Novel Amiloride Derivatives That Inhibit Bacterial Motility across Multiple Strains and Stator Types.

The bacterial flagellar motor (BFM) is a protein complex that confers motility to cells and contributes to survival and virulence. The BFM consists of stators that are ion-selective membrane protein complexes and a rotor that directly connects to a large filament, acting as a propeller. The stator complexes couple ion transit across the membrane to torque that drives rotation of the motor. The most common ion gradients that drive BFM rotation are protons (H+) and sodium ions (Na+). The sodium-powered stators, like those in the PomA/PomB stator complex of Vibrio spp., can be inhibited by sodium channel inhibitors, in particular, by phenamil, a potent and widely used inhibitor. However, relatively few new sodium motility inhibitors have been described since the discovery of phenamil. In this study, we characterized two possible motility inhibitors, HM2-16F and BB2-50F, from a small library of previously reported amiloride derivatives. We used three approaches: effect on rotation of tethered cells, effect on free-swimming bacteria, and effect on rotation of marker beads. We showed that both HM2-16F and BB2-50F stopped rotation of tethered cells driven by Na+ motors comparable to phenamil at matching concentrations and could also stop rotation of tethered cells driven by H+ motors. Bead measurements in the presence and absence of stators confirmed that the compounds did not inhibit rotation via direct association with the stator, in contrast to the established mode of action of phenamil. Overall, HM2-16F and BB2-50F stopped swimming in both Na+ and H+ stator types and in pathogenic and nonpathogenic strains. IMPORTANCE Here, we characterized two novel amiloride derivatives in the search for antimicrobial compounds that target bacterial motility. These compounds were shown to inhibit flagellar motility at 10 µM across multiple strains: from nonpathogenic Escherichia coli with flagellar rotation driven by proton or chimeric sodium-powered stators, to proton-powered pathogenic E. coli (enterohemorrhagic E. coli or uropathogenic E. coli [EHEC or UPEC, respectively]), and finally, sodium-powered Vibrio alginolyticus. Broad antimotility compounds such as these are important tools in our efforts to control virulence of pathogens in health and agricultural settings.

Making a move in exercise referral: co-development of a physical activity referral scheme.

Background: Translational research is required to ensure exercise referral schemes (ERSs) are evidence-based and reflect local needs. This article reports process data from the co-development phase of an ERS, providing an insight into (i) factors that must be considered when translating evidence to practice in an ERS setting, and (ii) challenges and facilitators of conducting participatory research involving multiple stakeholders. Methods: An ERS was iteratively co-developed by a multidisciplinary stakeholder group (commissioners, managers, practitioners, patients and academics) via five participatory meetings and an online survey. Audio data (e.g. group discussions) and visual data (e.g. whiteboard notes) were recorded and analysed using NVivo-10 electronic software. Results: Factors to consider when translating evidence to practice in an ERS setting included (i) current ERS culture; (ii) skills, safety and accountability; and (iii) resources and capacity. The co-development process was facilitated by needs-analysis, open questions, multidisciplinary debate and reflective practice. Challenges included contrasting views, irregular attendance and (mis)perceptions of evaluation. Conclusion: The multidisciplinary co-development process highlighted cultural and pragmatic issues related to exercise referral provision, resulting in an evidence-based intervention framework designed to be implemented within existing infrastructures. Further work is required to establish the feasibility and effectiveness of the co-developed intervention in practice.

Comparison of controlled-release and immediate-release oxycodone tablets in patients with cancer pain.

PURPOSE: This study compared the clinical efficacy of oxycodone hydrochloride controlled-release (CR) tablets administered every 12 hours with immediate-release (IR) oxycodone tablets administered four times daily in patients with cancer-related pain. PATIENTS AND METHODS: Cancer patients who required therapy for moderate to severe pain were randomized to CR oxycodone every 12 hours (n=81) or IR oxycodone four times daily (n=83) for 5 days in a multicenter, double-blind study. Pain intensity was assessed four times daily (categorical scale of none, slight, moderate, and severe); acceptability of therapy was assessed twice daily (categorical scale of very poor, poor, fair, good, and excellent). RESULTS: Pain intensity remained slight during the study, with mean oxycodone doses of 114 mg/d (range, 20 to 400 mg/d) for CR and 127 mg/d (range, 40 to 640 mg/d) for IR. Acceptability of therapy was fair to good with both treatments. While standard conversion ratios provided an acceptable dose for many patients, a protocol amendment that allowed initial titration and use of rescue medication reduced the discontinuation rate for lack of acceptable pain control (from 34% to 4% with CR and from 31% to 19% with IR before and after amendment, respectively) without increasing the discontinuation rate for adverse events (from 8% to 7% with CR and from 13% to 11% with IR). Fewer adverse events were reported with CR (109) than with IR (186) oxycodone (P=.006). CONCLUSION: CR oxycodone every 12 hours was as effective as IR oxycodone four times daily in managing moderate to severe cancer-related pain and was associated with fewer reports of adverse events.

