Release Of Oxygen Free Radicals Research Articles

Nitric oxide and cardioprotection during ischemia-reperfusion.

Coronary artery reperfusion is widely used to restore blood flow in acute myocardial infarction and limit its progression. However, reperfusion of ischemic myocardium results in reperfusion injury and persistent ventricular dysfunction even when achieved after brief periods of ischemia. Normally, small amounts of nitric oxide (NO) generated by endothelial NO synthase (eNOS) regulates vascular tone. Ischemia-reperfusion triggers the release of oxygen free radicals (OFRs) and a cascade involving endothelial dysfunction, decreased eNOS and NO, neutrophil activation, increased cytokines and more OFRs, increased inducible NO synthase (iNOS) and marked increase in NO, excess peroxynitrite formation, and myocardial injury. Low doses of NO appear to be beneficial and high doses harmful in ischemia-reperfusion. eNOS knock-out mice confirm that eNOS-derived NO is cardioprotective in ischemia-reperfusion. iNOS overexpression increases peroxynitrite but did not cause severe dysfunction. Increased angiotensin II (AngII) after ischemia-reperfusion inactivates NO, forms peroxynitrite and produces cardiotoxic effects. Beneficial effects of angiotensin-converting-enzyme inhibition and AngII type 1 (AT(1)) receptor blockade after ischemia-reperfusion are partly mediated through AngII type 2 (AT(2)) receptor stimulation, increased bradykinin and NO. Interventions that enhance NO availability by increasing eNOS might be beneficial after ischemia-reperfusion.

Protective effects of immunosuppressants and steroids against ischemia-reperfusion injury in cremaster muscle flap at microcirculatory level.

The sequential events between leukocytes and endothelium have significant implications in surgical procedures, trauma, vascular injury, and wound healing. These sequential events are mediated by free oxygen radicals, leukocytes, red blood cells, and endothelial cells. In this study, we investigated the protective effects of steroids and immunosuppressants against ischemia-reperfusion injury in cremaster muscle flaps at the microcirculatory level. In this experimental study, 50 male Sprague-Dawley rats, weighing about 120-130 g, were used. The rats were divided into five groups of 10 rats each: the control group (group 1, n = 10), methyl prednisolone group (group 2, n = 10), dexamethasone group (group 3, n = 10), cyclosporin A group (group 4, n = 10), and azathioprine (group 5, n = 10). Surgical procedures were divided into two stages. In the first stage, a cremaster muscle end-organ tube flap was created by extracting the testes and spermatic cord. The flap was placed into a subcutaneous tunnel in the anteromedial aspect of the ipsilateral limb. In the experimental groups, ischemia was performed by clamping the femoral artery and vein above and below the cremaster pedicle for 4 h. Then the clamps were removed, and perfusion of the cremaster muscle was allowed for 24 h. In the second stage, a round flap was obtained from the cremaster muscle tube-flap to evaluate microcirculation after 24 h of reperfusion, and dissected along its front wall using cauterization. Vessel diameter, red blood cell velocities, leukocyte activation, perfused capillaries, and endothelial edema index were measured and evaluated statistically. There was a significant decrease in the number of rolling, sticking, and transmigrating neutrophils of groups 2 (cyclosporin A), 4 (methylprednisolone), and 3 (azathioprine) (P < 0.05), whereas those of group 5 (dexamethasone) were not decreased (P > 0.05). There was a significant increase in the number of perfused capillaries of groups 2 (cyclosporin A), 4 (methylprednisolone), and 3 (azathioprine) (P < 0.05), nearly 0.75-, 0.5-, and 0.75-fold, respectively. We conclude that cyclosporin and azathioprine showed a protective effect on muscle tissue against ischemia-reperfusion injury by inhibiting leukocyte infiltration, and that methylprednisolone had a beneficial effect against ischemia-reperfusion injury by reducing the synthesis of eicosanoids, edema formation, and leukocyte infiltration. However, we believe that dexamethasone might have a salutary effect due to reducing the synthesis of eicosanoids, edema formation, and release of free oxygen radicals, but not due to leukocyte infiltration.

