Predictive value of some pro-oxidants in type 2 diabetes mellitus with vascular complications.

The study aims to analyze oxidative stress levels in circulation of some reactive molecules and products of biomolecular modification in type 2 diabetes mellitus (T2DM) with diabetes-specific vascular complications in order to determine their predictive value. Also, the alterations of their serum concentration with reference to disease characteristics were assessed. Reactive oxygen species (ROS), nitric oxide radicals (•NO), malondialdehyde (MDA), protein carbonyl (CO) and 8-hydroxydeoxyguanosin (8-OHdG) in serum were measured in 93 patients with T2DM with vascular complications, 94 control subjects and 16 diabetic patients who had no evidence of vascular disease. T2DM patients with clinically manifest vascular disease exhibit significantly elevated concentrations of all pro-oxidants in comparison to healthy subjects, with the highest degree of increase of •NO radicals. The levels of carbonylated proteins, ROS and 8-OHdG were significantly increased in insufficiently compensated diabetes as compared to good glycemic control state. Also, serum MDA, protein CO and 8-OHdG showed an association with glycemic control parameters. MDA, ROS and 8-OHdG correlated mostly with microvascular complications. Significant area under the curve (AUC) from plotted receiver operating characteristic (ROC) curves were obtained for all studied biomarkers, as for nitric oxide it was substantially bigger compared to those for the other pro-oxidants. Correspondingly, positive and negative predictive values related to the disease were in favor of the •NO radicals. The cutoff values of oxidative biomarkers may serve as an indicator of clinical reference for detecting T2DM with associated vascular complications, as nitric oxide radicals were the most reliable indicator.

Reducing oxidative toxicity of L-dopa in combination with two different antioxidants: an essential oil isolated from Rosa Damascena Mill., and vitamin C.

Parkinson disease (PD) is a multifactorial disease that takes a leading place among contemporary frequent diseases of the central nervous system (CNS) with not well-established mechanism. One of the most popular and effective therapy for patients with PD is Levodopa (L-dopa), but clinical effect of the drug diminished by motor complications resulting from prolonged treatment. Due to the L-dopa neurotoxic effect in the disease treatment, the L-dopa administration is delayed as long as possible in order to avoid side effects. In addition, combining L-dopa therapy with antioxidants, may decrease side-effects and provide symptomatic relief. The aim of the current research was to explore the possibility to reduce the oxidative stress (OS) induced by the L-dopa after its combining with two different antioxidants an essential oil isolated from Rosa damascena Mill., and vitamin C through experimental model of healthy mice. For this purpose, some oxidative stress indicators were evaluated - the lipid and protein oxidation end products - such as lipid peroxidation products measured as malondialdehyde (MDA) levels, protein carbonyl content (PCC), and advanced glycation end products (AGEs) in blood plasma of the experimental mice. For this purpose, was studied blood isolated from healthy mice after i.p. treatment with L-dopa (100 mg/kg). The groups with combining therapy were pre-treated first with Ascorbic acid (400 mg/kg), Rose oil (400 mg/kg). Statistically significant increased MDA levels, PCC and AGEs were found in the blood L-dopa treated mice compared to the controls, while the same parameters were significantly decreased in group pre-treated with antioxidants compared to the same controls. As a conclusion, the studied antioxidants can protect organisms from induced L-dopa oxidative toxicity and may play a key role in end products protection.

Hepatoprotective properties of Curcuma longa L. extract in bleomycin-induced chronic hepatotoxicity.

