Oxygen Radical Production Research Articles

Reactive Oxygen Species and Antioxidant Defense in Plants under Abiotic Stress: Revisiting the Crucial Role of a Universal Defense Regulator.

Global climate change and associated adverse abiotic stress conditions, such as drought, salinity, heavy metals, waterlogging, extreme temperatures, oxygen deprivation, etc., greatly influence plant growth and development, ultimately affecting crop yield and quality, as well as agricultural sustainability in general. Plant cells produce oxygen radicals and their derivatives, so-called reactive oxygen species (ROS), during various processes associated with abiotic stress. Moreover, the generation of ROS is a fundamental process in higher plants and employs to transmit cellular signaling information in response to the changing environmental conditions. One of the most crucial consequences of abiotic stress is the disturbance of the equilibrium between the generation of ROS and antioxidant defense systems triggering the excessive accumulation of ROS and inducing oxidative stress in plants. Notably, the equilibrium between the detoxification and generation of ROS is maintained by both enzymatic and nonenzymatic antioxidant defense systems under harsh environmental stresses. Although this field of research has attracted massive interest, it largely remains unexplored, and our understanding of ROS signaling remains poorly understood. In this review, we have documented the recent advancement illustrating the harmful effects of ROS, antioxidant defense system involved in ROS detoxification under different abiotic stresses, and molecular cross-talk with other important signal molecules such as reactive nitrogen, sulfur, and carbonyl species. In addition, state-of-the-art molecular approaches of ROS-mediated improvement in plant antioxidant defense during the acclimation process against abiotic stresses have also been discussed.

Possible mechanisms responsible for acute coronary events in COVID-19

The novel coronavirus (SARS-CoV-2) is primarily a respiratory pathogen and its clinical manifestations are dominated by respiratory symptoms, the most severe of which is acute respiratory distress syndrome (ARDS). However, COVID-19 is increasingly recognized to cause an overwhelming inflammatory response and cytokine storm leading to end organ damage. End organ damage to heart is one of the most severe complications of COVID-19 that increases the risk of death. We proposed a two-fold mechanism responsible for causing acute coronary events in patients with COVID-19 infection: Cytokine storm leading to rapid onset formation of new coronary plaques along with destabilization of pre-existing plaques and direct myocardial injury secondary to acute systemic viral infection. A well-coordinated immune response is the first line innate immunity against a viral infection. However, an uncoordinated response and hypersecretion of cytokines and chemokines lead to immune related damage to the human body. Human Coronavirus (HCoV) infection causes infiltration of inflammatory cells that cause excessive production of cytokines, proteases, coagulation factors, oxygen radicals and vasoactive molecules causing endothelial damage, disruption of fibrous cap and initiation of formation of thrombus. Systemic viral infections also cause vasoconstriction leading to narrowing of vascular lumen and stimulation of platelet activation via shear stress. The resultant cytokine storm causes secretion of hypercoagulable tissue factor without consequential increase in counter-regulatory pathways such as AT-III, activated protein C and plasminogen activator type 1. Lastly, influx of CD4+ T-cells in cardiac vasculature results in an increased production of cytokines that stimulate smooth muscle cells to migrate into the intima and generate collagen and other fibrous products leading to advancement of fatty streaks to advanced atherosclerotic lesions. Direct myocardial damage and cytokine storm leading to destabilization of pre-existing plaques and accelerated formation of new plaques are the two instigating mechanisms for acute coronary syndromes in COVID-19.

Adelmidrol: A New Promising Antioxidant and Anti-Inflammatory Therapeutic Tool in Pulmonary Fibrosis.

Background: Chronic pulmonary diseases are characterized by airway remodeling due to complex multicellular responses and the production of free oxygen radicals. They lead to a progressive decline of pulmonary functions. Adelmidrol is an analogue of palmitoylethanolamide (PEA), which is a well-known anti-inflammatory and anti-oxidant compound. In this study, we investigated the efficacy of adelmidrol (10 mg/Kg) for bleomycin-induced pulmonary fibrosis in mice. Methods: Bleomycin intratracheal administration was performed on the first day and for the following twenty-one days, mice were treated with adelmidrol (10 mg/Kg). Results: The survival rate and body weight gain were recorded daily. At the end of the experiment, adelmidrol-administered animals showed reduced airway infiltration by inflammatory cells, Myeloperoxidase (MPO) activity, and pro-inflammatory cytokine overexpression (IL,6 IL-1β, TNF-α, and TGF-1β). Moreover, adelmidrol treatment was able to manage the significant incapacity of antioxidants and elevation of the oxidant burden, as shown by the MDA, SOD, and GSH levels and decreased nitric oxide production. It was also able to significantly modulate the JAK2/STAT3 and IκBα/NF-kB pathway. Histologic examination of the lung tissues showed reduced sample injury, mast cell degranulation, chymase activity, and collagen deposition. Conclusions: In sum, our results propose adelmidrol as a therapeutic approach in the treatment of pulmonary fibrosis.