Pharmacological tools for the limitation of myocardial reperfusion injury Experimental evidence

Prompt re-establishment of coronary flow following coronary occlusion is mandatory for the preservation of myocardial tissue. The establishment of reperfusion treatment, originally by thrombolysis and subsequently by coronary interventions, was a major improvement in the management of patients with myocardial infarction. Myocardial damage is still seen even after very early initiation of successful reperfusion. Some of this damage has been ascribed to what has been labelled as reperfusion injury. Clinical manifestations of reperfusion injury include ventricular arrhythmia, compromised myocardial contractility and, most important for the subsequent prognosis, extension of the final infarct size. Many factors may contribute to the reperfusion injury. Among them is the release of free oxygen radicals with subsequent lipid peroxidation of cellular membranes. Intracellular calcium overload may cause excessive myofilament activation and impaired mitochondrial function, limit metabolic recovery and also activate proteases. Inflammatory activity and neutrophil activation are other factors of importance, as are endothelial dysfunction causing disturbances in nitric oxide (NO) availability and a compromised microvascular blood flow (the ‘noreflow’ phenomenon). Moreover, there is evidence that activation of the renin-angiotensin system may cause harm, e.g., by increasing the calcium content in the myocytes, by coronary vasoconstriction, and by modulation of the cardiac and vascular sympathetic activity. Some of these mechanisms may be interrelated. From considerations of their nature, an attractive hypothesis has been that some or several of these harmful effects may be counteracted by pharmacological interventions. This hypothesis has mainly been studied in the experimental setting. On the basis of the multiplicity of mechanisms that seem to be involved in the creation of reperfusion injury, a number of pharmacological agents have been investigated for their protective effect. Results from studies of some of them, namely beta-blockers, free oxygen radical scavengers, anti-oxidants, calcium channel blockers, inhibitors of the renin-angiotensin system and the combination of several of these drugs, are reviewed. Apart from beta-blockers, all these compounds have given effective protection from myocardial reperfusion damage. By studying them in different experimental settings and protocols, insight has been gained about important factors for possible future clinical applications of some or several of these drugs.

Aspirin treatment reduces oxygen free radical release by monocytes

Aspirin treatment reduces oxygen free radical release by monocytes

Protein Kinase Activation by Warm And Cold Hypoxia- Reoxygenation in Primary-Cultured Rat Hepatocytes–JNK 1/SAPK 1 Involvement in Apoptosis

Ischemia-reperfusion procedures induced severe hepatic damages owing to different processes related to hypoxia and reoxygenation (H/R) phases, including the consecutive oxygen free radical (OFR) release. Stress-activated protein kinases (SAPKs) could be activated by extracellular stimuli. The aim of this study was to show whether H/R stress conditions could stimulate these kinases, and especially c-jun-N-terminal kinase (JNK 1 /SAPK 1 ), to reveal a potential role of JNK 1 /SAPK 1 in the control of hepatocyte apoptosis. Primary cultured rat hepatocytes, isolated from other liver cells and blood flow, were subjected to warm and cold hypoxia-reoxygenation phases mimicking surgical and transplant conditions. The activation status of SAPKs was evaluated by immunoprecipitation or Western-blotting experiments, whereas apoptosis was assessed by measuring caspase activation and internucleosomal DNA fragmentation in vitro and by TUNEL reaction, in vivo. Hypoxia, and especially hypoxia-reoxygenation, significantly increased JNK 1 /SAPK 1 activation in cultured hepatocytes. Either in warm or cold conditions, OFR scavengers (N-Acetylcystein, Di-Phenyleneiodonium, Deferoxamine) decreased this stimulation. Warm ischemia-reperfusion also led to JNK activation. Hypoxia and especially hypoxia-reoxygenation induced programmed cell death in vivo and in vitro. This last phenomenon was inhibited when hepatocytes were treated with SB 202190, which was described as a potent inhibitor of p38 and JNK activities. Altogether, these results confirmed that JNK 1 /SAPK 1 was activated during the hypoxia-reoxygenation process, and that this activity participated in the onset of the apoptosis program. (Hepatology 2000;32:1029-1036.)