Curcuma longa L. (CLL) extract has previously been reported to alleviate liver damage. The current study examined the antioxidant activity of CLL by which the extract protects the liver against bleomycin (BLM)-induced hepatotoxicity in mice. The hypothesis was that CLL extract would protect the liver by reducing oxidative stress (induced superoxide dismutase (SOD) and catalase (CAT) activity), inhibiting lipid peroxidation, lowering biochemical parameters, and decreasing ROS production. Hepatic toxicity was induced by intraperitoneal injection of mice once daily with BLM (0.069 U/mL; 0.29 U/kg bw.) for a period of 4 weeks. The CLL was administered once a day for 4 weeks, 2 h prior at dose (40 mg/mL; 0.187 mg/kg/day). CLL extract significantly protected the liver, it decreased plasma bilirubin (BL) and gamma glutamyl transpeptidase (GGT), and it reduced lipid peroxidation levels. BLM intoxication produced oxidative stress, in which the antioxidant system functioned incorrectly and ROS production significantly increased. The CLL extract provided significant hepatic protection against BLM toxicity by improving SOD, CAT (p < 0.05), and MDA levels and decreasing ROS in the group receiving BLM (p < 0.05), leading to reduced membrane lipid peroxidation. Throughout this study, the CLL extract facilitated recovery from BLM-induced hepatic injury by suppressing oxidative stress. Therefore, the CLL extract has the potential to serve as an antioxidant compound to treat chronic hepatotoxicity.

Antioxidant and prooxidant effects of Piptadeniastrum africanum as the possible rationale behind its broad scale application in African ethnomedicine.

ETHNOPHARMACOLOGICAL RELEVANCE: Piptadeniastrum africanum is widely used in treating oxidative stress related diseases. Oxidative stress, defined as the disturbance in the balance between the production of free radicals and antioxidant defenses, is the root cause of many pathophysiological conditions. Based on the dual properties of prooxidants as toxic and beneficial compounds, both prooxidants and antioxidants may be effective in the treatment of these conditions when the right dose is given to the right subject at the right time for the right duration. AIM OF THE STUDY: This study was aimed at investigating the in vitro and ex vivo anti- and pro-oxidative effects of P. africanum. MATERIALS AND METHODS: Total phenolic and flavonoid contents of methanol and aqueous extracts of P. africanum stem back were quantified spectrophotometrically. The methanol extract, ascorbate radicals and reactive oxygen species in brain and liver homogenates of mice treated with the methanol stem bark extract were analyzed by electron paramagnetic resonance (EPR) spectroscopy. Free radical scavenging of DPPH was determined by spectrophotometric and EPR assays. RESULTS: The methanol extract was richer in both phenolic and flavonoid contents compared to the aqueous extracts and also showed better DPPH radical scavenging capacity. The EPR spectroscopy in vitro analysis exhibited high DPPH scavenging capacity before and after UV irradiation (99.5% and 98.76%) at 40 µg/ml extract. The ex vivo EPR spectroscopy studies demonstrated increased levels of ascorbate radicals (•Asc) in liver and brain homogenates of healthy mice treated with P. africanum in comparison with those of the non treated controls (0.6141 ±â€¯0.026 vs 0.1800 ±â€¯0.0073 arb. units for liver homogenates and 0.9605 ±â€¯0.0492 vs 0.3375 ±â€¯0.0062 arb. units for brain homogenates, correspondingly). Considerably, higher levels of reactive oxygen species (ROS) were measured in mice liver and brain homogenates after treatment with P. africanum extract compared to the control group, as well (1.9402 ±â€¯0.1200 vs 0.6699 ±â€¯0.062 arb. units for liver homogenates and 1.7325 ±â€¯01503 vs 0.3167 ±â€¯0.0403 arb.units, respectively). CONCLUSION: Therefore, P. africanum exhibited antioxidant and pro-oxidant properties which may explain its broad spectrum use in a wide variety of ailments.

Epidermal barrier and oxidative stress parameters improve during in 311 nm narrow band UVB phototherapy of plaque type psoriasis.