Simultaneous Electrochemical Generation of Ferrate and Oxygen Radicals to Blue BR Dye Degradation

In this study, electro-oxidation (EOx) and in situ generation of ferrate ions [Fe(VI)] were tested to treat water contaminated with Blue BR dye (BBR) using a boron-doped diamond (BDD) anode. Two electrolytic media (0.1 M HClO4 and 0.05 M Na2SO4) were evaluated for the BDD, which simultaneously produced oxygen radicals (•OH) and [Fe(VI)]. The generation of [Fe(VI)] was characterized by cyclic voltammetry (CV) and the effect of different current intensity values (e.g., 7 mA cm−2, 15 mA cm−2, and 30 mA cm−2) was assessed during BBR degradation tests. The discoloration of BBR was followed by UV-Vis spectrophotometry. When the EOx process was used alone, only 78% BBR discoloration was achieved. The best electrochemical discoloration conditions were found using 0.05 M Na2SO4 and 30 mA cm−2. Using these conditions, overall BBR discoloration values up to 98%, 95%, and 87% with 12 mM, 6 mM, and 1 mM of FeSO4, respectively, were achieved. In the case of chemical oxygen demand (COD) reduction, the EOx process showed only a 37% COD reduction, whereas combining [Fe(VI)] generation using 12 mM of FeSO4 achieved an up to 61% COD reduction after 90 min. The evolution of reaction byproducts (oxalic acid) was performed using liquid chromatography analysis.

P0010INCREASED SERUM URIC ACID LEVELS ARE ASSOCIATED TO RENAL ARTERIOLOPATHY AND PREDICT POOR OUTCOME IN IGA NEPHROPATHY

Abstract Background and Aims IgA nephropathy (IgAN) is the most common primary glomerulonephritis worldwide and a leading cause of End Stage Renal Disease (ESRD). In addition to the classical progression factors, including hypertension, proteinuria, and decreased renal function, other atherosclerosis-related factors, such as hypertriglyceridemia, have been reported to the renal progression of IgAN. Above all, many studies have indicated an association between hyperuricemia and progression of IgAN. Increased serum uric acid (SUA) levels are also well known to be associated with hypertension, endothelial dysfunction and development of cardiovascular and kidney diseases both in general population and high-risk patients. Previous studies suggested that uric acid induces proliferation of vascular smooth muscle cells, activates the renin–angiotensin–aldosterone system (RAAS) and also reduces the synthesis of nitric oxide. Furthermore, there is evidence that uric acid can promote the oxygenation of low-density lipoproteins (LDL) and increase the production of free oxygen radicals, increasing inflammation and oxidative stress. In consequence of these processes, SUA might specifically promote atherosclerosis and arteriolar damage, possibly playing a major role in pathogenesis and progression of IgAN. The aim of the present study was to explore the correlation between SUA levels, renal damage and its implication for renal outcome and all cause death in IgAN patients. Method The data for clinical features, laboratory and renal pathological examination were collected from 145 renal biopsy-proven IgAN patients and were retrospectively analyzed to determine the correlation between SUA levels, renal damage and overall outcome. Hyperuricemia (HU) was defined as the highest SUA gender-specific tertile. Biopsy-proven arteriolar damage was defined by the presence of arteriolar hyalinosis or intimal thickening. The primary outcome was death or ESRD. Results The mean baseline serum uric acid levels of the 42 female and 103 male patients at the time of kidney biopsy were 5.4 ±1.7 and 7.2 ±1.8 mg/dL, respectively. HU was >7.7 mg/dl for males and > 6.2 mg/dl for females. Patients were stratified on the basis of baseline gender-specific SUA level tertiles. Clinical and histologic characteristics are described in table 1. The higher the gender-specific SUA tertile, the greater the prevalence of arteriolar damage (p=0.02). No other histologic feature was significantly correlated with uric acid levels, therefore we analyzed laboratory and clinical characteristics on the basis of the presence/absence of biopsy-proven arteriolar damage (Table 2). At logistic regression analyses SUA was associated with arteriolar damage at univariate (OR 1.45 CI [1.12-1.87], p=0.004) and multivariate analyses (OR 1.75 CI [1.10-2.93], p=0.03), recorded in Table 3. Receiver Operating Curve (ROC) was performed and the sensitivity and specificity of SUA for predicting arteriolar damage were showed in Figure 1. The area under the ROC curve was 0.67 (IC95% 0.61 to 0.73) indicating that SUA is a fair test for detecting arteriolar damage. Patients with arteriolar damage had a worst outcome compared to patients without it (Kaplan Maier survival analysis, log rank test p=0.002, Figure 2a). HU and arteriolar damage had a synergic impact on progression of IgAN. Patients having both arteriolar damage and HU, showed a reduced survival free from the primary outcome as compared to those having only one risk factor or neither (log rank test p=0.003, Figure 2b). Conclusion SUA levels are directly associated with arteriolar damage and poor prognosis in patients with IgAN. Our study suggests the presence of higher SUA levels in IgAN patients identifies a sub-population at increased risk of progressing to ESRD or death, for whom a particular surveillance is warranted, possibly because of the pathogenetic role of SUA on arteriolar damage.