Liposome-mediated augmentation of catalase in alveolar type II cells protects against H2O2 injury.

Oxidant injury to the alveolar epithelium can be mediated by exposure to oxidant gases such as O2 at high concentrations and O3, inflammatory cell-derived reactive O2 species, and the intracellular metabolism of xenobiotics such as paraquat. An in vitro model of alveolar epithelial oxidant injury was developed based on exposure of cultured rat type II pneumocytes to superoxide and hydrogen peroxide (H2O2) enzymatically generated in the culture medium. Cytotoxicity was assessed by the release of lactate dehydrogenase (LDH) into the culture medium, which was a more reliable indicator of damage than release of 51Cr by prelabeled cells. Incubation of cells for 6-8 h with xanthine plus xanthine oxidase and glucose plus glucose oxidase induced the release of greater than 50% of total intracellular LDH. Oxidant exposure also resulted in significant detachment of cells from culture dishes. Modulation of oxidant damage was accomplished using liposomes as vectors for the delivery of catalase. Treatment of cells with catalase liposomes for 2 h resulted in augmentation of cellular catalase specific activities up to 631% of controls. Catalase was partitioned into intracellular and surface-associated compartments in catalase liposome-treated cells. Partial and complete protection against oxidant injury, induced by xanthine plus xanthine oxidase and glucose plus glucose oxidase, respectively, was achieved by pretreatment of cells with catalase liposomes. LDH release during oxidant exposure was inversely related to augmentation of cellular catalase activities. Catalase liposome-treated cells also exhibited an enhanced ability to scavenge enzymatically generated H2O2 from the culture medium. These observations suggest a useful approach to modulation of alveolar injury induced by reactive O2 species.

Controlled-release oxycodone compared with controlled-release morphine in the treatment of cancer pain: a randomized, double-blind, parallel-group study.

Controlled-release oral formulations of oxycodone and morphine are both suitable analgesics for moderate to severe pain. They were compared in cancer-pain patients randomized to double-blind treatment with controlled-release oxycodone (n = 48) or controlled-release morphine (n = 52) every 12 h for up to 12 days. Stable analgesia was achieved by 83% of controlled-release oxycodone and 81% of controlled-release morphine patients in 2 days (median). Following titration to stable analgesia, pain intensity (0=none to 3=severe) decreased from baseline within each group (p

The use of controlled-release oxycodone for the treatment of chronic cancer pain: a randomized, double-blind study.

To compare the effectiveness and safety of controlled-release (CR) oxycodone tablets with immediate-release (IR) oxycodone in patients with chronic cancer pain, a multicenter, randomized, double-blind, parallel-group study was performed in 111 patients with cancer pain. Patients were treated with 6 to 12 tablets or capsules of fixed-combination opioid/nonopioid analgesics per day at study entry. Patients received 30 mg of CR oxycodone tablets every 12 hr or 15 mg of IR oxycodone four times daily for 5 days. No titration or supplemental analgesic medications were permitted. The mean (+/- SE) baseline pain intensity (0 = none, 1 = slight, 2 = moderate, 3 = severe) was 1.5 +/- 0.1 for the CR oxycodone-treated group and 1.3 +/- 0.1 for the group given IR oxycodone (P > 0.05). The 5-day mean pain intensity was 1.4 +/- 0.1 and 1.1 +/- 0.1 for the CR and IR groups, respectively (P > 0.05). Discontinuation rates were equivalent (33%). There was no significant difference between treatment groups in the incidence of adverse events. This study demonstrates that cancer pain patients given 6 to 12 tablets or capsules of fixed-dose combination analgesics can be equally well treated with CR oxycodone administered every 12 hr or IR oxycodone four times daily at the same total daily dose. CR oxycodone offers the benefits of twice daily dosing.

Oxidant injury to the alveolar epithelium: biochemical and pharmacologic studies.