BACKGROUND: Psoriasis is a multi-systemic inflammatory disease that results from dysregulation between epidermal keratinocyte homeostasis and both innate and acquired immunity. Epidermal barrier defect has been described in psoriatic lesions. Furthermore an imbalance between pro-oxidative stress and antioxidant defense mechanisms are known in psoriasis patients. AIM: The aim of this study was to address the link between disease activity, epidermal barrier and systemic oxidative stress in the course of 311 nm narrow band ultraviolet B (NB-UVB) therapy of psoriasis. The dynamic of systemic oxidative stress parameters as well as local transepidermal water loss (TEWL) and stratum corneum hydration (SCH) was characterized before and after 311 nm NB-UVB therapy on the plaques of psoriasis vulgaris in comparison to untreated non-affected volar forearm sites of the same patients. MATERIAL AND METHODS: 22 patients with plaque type psoriasis vulgaris and 25 gender- and age-matched healthy controls were enrolled. We assessed the psoriasis area and severity index (PASI) and the dermatology life quality index (DLQI) for monitoring disease activity, severity and self-perceived DLQI impact as patient related outcome parameter. We measured non-invasively TEWL (Tewameter TM 300) and SCH (Corneometer CM 825) and the end product of lipid peroxidation - malondialdehyde (MDA), Reactive oxygen species (ROS), ascorbyl radicals (Asc) and detoxifying activity of catalase (CAT) were measured in the peripheral blood with spectrophotometric and EPR spectroscopy methods. RESULTS: Disease activity improved in all patients compared to baseline witnessed by significant decrease in PASI; (from 14.1 to 10.4; p < 0.0001) and DLQI (from 11.7 to 8.1; p < 0.0001). At baseline TEWL-values were significantly (p < 0.0001) higher on psoriatic plaques (16.8 g/h/m2) in comparison to uninvolved skin (5.3 g/h/m2); with a decrease at both sites after NB-UVB phototherapy. SCH was significantly lower at psoriatic plaque s (4.7AU) compared to uninvolved sskin (42.4AU) and increased after treatment (8.6AU) (p < 0.0001). Interestingly, SCH decrease slightly during therapy at uninvolved skin (40.6AU). ROS and Asc declined during therapy in parallel to a decrease in MDA. A mild decrease in the antioxidative enzyme CAT activity which did not reach the significance was observed. CONCLUSION: The presented data is shows that a clinical improvement of psoriatic plaques under NB-UVB therapy, shown in with a decreased PASI and reflected by an increase in quality of life has beneficial effects on epidermal barrier function, SCH and improvement of systemic oxidative stress parameters (ROS, MDA and Asc). We assume that the general improvement in the oxidative stress parameters along with epidermal barrier parameters reflects mainly the improvement of disease activity which overwrites the possible negative pro-oxidative effects of the UV treatment.

Influence of glycemic control on some real-time biomarkers of free radical formation in type 2 diabetic patients: An EPR study.

BACKGROUND: The pathology of diabetes is associated with several mechanisms, one of which is oxidative stress (OS). The relationship between OS and diabetic complications has been extensively investigated. OS has been suggested to be involved in the genesis of both macroand microangiopathy. In contrast, the relationship between OS and insulin action is a neglected research area. OBJECTIVES: The aim of this study is to elucidate the effect of glycemic control in type 2 diabetic patients by following the serum levels of some real-time oxidative stress biomarkers. MATERIAL AND METHODS: The study group consisted of 53 type 2 diabetic patients (31 with poor glycemic control and 22 with good glycemic control) and 24 healthy control subjects. The oxidative stress biomarkers (ROS, Asc• and •NO) were measured by using electron paramagnetic resonance spectroscopy (EPR) methods and compared with clinical parameters. RESULTS: The statistically significantly higher levels of ROS products and •NO in type 2 diabetic patients in both groups compared to controls mean that the oxidation processes take place at the time the survey is performed. Free radical overproduction persists after the normalization of the glucose levels, and oxidative stress may be involved in the "metabolic memory" effect. This is confirmed by the positive correlation between ROS levels/•NO and average blood glucose levels, triglycerides, and total cholesterol. Furthermore, the low level of the ascorbate radical in both diabetes groups compared to controls confirmed an increase in oxidation processes. CONCLUSIONS: Higher levels of real-time biomarkers show that intensive insulin treatment does not lead to the expected decrease in oxidative processes involving ROS and •NO, probably due to "metabolic memory".

Protective effect of two essential oils isolated from Rosa damascena Mill. and Lavandula angustifolia Mill, and two classic antioxidants against L-dopa oxidative toxicity induced in healthy mice.