CoQ10 fails to protect brain against focal and global ischemia in rats

Objective: Release of oxygen free radicals occurs following cerebral ischemia. Studies show that oxygen free radicals mediate ischemic brain injury. CoQ10 is a potent free radical scavenger and may offset brain injury associated with reperfusion. We tested exogeneous CoQ10 as a neuroprotectant in rats following both global and focal ischemic insults. Methods: Rats were subjected to either 4-vessel occlusion ischemia (4-VO, 10 min occlusion, 7-day survival) or middle cerebral artery occlusion (MCAO, 120 min-occlusion, 22.5 h survival). Regional cerebral blood flows (rCBF) and physiological variables such as blood pressure, pO2, pCO2, plasma glucose and hematocrit were monitored and measured in focal ischemia. The animals were randomized to receive treatments of either phosphate buffered saline (PBS) vehicle or CoQ10 following global or focal ischemia. Injection times were at the end of ischemia and 3 h later for both models of ischemia. Histological outcomes are expressed as a percentage of hippocampal CA 1 cell injury in global ischemia or percentage of cortical infarct over that of non-ischemic hemisphere in focal ischemia. Results: In global ischemia, animals treated with PBS vehicle and CoQ10 had 86±5% ( n=8) and 83±10% ( n=8), respectively, of hippocampal CA 1 cell injury ( P>0.05). The percentage of infarct volumes in animals following focal ischemia were 23±9% (control, n=10) and 25±9% (CoQ10, n=10). There were no temperature or physiological differences between the two treatment groups. Conclusion: Acute treatment with CoQ10 via intraperitoneal injection does not prevent neuronal injuries following global and focal ischemia.

Kinetic scavenging effect of chinonin on no and oxygen free radicals generated from ischemia reperfusion myocardium and its protective effect on the myocardium

The kinetic scavenging effect of chinonin on NO and oxygen free radicals generated from ischemia reperfusion myocardium and its protective effect on the myocardium were studied by electron spin resonance (ESR) spin trapping technique. It was found that after 20 minutes ischemia, the first peak of oxygen free radical appeared at about 0.5 minutes after the beginning of reperfusion, then the release of oxygen free radicals decreased with time. The second peak appeared at about 3 minutes. Similarly, there were two peaks of creatine kinase (CK) release, which indicated the myocardial damage, the first one appeared concomitantly with the first oxygen free radical peak but the second one appeared later about 1 minute after appearance of the second peak of oxygen free radicals. The release of NO free radicals was not significant in the absence of L-arginine. However, it increased significantly in the presence of L-arginine and it also possesses a biphase profile. It could protect the ischemia-reperfusion damage in the presence of low concentration of L-arginine (0.1 mM), but in high L-arginine concentration (10 mM) it generated higher concentrations of NO leading to a more serious ischemia-reperfusion damage. Addition of chinonin could scavenge the free radicals and protect the ischemia-reperfusion injury, especially in the second phase. The reduction stoichiometry of chinonin for Fe(III) was measured.

Carnitines inhibits oxygen free radical release by endothelial cell and leukocytes

Carnitines inhibits oxygen free radical release by endothelial cell and leukocytes

The role of transforming growth factor beta in chronic renal allograft nephropathy.

The role of transforming growth factor beta in chronic renal allograft nephropathy.

Cascading glia reactions: a common pathomechanism and its differentiated control by cyclic nucleotide signaling.

A pathological glia activation, stimulated by inflammatory proteins, beta-amyloid, or brain ischemia, is discussed as a common pathogenic factor for progressive nerve cell damage in vascular and Alzheimer dementia. A critical point seems to be reached, if the cytokine-controlled microglial upregulation causes a secondary activation of astrocytes which loose the negative feedback control, are forced to give up their physiological buffering function, and may add to neuronal damage by the release of nitric oxide (NO) and by promoting toxic beta-amyloid formation. A strengthening of the cyclic adenosine-5',3'-monophosphate (cAMP) signaling exerted a differential inhibition of the stimulatory cytokines tumor necrosis factor-alpha (TNF-alpha) and interleukin-1 beta (IL-1 beta) released from cultured rat microglia, but maintained the negative feedback signal IL-6; cAMP inhibited also the release of free oxygen radicals (OR) but not of NO. Reinforcement of the NO-induced cyclic guanosine monophosphate (cGMP) increase by blockade of the phosphodiesterase (PDE) subtype-5 with propentofylline counterbalanced the toxic NO action that causes with OR neuronal damage by peroxynitrate formation. In rat cultured astrocytes, a prolonged cAMP elevation favored cell differentiation, the expression of a mature ion channel patter, and an improvement of the extracellular glutamate uptake. Cyclic AMP signaling could be strengthened by PDE blockade and by raising extracellular adenosine, which stimulates A2 receptor-mediated cAMP synthesis. Via an A1 receptor-mediated effect, elevated adenosine was found to overcome a deficient intracellular calcium mobilization resulting from an impaired muscarinic signaling at pathologically decreased acetylcholine concentrations. We suggest that pharmaca, which elevate extracellular adenosine and/or block the degradation of cyclic nucleotides, may be used to counteract glia-related neuronal damage in dementing processes.

