Paolo Murabito

Propofol attenuates peroxynitrite-mediated DNA damage and apoptosis in cultured astrocytes: an alternative protective mechanism.

Background:The concentration of peroxynitrite in the brain increases after central nervous system injuries. The authors hypothesized that propofol, because of its particular chemical structure, mitigates the effects of peroxynitrite-mediated oxidative stress and apoptosis by the induction of heme oxygenase (HO)-1 in primary cultured astroglial cells. Methods:Primary cultured astroglial cells were incubated for 18 h with a known peroxynitrite donor (3 mm SIN-1) in the presence or absence of propofol (40 &mgr;m, 80 &mgr;m, 160 &mgr;m, and 1 mm). The protective effects of propofol were evaluated by 3(4,5-dimethyl-thiazol-2-yl)2,5-diphenyl-tetrazolium bromide cytotoxicity assay, lactic dehydrogenase release, DNA ladderization by Comet assay, and caspase-3 activation by Western blot analysis. Results:Appropriate propofol concentrations (ranging from 40 &mgr;m to 1 mm) significantly increased HO-1 expression and attenuated SIN-1–mediated DNA ladderization and caspase-3 activation. The protective effects of propofol were mitigated by the addition of tin mesoporphyrin, a potent inhibitor of HO activity. The addition of a specific synthetic inhibitor of nuclear factor &kgr;B abolished propofol-mediated HO-1 induction, suggesting a possible role of this nuclear transcriptional factor in our experimental conditions. Conclusions:The antioxidant properties of propofol can be partially attributed to its scavenging effect on peroxynitrite as well as to its ability to increase HO-1 expression at higher concentrations, a property that might be relevant to neuroprotection during anesthesia.

Potential therapeutic effects of natural heme oxygenase-1 inducers in cardiovascular diseases.

SIGNIFICANCE Many physiological effects of natural antioxidants, their extracts or their major active components, have been reported in recent decades. Most of these compounds are characterized by a phenolic structure, similar to that of α-tocopherol, and present antioxidant properties that have been demonstrated both in vitro and in vivo. Polyphenols may increase the capacity of endogenous antioxidant defenses and modulate the cellular redox state. Such effects may have wide-ranging consequences for cellular growth and differentiation. CRITICAL ISSUES The majority of in vitro and in vivo studies conducted so far have attributed the protective effect of bioactive polyphenols to their chemical reactivity toward free radicals and their capacity to prevent the oxidation of important intracellular components. One possible protective molecular mechanism of polyphenols is nuclear factor erythroid 2-related factor (Nrf2) activation, which in turn regulates a number of detoxification enzymes. RECENT ADVANCES Among the latter, the heme oxygenase-1 (HO-1) pathway is likely to contribute to the established and powerful antioxidant/anti-inflammatory properties of polyphenols. In this context, it is interesting to note that induction of HO-1 expression by means of natural compounds contributes to prevention of cardiovascular diseases in various experimental models. FUTURE DIRECTIONS The focus of this review is on the role of natural HO-1 inducers as a potential therapeutic strategy to protect the cardiovascular system against various stressors in several pathological conditions.

Antioxidant potential of different melatonin-loaded nanomedicines in an experimental model of sepsis.

Oxidative stress has been shown to play a major role in the complex pathophysiological processes leading to organ failure during sepsis. The aim of the present research was to evaluate the effect of different melatonin nanoparticle (NP) carriers in an experimental animal model of sepsis. Poly-d,l-lactide-co-glycolide (PLGA [NP-A]) and polyethylene glycol-co-(poly-d,l-lactide-co-glycolide) (PLGA-PEG [NP-B]) were used to obtain melatonin-loaded nanocarriers (10 mg/kg). Oxidative stress was measured in tissue homogenates by measuring heme oxygenase-1 (HO-1) expression, total thiol groups and lipid hydroperoxides (LOOH). In vitro NPs showed a long lag time followed by a controlled release of melatonin. All the different melatonin formulations restored total thiol group levels to those of controls in all the examined organs, with no significant changes among them. Both melatonin NP formulations significantly decreased LOOH levels when compared with sepsis vehicle animals. The stealth formulation NP-B was able to produce a more significant reduction in LOOH levels in the heart, lung and liver when compared with NP-A. No significant changes were observed between the two NP formulations in the kidney. Interestingly, HO-1 expression was differently affected following treatment with various melatonin formulations. The NP-B formulation was more effective in inducing HO-1 protein compared with free melatonin and NP-A, with the exception of the kidney. Taken together, our results show that melatonin possesses a significant antioxidant activity during sepsis and that it is possible to improve this ability by delivering the compound with specific drug delivery systems.