P0622FIGHTING AGAINST PROXIMAL AND DISTAL TUBULAR DAMAGE BY REDUCING INFLAMMATORY BURDEN AFTER CARDIAC SURGERY

Abstract Background and Aims Going on cardiopulmonary bypass (CPB) during cardiac surgery causes systemic inflammatory response syndrome. In different studies Pentoxifylline and Calcitriol have shown inhibitory effects on pro-inflammatory cytokines such as tumor necrosis factor (TNF) and interleukin-6 (IL-6) accompanying with production of free oxygen radicals in the ischemic reperfusion injury. On the other hand, they have positive effects on reducing oxidative stress. To investigate the effects of these drugs on reducing tubular injury after CABG surgery, by their anti-inflammatory properties we compared pre- and postoperative levels of urinary NGAL (as a proximal tubular damage marker), KIM-1 level (as a distal tubular damage marker), serum level of Malondialdehyde (MDA) (as an oxidative stress marker) and serum inflammatory markers such as IL-6, IL-8, and TNF levels in two groups of patients undergoing CABG, those who received Pentoxifylline and Calcitriol before surgery and those who didn't Method After signing an informed consent, 150 consecutive adult patients undergoing elective on-pump CABG were enrolled in a randomized control trial. Patients with collagen-vascular disease or having a history of treatment with immunosuppressive agents, corticosteroids (> 3 days), Methylxantines, Diltiazem or sodium Nitroprusside and patients with a history of angiography in the past 7 days or hemorrhagic diathesis and coagulopathy, uncontrolled diabetes mellitus, sepsis, renal failure (sCr > 2 mg/dl), hepatic failure (AST or ALT > 40 U/L) or urinary tract infection were excluded. All patients received the same anesthetic regimen and routine CPB management. The patients were randomly assigned to one of two groups: PTX was administered as an oral 400 mg tab every 12hours plus IV Calcitriol (0.01 mcg/kg) daily starting from 3 days before surgery. In the control group placebo tab and IV injection of normal saline were prescribed in the same direction. Results Results demonstrated that after three days prophylactic use of oral pentoxifylline plus IV Calcitriol there is improvement in selected checked inflammatory markers in the intervention group. Moreover Pentoxifylline+ Calcitriol showed beneficial effects by controlling the oxidative stress burden and reducing the mean serum MDA level in the patients undergoing CABG who are at risk of developing AKI as a post surgery complication. Probably by reducing inflammation and oxidative stress combination of these two drugs before CABG could decrease proximal and distal tubular damage which presented with significantly lower levels of uNGAL and KIM-1, as markers of tubular injury, in the intervention group. Conclusion Combination of Pentoxifylline and Calcitriol showed a promising prophylactic role in patients undergoing CABG with anti-inflammatory, anti-oxidant properties. This study provided scope of add-on drug therapy for better prognosis in post CABG population, involving risk of endothelial dysfunction and renal tubular damage due to inflammatory milieu and renders support to future prospective clinical studies to demonstrate net control of inflammation and oxidative stress after cardiac surgery.