This multifaceted study involved a combined biochemical and cellular analysis of oxidant metabolism by a lung cell at risk from injury by endogenous and environmental oxidants, the pulmonary alveolar type II epithelial cell. Within the framework of this study, a method was developed for effectively delivering antioxidant enzymes and alpha-tocopherol to the intracellular compartment of alveolar epithelial cells. Alveolar type II cells are key sources of pulmonary surfactant phospholipids and apoproteins and serve as progenitors of type I alveolar epithelium, thus playing an important role in the re-epithelialization of the lung alveolus after exposure to pulmonary oxidants. The type I and II pulmonary epithelium also play an essential collaborative role in maintaining the integrity of the air-blood barrier of the lung. Because of these critical properties of the alveolar epithelium and their recognized sensitivity to oxidant stress derived from diverse sources, such as activated inflammatory cells, hyperoxia, the environmental oxidants and nitrogen dioxide, and surgical procedures, such as cardiopulmonary bypass and lung transplantation, we endeavored to understand more about the oxidant metabolism and antioxidant pharmacology of these cells. In our experiments, we made the observation that loss of differentiated oxidant generation and antioxidant properties of type II cells occurs very rapidly in vitro. For example, we observed a 50% to 75% reduction in the specific activities of type II cell superoxide dismutase, catalase, and glutathione peroxidase, all critical scavengers of cell superoxide and hydrogen peroxide and key enzymes in the attenuation of hydroxyl radical formation. Although the differentiated characteristics of the type II cell antioxidant defenses changed in vitro, they may have become more reflective of type I alveolar epithelial cells. The type I cell is the most vulnerable for oxidant damage in the alveolus because of its large surface area and the possibility of a reduced antioxidant capacity compared to type II alveolar epithelium. In spite of this limitation, we were able to culture type II cells and study their adaptive and toxic responses to exogenously administered oxidant stress. We also observed that a significant source of self-generated oxidants in type II cells was the enzyme xanthine oxidase. Normal rates of oxidant production by this enzyme had an inhibitory effect on incorporation of biosynthetic precursors into surfactant phospholipids; these effects were eliminated by the xanthine oxidase inhibitor, allopurinol.(ABSTRACT TRUNCATED AT 400 WORDS)

Cardiac effects of esophageal stimulation: possible relationship between gastroesophageal reflux (GER) and sudden infant death syndrome (SIDS).

Twenty-six artificially ventilated newborn pigs were subjected to simulated gastroesophageal reflux; saline (10 cc) of varying pH was flushed through the esophagus from below. At a given pH threshold, reflex bradycardia, which could be blocked by atropine, was elicited. Transecting of the superior laryngeal nerves, the recurrent laryngeal nerves, and the pharyngeal plexus nerves did not block the reflex bradycardia. However, bypassing the regions superior to the esophagus with a shunt prevented the bradycardia. These results indicate that bradycardia caused by gastroesophageal reflux is independent of changes in ventilation and may be an important cause of sudden infant death.

Effects of acute hypoxemia on responses to dopamine and dobutamine in neonatal swine.

The effects of acute hypoxemia on cardiovascular responses to dopamine (DA) and dobutamine (DB) were studied in 2-4 day (n = 21) and 13-17 day (n = 27) old swine under sodium pentobarbital anesthesia. Sequential 10-min infusions of 2, 5 and 15 mu/kg/min of DA or DB or normal saline were administered under conditions of normoxemia and repeated during hypoxemia. Data are presented as mean percent of baseline value +/- SE. During normoxemia, DA increased aortic pressure, cardiac output, heart rate, LV dP/dt max and renal resistance and decreased mesenteric resistance in both ages, while DB increased heart rate, LV dP/dt max and cardiac output in both age groups and decreased total arterial resistance and renal resistance in the youngest. Increases in LV dP/dt max were larger with DA as compared to DB (p less than 0.05) with the highest dose in younger (152 +/- 5% vs. 124 +/- 4%) and older (201 +/- 29% vs. 157 +/- 9%) animals. Hypoxemia reduced heart rate responses to DA and DB in older piglets, contractility responses to DB in older animals, aortic pressure responses to DA in all animals and renal resistance responses to DA in older animals. Hypoxemia had little or no effect on cardiac output and total arterial, mesenteric and carotid resistance responses to DA and DB. Thus, in the newborn: 1) DA is a stronger inotropic agent during normoxemia and hypoxemia while the chronotropic effects of the two drugs did not differ. 2) Changes in cardiac output with both drugs did not differ significantly during normoxemia and were not affected by hypoxemia.(ABSTRACT TRUNCATED AT 250 WORDS)

Analgesic efficacy and safety of two oral controlled-release morphine preparations in orthopedic postoperative pain.