Levodopa (L-dopa) is a "gold standard" and most effective symptomatic agent in the Parkinson's disease (PD) treatment. The several treatments have been developed in an attempt to improve PD treatment, but most patients were still levodopa dependent. The issue of toxicity was raised in vitro studies, and suggests that L-dopa can be toxic to dopaminergic neurons, but it is not yet entirely proven. L-dopa prolonged treatment is associated with motor complications and some limitations. Combining the L-dopa therapy with antioxidants can reduce related sideeffects and provide symptomatic relief. The natural antioxidants can be isolated from any plant parts such as seeds, leaves, roots, bark, etc., and their extracts riched in phenols can retard the oxidative degradation of the lipids, proteins and DNA. Thus, study suggests that combination of essential oils (Rose oil and Lavender oil), Vitamin C and Trolox with Ldopa can reduce oxidative toxicity, and may play a key role in ROS/RNS disarm.

Biological evaluation of new potential anticancer agent for tumour imaging and radiotherapy by two methods: 99mTc-radiolabelling and EPR spectroscopy.

Recently, a new class of in vitro and ex vivo radiotracers/radioprotectors, the nitroxyl-labelled agent 1-ethyl-1-nitroso-3-[4-(2,2,6,6-tetramethylpiperidine-1-oxyl)]-urea (SLENU), has been discovered. Our previous investigations demonstrated that SLENU is a low-molecular-weight stable free radical which is freely membrane permeable, easily crosses the blood brain barrier and exhibited in/ex vivo the lowest general toxicity and higher anticancer activity against some experimental tumour models. Further investigation was aimed to develop a 99mTc-labelled SLENU (97%) as a chelator and evaluate its labelling efficiency and potential use as a tumour seeking agent and for early diagnosis. Tissue biodistribution of 99mTc-SLENU was determined in normal mice at 1, 2 and 24 h (n = 4/time interval, route of administration i.v.). The distribution data were compared using male albino non-inbred mice and electron paramagnetic resonance investigation. The imaging characteristics of 99mTc-SLENU conjugate examined in BALB/c mice grafted with Ehrlich Ascitis tumour in the thigh of hind leg demonstrated major accumulation of the radiotracer in the organs and tumour. Planar images and auto-radiograms confirmed that the tumours could be visualized clearly with 99mTc-SLENU. Blood kinetic study of radio-conjugate showed a bi-exponential pattern, as well as quick reduced duration in the blood circulation. This study establishes nitroxyls as a general class of new spin-labelled diagnostic markers that reduce the negative lateral effects of radiotherapy and drug damages, and are appropriate for tumour-localization.

Tissue redox activity as a sensing platform for imaging of cancer based on nitroxide redox cycle.

The experience in free radical biology and medicine shows the crucial role of redox signalling in carcinogenesis. The cells and tissues of healthy mammals are characterised by a low level of reactive oxygen species (ROS) and some constant (reference) level of reducing equivalents. Increasing of ROS above the critical level provokes genomic instability. The present study describes universal methodology for direct imaging of tissue redox activity in carcinogenesis, which allows a differentiation of cancer development from normal condition. The experiments were conducted on: neuroblastoma-bearing mice; colon cancer-bearing mice; and healthy mice. The tissue redox activity was visualised in vivo by nitroxide-enhanced magnetic resonance imaging (MRI) on anesthetised animals. The method is based on nitroxide redox cycle, coupled with appearance/disappearance of MRI signal. The half-life (τ1/2) of nitroxide-enhanced MRI signal in the respective tissue was used as a diagnostic marker. The study provides direct evidence that healthy and cancer-bearing mammalian tissues are characterised by different redox activities - a basis for cancer diagnosis. The tissues (cancer and 'normal') of cancer-bearing mammals were characterised by a long-lived MRI signal (τ1/2>14 min), indicating a high oxidative activity. The tissues of healthy organism were characterised by a short-lived MRI signal (τ1/2=1-3 min), indicating a high reducing activity. The study shows that tissue redox activity is a sensing platform for imaging of cancer using nitroxide-enhanced MRI. It also suggests that 'normal' tissues of cancer-bearing organism are susceptible to oxidative damage.

In vitro studies on radioprotective efficacy of silymarin against γ-irradiation.