Energy-transfer-mediated oxygen activation in carbonyl functionalized carbon nitride nanosheets for high-efficient photocatalytic water disinfection and organic pollutants degradation

Polymeric photocatalysts are promising candidates for water purification, however their catalytic performance are still unsatisfactory due to the fast charge recombination that leads to low reactive oxygen radicals production. In this study, a conceptual energy-transfer-mediated photocatalytic oxygen activation system over polymeric carbon nitride without the need of electron-hole separation is proposed, exhibiting remarkable singlet oxygen triggered bacteria inactivation performance as well as organic pollutants degradation. By structure and excitonic effect modulation, the oxygen activation process changes from the traditional electron-transfer mechanism to the final energy-transfer pathway, leading to the selective generation of singlet oxygen with high efficiency. The generated singlet oxygen is found to fervently attack the bacteria membrane, creating irreparable pores or holes on the cell membrane for cytoplasmic contents leaking out to accelerate bacteria destruction. The work demonstrated here offers a new photocatalytic oxygen activation pathway for achieving high-efficient reactive oxygen species generation performance without the need of charge separation.

P2X7 receptor as an independent prognostic indicator in gastric cancer.

Gastric cancer (GC) is one of the foremost causes of cancer-related death around the world. The P2X7 receptor (P2X7R), a member of the P2X7R subfamily of P2 receptors, is a unique molecule that has been shown to affect tumor growth and progression as well as various inflammatory processes, including proliferation of T lymphocytes, release of cytokines, and production of free oxygen radicals. P2X7R has been established as a prognostic parameter in some cancers, and recently, it has been investigated in the development of new targeted therapies. In the present study, we aimed to investigate the prognostic value of P2X7R expression in GC. The expression profile of P2X7R was evaluated immunohistochemically in 156 paraffin-embedded human GC specimens. P2X7R expression was higher in patients with lymph node metastasis than in those without (p < 0.001). P2X7R overexpression was closely related with tumor-infiltrating lymphocytes (TILs) (p = 0.001), vascular invasion (p = 0.006), depth of invasion (p < 0.001), distant metastasis (p < 0.001), and advanced tumor, node, metastasis stage (p < 0.001). Moreover, univariate (hazard ratio [HR] 3.98; 95% confidence interval (CI) 1.89-11.82; p < 0.001) and multivariate (HR 2.24; 95% CI 3.53-12.50; p < 0.001) Cox regression analysis showed that upregulated P2X7R expression clearly correlated with worsened overall survival. In summary, our data revealed that P2X7R may serve as a reliable prognostic parameter and promising therapeutic target for GC.

6\u03b2-Hydroxytestosterone, a metabolite of testosterone generated by CYP1B1, contributes to vascular changes in angiotensin II-induced hypertension in male mice

BackgroundPreviously, we showed that 6β-hydroxytestosterone (6β-OHT), a cytochrome P450 1B1 (CYP1B1)-derived metabolite of testosterone, contributes to angiotensin II (Ang II)-induced hypertension in male mice. This study was conducted to test the hypothesis that 6β-OHT contributes to increased vascular reactivity, endothelial dysfunction, vascular hypertrophy, and reactive oxygen species production associated with Ang II-induced hypertension.MethodsEight- to 10-week-old intact or castrated C57BL/6 J (Cyp1b1+/+ and Cyp1b1−/−) mice were anesthetized for implantation of a micro-osmotic pump which delivered Ang II (700 ng/kg/day) or saline for 14 days. Mice were injected with 6β-OHT (15 μg/g b.w every third day), flutamide (8 mg/kg every day), or its vehicle. Blood pressure was measured via tail-cuff. Vascular reactivity, endothelial-dependent and endothelial-independent vasodilation, media to lumen ratio, fibrosis by collagen deposition, and reactive oxygen species production by dihydroethidium staining were determined in the isolated thoracic aorta.ResultsThe response of thoracic aorta to phenylephrine and endothelin-1 was increased in Ang II-infused Cyp1b1+/+ mice compared to intact Cyp1b1−/− or castrated Cyp1b1+/+ and Cyp1b1−/− mice; these effects of Ang II were restored by treatment with 6β-OHT. Ang II infusion caused endothelial dysfunction, as indicated by decreased relaxation of the aorta to acetylcholine in Cyp1b1+/+ but not Cyp1b1−/− or castrated Cyp1b1+/+ and Cyp1b1−/− mice. 6β-OHT did not alter Ang II-induced endothelial dysfunction in Cyp1b1+/+ mice but restored it in Cyp1b1−/− or castrated Cyp1b1+/+ and Cyp1b1−/− mice. Ang II infusion increased media to lumen ratio and caused fibrosis and reactive oxygen species production in the aorta of Cyp1b1+/+ mice. These effects were minimized in the aorta of Cyp1b1−/− or castrated Cyp1b1+/+ and Cyp1b1−/− mice and restored by treatment with 6β-OHT. Treatment with the androgen receptor antagonist flutamide reduced blood pressure and vascular hypertrophy in castrated Ang II-infused mice injected with 6β-OHT.Conclusions6β-OHT is required for the action of Ang II to increase vascular reactivity and cause endothelial dysfunction, hypertrophy, and increase in oxygen radical production. The effect of 6β-OHT in mediating Ang II-induced hypertension and associated hypertrophy is dependent on the androgen receptor. Therefore, CYP1B1 could serve as a novel target for the development of therapeutics to treat vascular changes in hypertensive males.