This single-dose, double-blind, randomized, parallel-group study compared the analgesic efficacy and safety of MS Contin (MSC) and Oramorph SR (OSR), two controlled-release preparations of oral morphine sulfate, in patients following orthopedic surgery. One hundred patients received MSC 30 mg, MSC 60 mg, or OSR 60 mg (two 30-mg tablets) when postoperative pain became moderate or severe. Patients self-rated pain intensity and relief on categorical (CAT) and visual analogue scales (VAS) hourly for up to 12 hours. MSC 60 mg produced the greatest peak analgesic effect and was more efficacious than OSR 60 mg through the sixth hour, with statistical significance achieved at 1, 2, and 3 hours postdosage. Compared with OSR 60 mg, both MSC dosages provided significantly more rapid times to peak effect by CAT and VAS ratings. The OSR group experienced almost twice as many adverse events as did the two MSC groups and also reported somnolence and dizziness more frequently. MSC 60 mg provided more rapid and greater peak analgesia with fewer adverse effects than did OSR 60 mg.

Chest pain, dyspnea on exertion, and exercise induced asthma in children and adolescents.

UNLABELLED: The contribution of maximal exercise tests to the evaluation of 180 patients with chest pain associated with exercise (n = 147) or dyspnea on exertion (DOE, n = 33) was examined. The ages ranged from 5 to 22 (mean 13.2) years, and 68 patients were females. All patients had a normal cardiovascular examination, electrocardiogram, chest x-ray, and 2D-echocardiogram. Maximal exercise tests were performed on a treadmill or bicycle ergometer, and flow volume loops were performed before and after exercise (n = 65). Exercise tests did not reveal any cardiovascular abnormalities, but 14 patients with chest pain (9.5%) and seven patients with DOE (21.2%) developed exercise-induced asthma. Postexercise decrease in peak expiratory flow rate was 26.2 +/- 3.7 percent in patients with chest pain and 39.4 +/- 8.9 percent in those with DOE. Only five patients had a personal history and four others had a family history of asthma. Seven patients had a personal or family history of allergies. IMPLICATIONS: exercise-induced asthma should be considered in pediatric patients with symptoms of chest pain or dyspnea on exertion; when exercise tests are performed, flow volume loops should be included before and after exercise; maximal exercise tests are unlikely to unmask any cardiovascular abnormalities in such patients.

Arachidonic acid stimulates protein tyrosine phosphorylation in vascular cells.

Arachidonic acid and its metabolites are important cellular mediators. In this study, we report a novel role for arachidonic acid in vascular cell signaling. We tested the effects of exogenous arachidonic acid on protein tyrosine phosphorylation in cultured vascular endothelial and smooth muscle cells. Arachidonic acid stimulated the phosphorylation of tyrosine-containing proteins of approximately 58, 93, and 120 kDa in the three cell types studied. This response was dose dependent, with a maximum effect observed with 40 microM arachidonic acid. Phosphorylation was rapid and transient, reaching a peak 0.5 min after the addition of arachidonic acid and returning to baseline by 8 min. A common set of protein substrates was phosphorylated in smooth muscle cells treated with the Ca(2+)-mobilizing agonist endothelin, concomitant with an increase in endogenous unesterified arachidonic acid. To determine whether the protein tyrosine phosphorylation was due to arachidonic acid or to a metabolite, we used inhibitors of cyclooxygenase, lipoxygenase, and epoxygenase pathways. Ibuprofen, nordihydroguaiaretic acid, eicosatriynoic and eicosatetraynoic acids, and 8-methoxypsoralen failed to inhibit the arachidonic acid-mediated response. We also found increased protein tyrosine phosphorylation after treatment with oleic, linolenic and gamma-linoleic acid. These results suggest a mechanism of protein tyrosine phosphorylation that is directly stimulated by unmetabolized unsaturated fatty acids.

Ca(2+)-independent arachidonic acid release by vascular endothelium requires protein synthesis de novo.