Platelets, oxidant stress and erectile dysfunction: an hypothesis.

Time for primary review 30 days. The successful use of sildenafil (Viagra™) to treat vasculogenic erectile dysfunction (VED) has heightened awareness of the widespread prevalence of this disorder [1–3]. Sildenafil acts through the augmentation of nitric oxide (NO) actions on vascular/corpus cavernosal smooth muscle cell relaxation [1–4] (Fig. 1). Thus, the therapeutic use of sildenafil has also highlighted the central role played by NO in mediating normal erection and in the pathogenesis of VED. Nevertheless, we still know little about the basic pathophysiology of VED. Since prevention is preferable to cure, it is imperative to improve our understanding of the pathophysiology of VED. In this brief review, we suggest that another major consequence of diminished NO formation is the increased adhesion of platelets and leukocytes, in particular neutrophils, which may counter erection through release of vasconstrictors, superoxide and other oxygen free radicals. We also suggest that erection itself may trigger events that contribute to exacerbation of the disorder in the long-term, again through the adhesion of platelets and leukocytes. Fig. 1 Key events in normal penile erection — the central role of NO. Upon sexual arousal, sympathetic drive initiates erection through both sympathetic and non-adrenergic, non-cholinergic (NANC) innervation of the corpus cavernosum and pudendal arteries. NANC fibres release NO and acetylcholine stimulates the release of NO from local endothelium by activation of NO synthase (NOS). NO promotes relaxation of arterial and cavernosal smooth muscle through activation of guanylate cyclase and suppression of activator Ca2+ which in turn results in erection. It is now well established that impairment of the above systems is central to the aetiology of vasculopathic erectile dysfunction. Sildenafil promotes erection through inhibition of type V phosphodiesterase which blocks the metabolism of cGMP to inactive GMP. Thus, cGMP accumulates thereby promoting relaxation of the relevant VSMCs. Patients with … * Corresponding author. Tel.: +44-117-928-3154; fax: +44-117-929-9737 j.y.jeremy{at}bristol.ac.uk

Influence of Peroperative Lavage Solutions on Peritoneal Defence Mechanisms in vitro

To find out the in vitro reaction of mesothelial cells and polymorphonuclear leucocytes (PMN) to incubation with seven commonly-used lavage solutions. Experimental study. Laboratories, The Netherlands. Cultured human peritoneal mesothelial cells and isolated PMN. Incubation of cells with clinically used lavage solutions (sodium chloride, Hartmann's solution, povidone-iodine, Dakin's solution, taurolidine, chlorhexidine, and hydrogen peroxide). Activation of monolayers of mesothelial cells and PMN measured by release of oxygen free radicals (chemiluminescence) and interleukin (IL)-8 concentrations and toxic effects measured by morphology, release of lactate dehydrogenase, failure of the restriction of the passage of inulin, and incorporation of propidium iodide. All solutions activated and killed mesothelial cells and PMN to some extent; the more concentrated the solution the greater the effect on these cells. Lavage solutions both poison and stimulate mesothelial cells and neutrophils, and some solutions are more potent than others.

The role of neutrophils in myocardial ischemia-reperfusion injury.