Antioxidant variability of wheat genotypes under salinity stress in situ

Crop cultivation under the open-field conditions depends on a variety of biotic and abiotic factors which cause plant stress and deterioration. Due to high soil salinity, some soils can be an unfavourable growing environment for most plants. As a low productivity soil type, solonetz soils are a stressful growing environment, causing plant deterioration. Plants have developed a complex antioxidative defense system as a precaution against oxidative stress caused by high soil salinity. The trial was set up on a halomorphic soil type-solonetz. The research included ten cultivars, one local population of hexaploid wheat (Triticum aestivum ssp. aestivum L.) and one cultivar of triticale (Triticosecale W.). The activity of enzymatic and non-enzymatic antioxidants in plant antioxidative defense system was detected during the trial, as well as lipid peroxidation. The analysis of biochemical markers was done in the flowering stage, and then in the milk maturity phase. Research results of the tested components revealed the highest stress tolerance exhibited by genotypes Banatka and Bankut 1205. Understanding the process of oxygen radical production by the plant tissue contributes to breeding wheat cultivars for better stress tolerance. Selection of genotypes better adapted to growing conditions in solonetz soils could facilitate a more economically justifiable wheat production, and promote utilization of the lower-quality soil types in agriculture.

Mechanisms of Anthracycline-Enhanced Reactive Oxygen Metabolism in Tumor Cells.

In this investigation, we examined the effect of anthracycline antibiotics on oxygen radical metabolism in Ehrlich tumor cells. In tumor microsomes and nuclei, doxorubicin increased superoxide anion production in a dose-dependent fashion that appeared to follow saturation kinetics; the apparent Km and Vmax for superoxide formation by these organelles was 124.9 μM and 22.6 nmol/min/mg, and 103.4 μM and 4.8 nmol/min/mg, respectively. In both tumor microsomes and nuclei, superoxide formation required NADPH as a cofactor, was accompanied by the formation of hydrogen peroxide, and resulted from the transfer of electrons from NADPH to the doxorubicin quinone by NADPH:cytochrome P-450 reductase (NADPH:ferricytochrome oxidoreductase, EC 1.6.2.4). Anthracycline antibiotics also significantly enhanced superoxide anion production by tumor mitochondria with an apparent Km and Vmax for doxorubicin of 123.2 μM and 14.7 nmol/min/mg. However, drug-stimulated superoxide production by mitochondria required NADH and was increased by rotenone, suggesting that the proximal portion of the electron transport chain in tumor cells was responsible for reduction of the doxorubicin quinone at this site. The net rate of drug-related oxygen radical production was also determined for intact Ehrlich tumor cells; in this system, treatment with doxorubicin produced a dose-related increase in cyanide-resistant respiration that was enhanced by changes in intracellular reducing equivalents. Finally, we found that in the presence of iron, treatment with doxorubicin significantly increased the production of formaldehyde from dimethyl sulfoxide, an indication that the hydroxyl radical could be produced by intact tumor cells following anthracycline exposure. These experiments suggest that the anthracycline antibiotics are capable of significantly enhancing oxygen radical metabolism in Ehrlich tumor cells at multiple intracellular sites by reactions that could contribute to the cytotoxicity of this class of drugs.