We investigated the mechanism by which the G-protein activators aluminium fluoride and vanadate stimulate arachidonic acid release in pig aortic endothelial cells. Our previous study demonstrated a novel Ca(2+)-independent pathway of phospholipase A2 (PLA2) activation stimulated by aluminium fluoride in this model. In the present study, we found that sodium metavanadate stimulated a rapid concentration-dependent release of [3H]arachidonic acid from prelabelled cells. A more than 3-fold enhancement of arachidonic acid release was achieved in cells treated with 1 mM vanadate for 20 min. Synthesis of prostaglandin products was similarly enhanced. The release of arachidonic acid was not dependent on the presence of extracellular Ca2+, but did require protein synthesis de novo. Both cycloheximide and actinomycin D completely blocked aluminium fluoride- and vanadate-stimulated arachidonic acid release. Because fluoride and vanadate are known protein tyrosine phosphatase inhibitors, it is possible that PLA2 activation occurred secondarily to changes in protein tyrosine phosphorylation. Both aluminium fluoride and vanadate stimulated the rapid phosphorylation of 58, 93 and 120 kDa tyrosine-containing protein substrates. However, in contrast with arachidonic acid release, this response was found to be sensitive to the presence of extracellular Ca2+ and insensitive to blockers of protein synthesis de novo. Furthermore H2O2 treatment resulted in rapid tyrosine phosphorylation of the same substrates without a concomitant increase in arachidonic acid release. These results suggest that the effects of aluminium fluoride and vanadate on PLA2 are not due to changes in protein tyrosine phosphorylation, but do require rapid protein synthesis de novo.

Glutathione redox status of control and cadmium oxide-exposed rat lungs during oxidant stress.

Activities of enzymes responsible for the maintenance of reduced glutathione (GSH) levels have been shown in a previous study to be increased in rat lungs following a 3-h exposure to cadmium oxide aerosols at 5.0 mg/m3. In this study, the ability of the lung to maintain levels of GSH during challenge with tert-butyl hydroperoxide (tBuOOH) was evaluated in isolated perfused lungs from control and cadmium oxide-exposed rats. Changes in glutathione redox status were indicated by measurements of nonprotein sulfhydryls (NPSH), total glutathione (1/2 GSH + GSSG), and glutathione disulfide (GSSG) in liquid nitrogen freeze-clamped lungs after 3-min infusions with 0-0.6 mM tBuOOH. In control and cadmium oxide-exposed lungs, levels of 1/2 GSH + GSSG remained constant over the range of 0-0.6 mM tBuOOH, indicating that no loss of glutathione from the system had occurred. In experiments with control lungs, levels of NPSH fell from 8.04 +/- 0.22 to 3.09 +/- 0.40 mumol/g dry weight when tBuOOH concentrations were increased from 0 to 0.6 mM (n = 20-23). In cadmium oxide-exposed lungs, NPSH levels also decreased proportionally to increases in GSSG. However, at concentrations of 0.075 and 0.15 mM tBuOOH, significantly smaller decreases in NPSH levels were observed in cadmium oxide-exposed lungs compared with controls. This protection against the GSH-depleting effects of tBuOOH might be explained by increased tissue levels of GSH-related enzymes.

Pulmonary cadmium oxide toxicity in the rat.

Although occupational exposures to cadmium have usually involved inhalation of insoluble cadmium oxide (CdO) particles, experimental studies of pulmonary cadmium toxicity have relied on aerosol exposures to soluble cadmium chloride particles. The present study describes a model of acute lung injury based on single 3-h exposures of rats to 0.5 and 5.3 mg/m3 CdO. Biochemical changes were correlated with pathological observations for 15 d postexposure to CdO. Four days following CdO exposure, histopathological observations included focal areas of epithelial hyperplasia, a mononuclear interstitial infiltrate, and increased numbers of alveolar macrophages. In the high-dose group, these changes were correlated with increases in tissue protein and DNA contents of 217% and 195% of controls, respectively. While lungs from the low-dose exposures had returned to a normal appearance by 15 d postexposure, high-dose-exposed lungs exhibited an increase in noncellular thickening of the interstitium and a continued general hypercellularity at this time. In the high-dose exposure group, activities of the enzymes glutathione peroxidase, glutathione reductase, and the dehydrogenase of glucose 6-phosphate and 6-phosphogluconate were significantly elevated two- to fivefold at 2-4 d postexposure. When a correction was made for changes in lung cell number, significant increases were observed only in activities of the pentose-cycle dehydrogenases at 180-238% of controls. These increases suggested an enhanced ability of CdO-exposed lungs to generate the pentose-cycle products NADPH and ribose 5-phosphate, which would be needed for lipid and nucleic acid biosynthesis expected during the proliferative stages of epithelial repair. This study has demonstrated that the response to CdO exposure includes the induction of enzymatic activities that are related to antioxidant defense and lung repair.

