by comparing the effects of isoniazid and phenobarbital pretreatment with no pretreatment under hypoxic conditions. Halothane caused 4- and 11-fold increases in plasma and liver F(2)-isoprostanes, respectively, in non-pretreated rats, whereas isoniazid pretreatment had no effect. Phenobarbital pretreatment potentiated halothane-induced lipid peroxidation with 9- and 20-fold increases in plasma and liver F(2)-isoprostanes, respectively. Alanine aminotransferase activity was increased only in this group. At ambient oxygen concentrations, halothane but not enflurane or desflurane, caused F(2)-isoprostanes to increase. CONCLUSIONS: Specific halothane-induced lipid peroxidation was demonstrated in Sprague-Dawley rats using quantification of F(2)-isoprostanes and was increased by hypoxia and phenobarbital pretreatment, but not isoniazid pretreatment.
Anesthetics, Inhalation↗
Ritual Samoan body tattooing and associated sporotrichosis.
A 36 year old Samoan male who developed cutaneous sporotrichosis as a result of traditional body tattooing is reported. It is considered that the tattooing procedure was responsible for introducing the infection. He was treated with itraconazole, which produced resolution of the lesions.
Adult↗