Using natural honey as an anti-oxidant and thermodynamically efficient cryoprotectant in embryo vitrification

Embryo cryopreservation is a common practice in reproductive biology and infertility treatments. Despite major improvements over years, the cryoprotectant solutions are still a major source of concern, mostly due to their chemical toxicity and suboptimal protection against cryoinjuries. In this work, we introduced natural honey as a non-permeating cryoprotectant to replace traditionally used sucrose in embryo vitrification. The proposed media were compared with conventional ones by evaluating vitrified/warmed mouse embryos based on their re-expansion, hatching rate and transcription pattern of selected genes involved in heat-shock response, apoptosis and oxidative stress. Despite the similar high re-expansion rate, molecular fingerprint of the cryopreservation is remarkably reduced when honey is used instead of sucrose. The biological response of the proposed media was explained from a fundamental point of view using antioxidant analysis, DSC and GC techniques. It was found that the proposed honey-based medium is less thermodynamically prone to ice formation, which along with its antioxidant capacity can control the production of oxygen radicals and minimize the stress-induced transcriptional response. Furthermore, this work tries to correlate the physico-chemical properties of the vitrification solutions with the cellular and molecular aspects of the cryopreservation and proposes the application of natural cryoprotectants in cryobiology.

Numerical modelling of plasmachemistry induced by cold atmospheric plasma jet near surface

Recently the cold plasma jets are widely used for the development of anticancer therapy. The main mechanism of atmospheric plasma jet action on cells is related to the production of nitrogen and oxygen radicals in plasma-stimulated chemical reactions, followed by the diffusion of radicals into the media with cells. The increase of the plasma jet treatment efficiency depends on a detailed study of the plasma-chemical processes that occur when the plasma jet interacts with the target. In this work, a numerical study of the plasma-chemical processes for dry air is carried out, the main ways of production and loss of RONS are detected and mixture composition near the surface during plasma jet processing is estimated.

Kinetics and reaction mechanism of NO removal from simulated flue with peroxymonosulfate solution activated by high temperature and metal ions ([formula omitted] and [formula omitted

A strategy for removing nitric oxide (NO) from simulated flue gas by transition metal ion (Fe2+,Co2+, and Fe3+)-induced production of sulfate (SO4−), hydroxyl (OH), and singlet oxygen radicals (1O2) from peroxymonosulfate (PMS) at high temperature was investigated, and the catalytic performances of the three metal ions were compared. The nitric oxide radical (NO) was detected in solution using electron spin-resonance (ESR) spectroscopy, and a pathway for reaction between NO and the radicals was further explored and refined. The metal ion-catalyzed decomposition of PMS could be considered a pseudo-first order reaction with good reliability (R2 > 0.97), and the rate constant, k0, for metal ion-catalyzed PMS degradation followed the sequence: Fe2+>Co2+>Fe3+. Specific experiments were optimized to select the optimal experimental parameters (PMS/metal molar ratio and solution pH) for maximizing the NO removal efficiency. ESR was employed to monitor the progress of PMS activation, and it was found that: 1) SO4−, OH, and 1O2 radicals were mainly produced during the activation of PMS by the three metal ions; 2) the three radicals synergistically removed NO; 3) the generation of 1O2 was more sensitive to temperature than the generation of the SO4− and OH radicals; 4) the capacity of the metals to catalyze the oxidation reaction followed the order: Fe2+>Co2+>Fe3+. These findings provide new insight into PMS activation by transition metal catalysts at high temperature.

Analysis of differences in the stability of proteins encoded in mitochondrial DNA of model organisms

The aim of this study was to describe the stability of proteins encoded in mtDNA, which are part of the OXPHOS system, in different model organisms and to define why certain proteins are more prone to be unstable than others. The in silico analyses involved 155 reference sequences of all proteins encoded in the mitochondrial DNA in twelve model organisms representing different phylogenetic groups. The amino acid sequences of the proteins were taken from the GenPept database. The bioinformatic analyses were performed in the ProtParam program. Thirty-eight of the 155 analyzed proteins exhibited instability. The greatest numbers of unstable mitochondrial proteins were detected in H. sapiens and A. mexicanum and the lowest levels were found in C. elegans. ND1 and ATP8 were the most unstable mitochondrial proteins. Proteins COX1 and COX3 did not exhibit instability in the examined group of organisms. The highest instability index values were recorded in the case of protein ATP8. Protein ND1 turned out to be stable in the representatives of the class invertebrates. The preliminary results of the pioneer investigations indicate that the type and number of unstable proteins encoded in mtDNA was species specific. Protein instability in lower organisms may be associated with resistance to oxidative stress. In higher organisms, in turn, protein instability may be related to the physiological production of free oxygen radicals, which play multiple roles in metabolic processes. The phenomenon of instability in the respiratory chain proteins may have a strategic function although it appears to be detrimental to the stability of the protein structure per se.