Effect of ischemia-reperfusion on renal expression and activity of N(G)-N(G)-dimethylarginine dimethylaminohydrolases.

Background:Asymmetric dimethylarginine (ADMA) is an endogenous inhibitor of nitric oxide synthase. It is degraded by the enzyme dimethylarginine dimethylaminohydrolase (DDAH). Methods:Rats (n = 50) underwent to 45 min of renal ischemia followed by 30 min, 1 h, and 3 h of reperfusion. Expression of endothelial nitric oxide synthase, inducible nitric oxide synthase, DDAH-1, DDAH-2, renal DDAH activity, plasma NO2−/NO3−, and ADMA levels were evaluated. Results:Inducible nitric oxide synthase expression increased, as confirmed by both plasma (11.89 ± 1.02, 15.56 ± 0.93, 11.82 ± 0.86, 35.05 ± 1.28, and 43.89 ± 1.63 nmol/ml in the control, ischemic, 30-min, 1-h, and 3-h groups, respectively) and renal (4.81 ± 0.4, 4.85 ± 1, 9.42 ± 0.7, 15.42 ± 0.85, and 22.03 ± 1.11 nmol/mg protein) formations of NO2−/NO3−. DDAH-1 expression decreased after reperfusion, whereas DDAH-2 increased after 30 min, returning to basal levels after 3 h. Total DDAH activity was reduced during all times of reperfusion. Both plasma (0.41 ± 0.03, 0.43 ± 0.05, 0.62 ± 0.02, 0.71 ± 0.02, and 0.41 ± 0.01 nmol/ml in the control, ischemic, 30-min, 1-h, and 3-h groups, respectively) and renal (1.51 ± 0.01, 1.5 ± 0.01, 1.53 ± 0.01, 2.52 ± 0.04, and 4.48 ± 0.03 nmol/mg protein in the control, ischemic, 30-min, 1-h, and 3-h groups, respectively) concentrations of ADMA increased. Conclusions:Results suggest that ischemia–reperfusion injury leads to reduced DDAH activity and modification of different DDAH isoform expression, thus leading to increased ADMA levels, which may lead to increased cardiovascular risk.

(+)-Pentazocine reduces oxidative stress and apoptosis in microglia following hypoxia/reoxygenation injury

BACKGROUND Sigma-1 receptors (σ1R) are highly expressed in neurons as well as microglia and have been shown to modulate the inflammatory response in the central nervous system and thus may serve as possible target for neuroprotective strategies. The aim of the present study was to test the effect of (+)-pentazocine, a putative σ 1R agonist, in an in vitro model of microglia activation. METHODS Microglia (BV2 cells) was exposed (3h) to 1% oxygen and reoxygenation was allowed for 24h. Cells were treated with different concentrations (1, 10, 25 and 50μM) of (+)-pentazocine in the presence or absence of NE-100 (1μM), a well established σ1R antagonist. Cell viability and apoptosis were measured by cytofluorimetric analysis, whereas oxidative stress was evaluated by reduced glutathione (GSH) content and mitochondrial potential analysis. RESULTS Our results showed that (+)-pentazocine was able to increase cell viability and restore mitochondrial potential at all concentrations whereas only 1 and 10μM were able to reduce significantly apoptotic cell death, to restore reduced glutathione intracellular content and prevent ERK1/2 phosphorylation. All these effects were abolished by concomitant treatment with NE-100. CONCLUSIONS (+)-pentazocine exhibits significant dose dependent protective effects in our in vitro model of microglial activation thus suggesting that σ1R may represent a possible target for neuroprotection.