Reperfusion of ischemic myocardium is necessary to salvage tissue from eventual death. However, reperfusion after even brief periods of ischemia is associated with pathologic changes that represent either an acceleration of processes initiated during ischemia per se, or new pathophysiological changes that were initiated after reperfusion. This 'reperfusion injury' shares many characteristics with inflammatory responses in the myocardium. Neutrophils feature prominently in this inflammatory component of postischemic injury. Ischemia-reperfusion prompts a release of oxygen free radicals, cytokines and other proinflammatory mediators that activate both the neutrophils and the coronary vascular endothelium. Activation of these cell types promotes the expression of adhesion molecules on both the neutrophils and endothelium, which recruits neutrophils to the surface of the endothelium and initiate a specific cascade of cell-cell interactions, leading first to adherence of neutrophils to the vascular endothelium, followed later by transendothelial migration and direct interaction with myocytes. This specific series of events is a prerequisite to the phenotypic expression of reperfusion injury, including endothelial dysfunction, microvascular collapse and blood flow defects, myocardial infarction and apoptosis. Pharmacologic therapy can target the various components in this critical series of events. Effective targets for these pharmacologic agents include: (a) inhibiting the release or accumulation of proinflammatory mediators, (b) altering neutrophil or endothelial cell activation and (c) attenuating adhesion molecule expression on endothelium, neutrophils and myocytes. Monoclonal antibodies to adhesion molecules (P-selectin, L-selectin, CD11, CD18), complement fragments and receptors attenuate neutrophil-mediated injury (vascular injury, infarction), but clinical application may encounter limitations due to antigen-antibody reactions with the peptides. Humanized antibodies and non-peptide agents, such as oligosaccharide analogs to sialyl Lewis, may prove effective in this regard. Both nitric oxide and adenosine exhibit broad spectrum effects against neutrophil-mediated events and, therefore, can intervene at several critical points in the ischemic-reperfusion response, and may offer greater benefit than agents that interdict at a single point in the cascade. The understanding of the molecular processes regulating actions of neutrophils in ischemic-reperfusion injury may be applicable to other clinical situations, such as trauma, shock and organ or tissue (i.e. vascular conduits) transplantation.

Antigenic stimulation in T-cell cultures in cardiomyopathies: differences in cytokine profiles.

Immune-mediated mechanisms are involved in the pathogenesis of cardiomyopathies. In this study, we investigate which pattern of immune response (Th1 or Th2) lies behind these diseases by analysing the basic cytokines secreted from PHA-cultured T lymphocytes and determining what differences, if any, exist between dilated cardiomyopathy (DMC) and hypertrophic cardiomyopathy (HCM). Two groups of patients were studied: 10 patients with DCM and 10 patients with HCM. Age- and sex-matched healthy individuals were used as controls. PHA-cultured T lymphocytes in the presence or absence of different myocardial antigen (MA) concentrations were measured. Interleukine-2 (IL-2), Interleukine-6 (IL-6) and Interferon-gamma (IFN-gamma) levels were measured in culture supernatants by an ELISA method. At the same time, delayed-type hyperactivity (DTH) against the same antigenic preparation was measured by the leukocyte migration inhibitory index technique. Patients were subdivided into DTH-positive and DTH-negative and re-examined for IL-2 cytokine expression. IL-6 levels were found to increase both in the presence and in the absence of MA in the patient groups compared to the controls. IL-2 levels were decreased in both groups, in an antigen dose-related manner. Anergic patients showed a further reduction in IL-2 levels for both groups of patients. IFN-gamma remained unaffected in the patient groups. Almost half of the patients exhibited anergy to the DTH reaction against MA. We conclude that, upon antigenic stimulation, the initially mounted immune response (increased IL-6) is somehow blocked/switched off in patients, resulting in an immunologic tolerance/unresponsiveness to MA (IL-2 decreased, IFN-gamma unchanged). Finally, increased IL-6 could lead to a perpetuation of immunologic injury through the release of oxygen-free radicals with a cytotoxic effect on the myocardium. We hypothesize an antigen-related, defective macrophage-Th1 cell reaction, which accounts for the differences in the IL-2 profile between the DCM and HCM groups, that might cause local immune responses to lead to immunosuppression (immune tolerance effect), thus contributing to the pathogenesis of cardiomyopathies.

Plasma-mediated stimulation of neutrophil superoxide anion production during coronary artery bypass grafting: role of endothelin-1.