THE IMPACT OF IONIZED WATER SUPPLEMENTED WITH GLUTATION AND VITAMIN C DURING ACUTE HYPERTHERMIC EXPOSURE ON THE CONCENTRATION OF TOTAL PROTEINS IN THE BLOOD SERUM AT WHITE LABORATORY RATS

Alkaline water is in the focus of scientific interest over the last decade, due to its ability to alkalize the organism as well as its antioxidant effect. The aim of our research is to determine the impact of ionized water by adding enzymatic and non-enzymatic antioxidants, glutathione and vitamin C, during hyperthermic stress, on the concentration of total proteins. The experiment performed on a white laboratory Wistar rats, a female sex, weighing 180-220 g, young rats, divided into three groups of 15 individuals. Oxidative stress was caused by acute hyperthermic exposure at 41˚C. The first group is the control group (KPM), the second group is treated with ionized water (TAM), and the third group with ionized water with added glutathione and vitamin C (TAD). The duration of treatment lasted 21 days. Acute hyperthermic exposure caused a significant difference in total protein concentration between the 7th and 14th day taken as the starting points and the 21st day of treatment, respectively, on the day of hyperthermic exposure of rats. Such a significant difference was observed in the KPM and the TAM group (p &lt;0.001), whereas the TAD group is an exception where no statistically significant difference was observed in the same period (p&gt; 0.05). Animals treated with ionized water on day 14 showed a significantly higher total protein concentration (p &lt;0.01) compared to the group receiving only natural water, which is consistent with the finding obtained by comparison between the KPM and TAD group at 21st day where the difference is also statistically significant (p &lt;0.05). The acute application of the treatment with ionized water, without or in combination with other antioxidants, in the absence of hyperthermic stress does not lead to a significant alteration in protein concentration. The hyperthermic exposure intensifies the production of oxygen radicals and the potential for oxidative modification of proteins and molecules with a protein structure. Individual action of ionized water as well as synergism with the added antioxidants caused a high protective effect on oxidative damage. Because of this, the effect of oxidative cleavage is less pronounced in rats treated with ionized water and antioxidants because they have a stronger antioxidant defense that prevents oxidative modification of proteins and all biomolecules in the body.

P1826Computational fluid dynamic analysis of acute aortic dissection

Abstract Background Mechanism of acute aortic dissection still is not well understood. It is very difficult to predict where, when, and how acute aortic dissection occurs. This time we focused on blood flow velocity (FV), wall shear stress (WSS), and oscillatory shear index (OSI) which is three dimensional fluctuations of WSS inside ascending aorta. It is well recognized that a higher or lower WSS may cause intimal dysfunction, which could result in the progression of atherosclerosis. Furthermore, because of the non-slip boundary mechanical equilibrium of forces, WSS will separate the inner and outer layers of the aorta. This sheer force could stress the medial tissue and result in degeneration. OSI has been reported as important hemodynamic parameter that is highly associated with the progression of atherosclerosis by inducing radical oxygen production of the endothelial cells. Therefore, OSI may be more closely associated with the degeneration of the media in comparison with WSS. This time FV, WSS, and OSI was evaluated using computational fluid dynamics to identify the location of acute aortic dissection entry. Method Using computed tomography data of three patients (Case A, B, C) who had acute aortic dissection, each three pre-dissection aorta model was made. Computer simulation images of four dimensional pulsatile blood flow was made and pulsatile cardiac flow from one cardiac cycle was simulated. A three-dimensional movie was made to evaluate FV, WSS, and OSI. One normal size aorta was evaluated as a control. Results In control, blood flow inside ascending aorta was laminar, on the contrary, spiral flow was observed in three dissection cases. FV was slightly higher in dissection case (control: 0.56m/s, A: 0.77m/s, B: 1.05m/s, C: 0.35m/s). Maximum WSS in ascending aorta was 3.24Pa in control. In dissection cases, there was patchy high (A: 18.6Pa, B: 25.6Pa, C: 4.0Pa,) WSS lesion in ascending aorta. In case A and B, these patchy high lesions are close to the entry site. In all three dissection case, OSI was high around entry site. Figure 1 Conclusion According to our computer simulation, patchy high WSS and high OSI lesion is related with acute dissection entry site at the ascending aorta. Acknowledgement/Funding None

The mTOR kinase inhibitor rapamycin enhances the expression and release of pro-inflammatory cytokine interleukin 6 modulating the activation of human microglial cells.