Pharmacologic induction of heme oxygenase-1: it is time to take it seriously*.

www.ccmjournal.org 1967 Propofol is an IV sedative-hypnotic agent commercially introduced in the United States in 1989 by Zeneca Pharmaceuticals. It was the first of a new class of IV anesthetic agents: the alkylphenols. It is indicated for induction and maintenance of general anesthesia as well as for sedation of intubated, mechanically ventilated adults in the ICU. Propofol is characterized by a phenolic structure similar to that of α-tocopherol and presents antioxidant properties that have been demonstrated in vitro and in vivo. Propofol has been reported to inhibit lipid peroxidation in various experimental models (1), to protect cells against oxidative stress, and to increase the antioxidant capacity of plasma in humans (2). Furthermore, we suggested a possible novel aspect in the mode of action of propofol, that is, the ultimate stimulation of the heme oxygenase (HO)-1 pathway is likely to account for the established and powerful antioxidant/anti-inflammatory properties of this compound (3). The study by Xu et al (4) in this issue of Critical Care Medicine evaluated the impact of propofol treatment on hyperglycemia-induced oxidative stress in cardiomyocytes and how HO-1 regulates such mechanism. HO isoforms catalyze the conversion of heme to carbon monoxide (CO) and biliverdin with a concurrent release of iron, which can drive the synthesis of ferritin for iron sequestration (5). HO is the physiologic pathway of heme degradation and, consequently, plays a critical role in the regulation of cellular heme-dependent enzyme concentrations (6). To date, two HO isoforms have been shown to be catalytically active in heme degradation, and each is encoded by a different gene. HO-1, the inducible isoform, is found ubiquitously and transiently expressed after exposure to various stressors (5). It is now widely accepted that induction of HO-1 expression represents an adaptive response that critically increases cell resistance to oxidative injury, and this pathway plays a critical role in various pathological conditions (7). By contrast, HO-2 is responsible for most HO constitutive activity and does not undergo modifications during most experimental conditions. Xu et al (4) showed that type 1 diabetic rats exhibited a significant increase of oxidative stress and myocardial damage as measured by 15-F2t-isoprostane, cardiomyocyte hypertrophy, apoptosis, and impaired left ventricular function. Interestingly, these findings were associated with reduced HO-1 activity and signal transducer and activator of transcription 3 (STAT3) activation despite an increase in HO-1 protein expression compared with control. Propofol infusion (900 μg/ kg/min) for 45 minutes significantly improved cardiac function with concomitantly enhanced HO-1 activity and STAT3 activation. Interestingly, propofol significantly elevated HO-1 activity in the diabetic heart despite only a slight increase in HO-1 protein expression. Furthermore, propofol did not have significant impact on either HO activity or HO-1 protein expression in control hearts. These results regarding HO-1 expression/HO activity may seem discrepant for the readers when analyzed at a first look. To this regard, authors’ data are consistent with previous studies showing that under certain conditions (i.e., hyperglycemia), there exists an uncoupling between HO-1 expression and HO activity (8). The clear significance of such phenomenon is not fully understood. It is possible to speculate that some posttranscriptional modification may occur under hyperglycemic experimental conditions. Little is known about the potential regulation of HO-1 by posttranslational modifications, although there is increasing appreciation that structural modifications, for example, truncation, underpin noncanonical functions of HO-1 (9). An in silico analysis of the human HO-1 protein predicts a number of potential sites for posttranslational modifications such as acetylation, ubiquitination, palmitoylation, and phosphorylation (9). In addition, peroxynitrite (ONOO) may also regulate HO activity by mechanisms that involve different interactions with this enzyme (10). ONOO, which results from the reaction between nitric oxide and superoxide anion, is an oxidant that can react with proteins, lipids, carbohydrates, and nucleic acids. Previous studies showed that propofol reacts with peroxynitrite, leading to the formation of a propofol-derived phenoxyl radical, and has therefore been hypothesized to be a peroxynitrite scavenger. Therefore, propofol itself, together with its ability to up-regulate HO-1, may be responsible for the increased HO activity. The study by Xu et al (4) concluded that the induction of HO-1/STAT3 Copyright