Tunicamycin increases intracellular calcium levels in bovine aortic endothelial cells.

Tunicamycin is a nucleoside antibiotic that inhibits protein glycosylation and palmitoylation. The therapeutic use of tunicamycin is limited in animals because of its toxic effects, particularly in cerebral vasculature. Tunicamycin decreases palmitoylation of the endothelial isoform of nitric oxide synthase, stimulates nitric oxide synthesis, and increases the concentration of intracellular calcium ([Ca2+]i) in bovine aortic endothelial cells (B. J. Buckley and A. R. Whorton. FASEB J. 11: A110, 1997). In the present study, we investigated the mechanism by which tunicamycin alters [Ca2+]i using the Ca2+-sensitive dye fura 2. We found that tunicamycin increased [Ca2+]i without increasing levels of inositol phosphates. When cells were incubated in the absence of extracellular Ca2+, [Ca2+]i rapidly rose in response to tunicamycin, although a full response was not achieved. The pool of intracellular Ca2+ mobilized by tunicamycin overlapped with that mobilized by thapsigargin. Extracellular nickel blocked a full response to tunicamycin when cells were incubated in the presence of extracellular Ca2+. The effects of tunicamycin on [Ca2+]i were partially reversed by washing out the drug, and the remainder of the response was inhibited by removing extracellular Ca2+. These results indicate that tunicamycin mobilizes Ca2+ from intracellular stores in a manner independent of phospholipase C activation and increases the influx of Ca2+ across the plasma membrane.

Adaptive responses to peroxynitrite: increased glutathione levels and cystine uptake in vascular cells.

We and others recently demonstrated increased glutathione levels, stimulated cystine uptake, and induced gamma-glutamylcysteinyl synthase (gamma-GCS) in vascular cells exposed to nitric oxide donors. Here we report the effects of peroxynitrite on glutathione levels and cystine uptake. Treatment of bovine aortic endothelial and smooth muscle cells with 3-morpholinosydnonimine (SIN-1), a peroxynitrite donor, resulted in transient depletion of glutathione followed by a prolonged increase beginning at 8-9 h. Concentration-dependent increases in glutathione of up to sixfold occurred 16-18 h after 0.05-2.5 mM SIN-1. Responses to SIN-1 were inhibited by copper-zinc superoxide dismutases and manganese(III)tetrakis(1-methyl-4-pyridyl)porphyrin pentachloride, providing evidence for peroxynitrite involvement. Because glutathione synthesis is regulated by amino acid availability, we also studied cystine uptake. SIN-1 treatment resulted in a prolonged increase in cystine uptake beginning at 6-9 h. Increases in cystine uptake after SIN-1 were blocked by inhibitors of protein and RNA synthesis, by extracellular glutamate but not by extracellular sodium. These studies suggest induction of the x(c)(-) pathway of amino acid uptake. A close correlation over time was observed for increases in cystine uptake and glutathione levels. In summary, vascular cells respond to chronic peroxynitrite exposure with adaptive increases in cellular glutathione and cystine transport.

Regulation of endothelial cell prostaglandin synthesis by glutathione.

Prostaglandin synthesis in in vitro systems is dependent on glutathione and peroxide concentrations. We tested the effects of glutathione depletion and H2O2 exposure on prostaglandin synthesis in cultured porcine aortic endothelial cells. Depletion of glutathione using buthionine sulfoximine (BSO), diethylmaleate, and 2,4-chlorodinitrobenzene increased prostaglandin synthetic capacity. Production of prostacyclin, but not prostaglandin E2, from exogenous arachidonic acid was significantly greater than in controls. Glutathione depletion also resulted in enhanced production of prostacyclin from exogenous prostaglandin H2. These responses were not due to direct effects of glutathione-depleting agents on prostaglandin synthetic enzymes. Exposure to H2O2 also altered prostaglandin synthetic capacity in endothelial cells. While 5 microM H2O2 stimulated prostaglandin production from exogenous arachidonate, 25 and 50 microM were found to be inhibitory. Prostaglandin synthetic capacity was greater in BSO-treated cells which were exposed to 5 and 10 microM H2O2 than in cells exposed to H2O2 alone. However, prostaglandin synthetic capacity was greatly reduced in BSO-treated cells exposed to 50 microM H2O2. Thus, normal levels of cellular glutathione exert an inhibitory influence on prostaglandin synthesis. However, glutathione depletion increases the sensitivity of prostaglandin synthesis to inhibition by 50 microM H2O2.