Emerging evidence suggests the potential use of rapamycin in treatment of several neurological disorders. The drug readily crosses the blood brain barrier and may exert direct immunomodulatory effects within the brain. Microglia are the main innate immune cells of the brain, thus critically involved in the initiation and development of inflammatory processes at this level. However, there are conflicting data from rodent studies about the pharmacological effects of rapamycin on microglial inflammatory responses. Considering that rodent microglia display relevant biochemical and pharmacological differences compared to human microglia, in the present study we studied the effects of rapamycin in an experimental model of human microglia, the human microglial clone 3 (HMC3) cell line. Rapamycin was tested in the nM range both under basal conditions and in cells activated with a pro-inflammatory cytokine cocktail, consisting in a mixture of interferon-γ and interleukin-1β (II). The drug significantly increased II stimulatory effect on interleukin-6 (IL-6) expression and release in the HMC3 cells, while reducing the production of free oxygen radicals (ROS) both under basal conditions and in cells activated with II. Consistently with its known molecular mechanism of action, rapamycin reduced the extent of activation of the so-called 'mechanistic' target of rapamycin complex 1 (mTORC1) kinase and the total amount of intracellular proteins. In contrast to rodent cells, rapamycin did not alter human microglial cell viability nor inhibited cell proliferation. Moreover, rapamycin did not exert any significant effect on the morphology of the HMC3 cells. All together these data suggest that the inhibition of mTORC1 in human microglia by rapamycin results in complex immunomodulatory effects, including a significant increase in the expression and release of the pro-inflammatory IL-6.

Drug-induced interstitial lung disease: role of pharmacogenetics in predicting cytotoxic mechanisms and risks of side effects.

The diagnosis of drug-induced interstitial lung disease (DI-ILD) is challenging and mainly made by exclusion of other possible causes. Toxicity can occur as a cause of drug(s) or drug-drug interactions. In this review, we summarize the possible role of pharmacogenetics of metabolizing enzymes in DI-ILD. Knowledge of the genetic predispositions of enzymes involved in drug metabolization and their relation with proposed cytotoxic mechanisms of DI-ILD, in particular direct cell toxicity and free oxygen radical production is increasing. The cytochrome P450 enzyme family and other enzymes play an important role in the metabolism of all sorts of ingested, injected, or inhaled xenobiotic substances. The liver is the major site for metabolism. Metabolic cytotoxic mechanisms have however also been detected in lung tissue. Polymorphisms in genes coding for enzymes that influence metabolic activity may lead to localized (toxic) reactions and tissue damage. This knowledge may be helpful in preventing the risk of DI-ILD. Drug toxicity can be the consequence of absence or very poor enzyme activity, especially if no other metabolic route is available. In the case of reduced enzyme activity, it is recommended to reduce the dose or to prescribe an alternative drug, which is metabolized by a different, unaffected enzyme system to prevent toxic side effects. However, enhanced enzyme activity may lead to excessive formation of toxic and sometimes reactive metabolites. Therefore, knowing a patient's drug-metabolizing profile before drug prescription is a promising way to prevent or explain DI-ILD.

Oxidative stress mediated by gyrophoric acid from the lichen Umbilicaria hirsuta affected apoptosis and stress/survival pathways in HeLa cells

BackgroundLichens produce a huge diversity of bioactive compounds with several biological effects. Gyrophoric acid (GA) is found in high concentrations in the common lichen Umbilicaria hirsuta, however evidence for biological activity was limited to anti-proliferative activity described on several cancer cell lines.MethodsWe developed and validated a new protocol for GA isolation, resulting in a high yield of highly pure GA (validated by HPLC and NMR) in an easy and time saving manner. Anti-proliferative and pro-apoptotic activity, oxygen radicals formation and stress/survival proteins activity changes was study by flow cytometry.ResultsThe highly purified GA showed anti-proliferative activity against HeLa (human cervix carcinoma) and other tumor cells. Moreover, GA threated cells showed a significant increase in caspase-3 activation followed by PARP cleavage, PS externalization and cell cycle changes mediated by oxidative stress. Production of oxygen radicals led to DNA damage and changes in stress/survival pathways activation.ConclusionsGA treatment on HeLa cells clearly indicates ROS production and apoptosis as form of occurred cell death. Moreover, DNA damage and changing activity of stress/survival proteins as p38MAPK, Erk1/2 and Akt mediated by GA treatment confirm pro-apoptotic potential. The pharmacological potential of U. hirsuta derived GA is discussed.