Sugammadex Use in the Reversal of Deep Neuromuscular Block in a Six-Year-Old Child after an Emergency Procedure in Ear, Nose and Throat Surgery Department

Tonsillectomy is a frequently performed procedure in children; its common postoperative complication is bleeding. We present a case report of a 6-year-old patient undergoing surgical haemostasis after an acute post-tonsillectomy bleeding. A rapid intubation sequence was performed to reduce the risk of blood inhalation and bronchospasm: fentanyl (1 mcg/kg) and propofol (3mg/Kg) were used for a rapid induction of anaesthesia, and rocuronium (1.2 mg/Kg) was used for neuromuscular block. Neuromuscular function was monitored using the TOF-Watch SX acceleromyography at the adductor thumb muscle. Surgical haemostasis lasted for 12 minutes after rocuronium administration, so the neuromuscular block was still deep (TOF 0, PTC 2). Sugammadex (4mg/Kg) was administered to allow a rapid reversal of the block. After the sugammadex bolus, we started a continuous monitoring with TOF-Watch SX until a TOF ratio of 0.9 was reached. A TOF ratio of 0.9 was reached 3 minutes after the administration of sugammadex. The patient was then extubated in the operating room and monitored in recovery room for 60 minutes. No further complications arose, either in the recovery room or in the ward.

Antioxidant Properties of Propofol

Propofol is an intravenous sedative-hypnotic agent indicated for induction and maintenance of general anesthesia as well as for sedation of intubated, mechanically ventilated adults in intensive care units (ICU). Propofol is characterized by a phenolic structure similar to that of α-tocopherol, and presents antioxidant properties that have been demonstrated both in vitro and in vivo. The aim of the present review is to evaluate the antioxidant properties of propofol in various models and whether or not it may be considered an efficient therapeutic tool in counteracting oxidative stress during general anesthesia and sedation in ICU.

Sigma-1 receptor and neuroprotection: current outlook and potential therapeutic effects

Oxidative stress plays a major role in neurodegenerative disease since central nervous system is particularly vulnerable to reactive oxygen species (ROS) due to several reasons: high consumption of O2; high production of ROS and nitrosative species, which originate from specific neurochemical reactions (e.g., dopamine oxidation); high deposition of metal ions in the brain with aging leading to Fentons reactions; high abundance of lipids which are particularly sensitive to oxidation. Therefore, we evaluated several pharmacological strategies directed at investigating the role of various antioxidants to prevent or treating neurodegenerative disorders in various experimental models (Caccamo et al., 2004; Campisi et al., 2004). However, despite several studies performed both in vitro and in vivo have shown promising results, none of them appear to be of great clinical significance. One possible explanation for such pharmacological profile of antioxidants may be dependent on the complex biochemical cascade underlying neuronal injury. In fact, oxidative stress represents just one of several mechanisms triggered by the pathogenic noxa and it may occur also as a late mechanism of injury. Therefore, several other strategies have been developed in order to obtain a broad range of effects, including the antioxidant effect, and which may also impact on early mechanisms underlying neuronal degeneration. Among such pharmacological strategies possessing pleiotropic effects, sigma receptors seem to play a major role.

Anesthetic Management in Mucopolysaccharidoses

Mucopolysaccharidoses (MPSs) are a group of uncommon genetic diseases of connective tissue metabolism. It is well established that the elective treatment of subjects affected by MPS is multidisciplinary and must be carried out by experienced personnel in highly specialist centers. However, there is the possibility to perform an anesthesia in a peripheral center, where anesthesiologists might not have a large experience of MPS. Various attempts to produce guidelines have been made for MPS. There has been an increasing interest in the topic as it is demonstrated by the number of scientific studies published in the last few years (peak in 2011). We want to provide a summary of anesthetic management, reviewing the current literature on the topic in a procedural algorithm for these high-risk patients, who require surgical procedures and diagnostic examinations under sedation with a higher frequency than the general population.