Activation of polymorphonuclear neutrophils (PMN) and subsequent release of free oxygen radicals, including the superoxide anion (O2-) has been shown to result in postischaemic myocardial dysfunction during coronary artery bypass grafting (CABG). Several neutrophil-oriented stimuli are known to be released from myocardium during ischaemia and reperfusion. Release of endothelin-1 has been documented during CABG. The aim of the current study was to evaluate plasma-mediated neutrophil stimulation and to verify whether endothelin-1, known to be a stimulus for PMN, is involved in plasma-mediated stimulation of PMN during coronary artery bypass grafting. Plasma samples from peripheral artery, peripheral vein, and coronary sinus were obtained from 21 patients undergoing CABG before aortic clamping (global ischaemia), immediately after beginning reperfusion, and 30 min after reperfusion as well as from healthy controls. Plasma was incubated with PMN isolated from healthy donors preincubated in the presence of saline or specific endothelin-1 receptor antagonist (ET-A). PMN O2- production was measured spectrophotometrically. Plasma samples taken from the coronary sinus at the beginning of reperfusion were capable of higher stimulation of neutrophil superoxide anion production (24.2 +/- 2.0 nmol/5 x 10(6)PMN/30 min) than plasma obtained before reperfusion (15.6 +/- 1.5; p < 0.05) or plasma taken from peripheral artery (17.1 +/- 1.7; p < 0.05). Preincubation of PMN with endothelin-1 receptor antagonist decreased superoxide anion production by cells exposed to plasma taken from coronary sinus at the beginning of reperfusion (17.6 +/- 2.0, p < 0.05). Transcardiac release of soluble stimuli for PMN occurs as a result of myocardial ischaemia during CABG. Endothelin-1 may be involved in the plasma-mediated stimulation of neutrophil superoxide anion production.

The hydroxyl radical scavenger Nicaraven inhibits glutamate release after spinal injury in rats

Neuronal degeneration after trauma is mediated in part by release of excitatory amino acids (EAAs) and oxygen free radicals (OFR). We evaluated the effect of i.v. treatment with a hydroxyl radical scavenger ((+/-)-N,N'-propylenedinicotinamide; AVS) and spinal hypothermia (33 degrees C) on spinal CSF glutamate release after spinal trauma. In a control group, spinal compression evoked at 10 min a significant increase (5-fold) in glutamate which declined over 4 h (2.1-fold). AVS treatment attenuated glutamate release but had no additive effect. These data suggest that this compound can be effective in modulating spinal excitotoxicity resulting from increased OFR synthesis and corresponding potentiation of EAA release.

Prevention of Adriamycin-Induced Skin Necrosis with Various Free Radical Scavengers

Infiltration of antitumor agents into subcutaneous tissues may either result in a local area of self-resolving inflammation or progress to full-thickness loss of skin and underlying vital structures. Inadvertent extravasation of adriamycin can result in severe tissue necrosis. The mechanism of this tissue damage is believed to be release of oxygen free radicals into the tissue. After adriamycin extravasation, the treatment groups were made up according to drugs used, EGb 761, pentoxifylline, α-tocopherol acetate, and α-tocopherol succinate in rats. To prevent the necrosis and to decrease the tissue malondialdehyde levels, the most effective agent was found to be EGb 761, and pentoxifylline was also effective (P< 0.001). No difference was found between topical lanoline and saline (P> 0.05). The maximum ulcer diameter was obtained in 2 weeks. The maximum tissue malondialdehyde levels were obtained in 24 h, and in comparison to the control group the treatment groups showed lower levels. Our aim is to show the role of free radicals in the formation of skin necrosis as a cause of adriamycin extravasation and to prevent or decrease the skin necrosis using various free radical scavengers.

Effects of phorbol myristate acetate (PMA) on functions of macrophages and microglia in vitro.

Peripheral macrophages infiltrating the central nervous system and resident microglia phagocytize myelin in cell-mediated demyelinating diseases, including experimental autoimmune encephalomyelitis and multiple sclerosis. A cascade of cytokines is believed to modulate the immunological sequence of events occurring in these conditions, and several of these mediate their effects through the protein kinase C pathway. Therefore, we compared the effects of phorbol myristate acetate (PMA), an activator of protein kinase C, on various functions of cultured macrophages and microglia. PMA at moderate concentrations induced apoptosis in macrophages, and this process appeared to be increased in the presence of myelin. In contrast, microglia were activated by PMA, and greatly increased their phagocytosis of myelin. Control macrophages released a considerable amount of proteolytic activity into the medium, as measured by the breakdown of myelin basic protein, and in the process of undergoing apoptosis from PMA-treatment, even higher amounts were released. The enzyme activity in control macrophage medium was inhibited mainly by PMSF and calpain inhibitors, while that from PMA-treated macrophages was inhibited by calpain inhibitors only. An ICE inhibitor was ineffective in inhibiting activity in medium from PMA-treated cells undergoing apoptosis. Medium from microglia contained very little proteolytic activity, and this was not increased by PMA. Cultured macrophages showed little evidence of oxygen free radical release as measured by the TBARS procedure, and PMA had no effect. Microglia, on the other hand, produced higher levels of reactive oxygen species, with a further increase of 18% by PMA. Thus major functions of these phagocytic cells appear to be modulated by the protein kinase C pathway, although the two cell types show very different responses to an activator of this signal.

Inhibition of Transforming Growth Factor β1 Induction by Dietary Vitamin E in Unilateral Ureteral Obstruction in Rats

Free radical species associated with bilateral ureteral obstruction (BUO) are considered important in the pathogenesis of the glomerular and tubulointerstitial injury in BUO rats. We seek to test the hypothesis that the use of an easily administered antioxidant, vitamin E, at sufficient plasma concentrations, can decrease this release of free oxygen radicals in kidney tissue and ameliorate the increase of the fibrogenic cytokine, transforming growth factor β-1 (TGFβ-1). We used the unilateral ureteral obstruction (UUO) rat model, because the presence of the uninjured contralateral kidney provides a nonuremic internal milieu, in contrast to the uremic, acidotic, and hypercholesterolemic BUO model. Compared to sham controls, the UUO animals showed a dramatic increase in renal cortical TGFβ-1 mRNA, as quantitated by Northern blot analysis with cyclophilin internal standards. This increase in TGFβ-1 mRNA was reversed in UUO rats treated with vitamin E. The plasma malondialdehyde (MDA) concentration, an index of lipid peroxidation and an indirect index of free radical release, was significantly elevated in UUO animals compared to sham animals. The vitamin E-treated UUO animals showed a significant decrease in both plasma and renal cortical tissue MDA content. Taken together, these findings provide evidence of the important biological role of reactive free radical species in the tubulointerstitial injury of UUO and the novel role of vitamin E in modulating the mRNA of the fibrogenic TGFβ-1 in obstructive uropathy.

Plasma vitamin C concentrations in patients with cystic fibrosis: evidence of associations with lung inflammation

Vitamin C status and possible associations with the disease process in cystic fibrosis (CF) patients were investigated. Plasma vitamin C concentrations in patients from two different mid-European populations (Swiss, n = 62; Austrian, n = 60) taking no or low-dose vitamin C from multivitamin supplements did not differ from each other or from control subjects (n = 34). Vitamin C concentrations decreased with age (5.05 mumol.L-1, y-1). When followed up for 12 mo, patients had the highest plasma vitamin C concentrations in February and the lowest in May and August (P < 0.01); the decrease in vitamin C was accompanied by increases in plasma malondialdehyde (P < 0.001) and tumor necrosis factor alpha concentrations (P < 0.01). During supplementation with vitamin E for 2 mo or beta-carotene for 12 mo vitamin C concentrations did not change. They correlated inversely with white blood cell count (r = -0.36, P = 0.008), bands (r = -0.36, P = 0.02), alpha 1-acid glycoprotein (r = -0.45, P = 0.002), interleukin 6 (r = -0.46, P = 0.0006), and neutrophil elastase/alpha 1-proteinase inhibitor complexes (r = -0.34, P = 0.02). In patients with vitamin C concentrations < 40 mumol/L, all indexes of inflammation were relatively high, whereas those with concentrations > 80 mumol/L (upper quartile of control subjects) showed clearly lower values. These results are consistent with the hypothesis that by scavenging oxygen free radicals vitamin C interacts with an inflammation-amplifying cycle of activation of alveolar macrophages and neutrophils, release of proinflammatory cytokines and oxygen free radicals, and inactivation of antiproteases.