UNLABELLED: Abstract Purpose: Silymarin has been widely exploited for its hepatoprotective activities. This study aimed to evaluate the protective efficacy of silymarin against γ-radiation. MATERIALS AND METHODS: The radioprotective properties of silymarin were studied using different assays. Cytotoxicity of silymarin on Human embryonic kidney (HEK) cells was evaluated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) assay. Protective efficacy against γ-radiation was assessed by studying reduction in micronuclei frequency and free radical generation using 2',7'-dichlorodihydroflurescin diacetate (H2DCFDA). Radiation-induced apoptosis was estimated by Annexin V-PI (propidium iodide) analysis and cell cycle analysis. γ-radiation induced changes in mitochondrial membrane potential (MMP) and DNA damage was estimated employing flow-cytometry and comet assay respectively. RESULTS: MTT assay and Annexin V-PI studies showed that pre-incubation of HEK cells with silymarin protected them from γ-irradiation. Significant reduction in apoptosis (76.36%) was observed. Silymarin also decreased the percentage of radiation-induced micronuclei (> 69%) (p < 0.05 ). Measurement of intracellular reactive oxygen species (ROS) by H2DCFDA revealed a reduction in ROS (21%) at 0.5 h. Cell cycle analysis revealed G1 block in the unirradiated control, which declined in the silymarin pretreated irradiated group (0.5 h). Silymarin treatment resulted in a significant increase in MMP (2 h) against the radiation control. Moreover, the presence of silymarin during irradiation significantly decreased the DNA damage (as measured by comet assay). CONCLUSIONS: Protection against radiation-induced cell-death and DNA damage by silymarin could be attributed to a reduction in ROS induced by γ-radiation. In vitro experiments on HEK cells explicitly prove that silymarin is a promising, effective and safe radiation countermeasure agent.

The impact of reactive oxygen species on anticancer therapeutic strategies.

Over 50 years of experience in free radical biology and medicine shows that normal cells of healthy mammals are characterized by a low steady-state level of reactive oxygen species (ROS) and a constant (reference) level of reducing equivalents. A lasting increase of ROS above the critical level leads to permanent oxidative stress in the cells. This could cause genomic instability and mutations, which are responsible for adaptation of cells to oxidative stress and their survival in an oxidative environment. In turn, these events could provoke malignancy. It is widely accepted that the balance between ROS and reducing equivalents in cells and tissues determines their redox status. The evaluation of tissue redox status has great diagnostic potential in cancer, as well as prognostic potential for cancer therapy, and could significantly contribute to the planning of appropriate treatment and to increasing the patients' quality of life. The conventional therapeutic strategy is based on drugs that increase ROS generation and induce apoptosis in cancer cells. However, this therapeutic approach has serious disadvantages: the expression of various toxic side effects in normal (non-cancer) tissues. The current review describes the basics of free radical biology in carcinogenesis. The authors emphasize the different redox status of normal and cancer cells, which permits the use of this parameter as a new therapeutic target. The authors also outline some directions for the development of promising therapeutic strategies based on the regulation of redox signaling using combined therapy. The review is intended for a broad readership - from non-specialists to researchers in the field of cancer biochemistry and pharmacy.

Cell-penetrating nitroxides as molecular sensors for imaging of cancer in vivo, based on tissue redox activity.

The present study shows that hydrophobic and cell-penetrating piperidine-type nitroxide radicals SLENU and TEMPOL, but not hydrophilic and partially penetrating or non-penetrating pyrrolidine-type nitroxides carbamoyl-PROXYL and carboxy-PROXYL, are appropriate contrast agents for magnetic resonance imaging (MRI) of cancer, based on its functionality - tissue redox activity. The experiments were conducted on anesthetized mice: healthy and neuroblastoma-bearing in a moderate stage of cancer development. The method is based on the nitroxide redox cycle, coupled with appearance or disappearance of the MRI signal. The half-life (τ(1/2)) of a nitroxide-enhanced MRI signal in the respective tissue was used as a marker to assess tissue redox activity to the nitroxide radical. In the case of SLENU and TEMPOL, there were large differences in the histograms between control and cancer-bearing mice. All tissues (cancer and non-cancer) of cancer-bearing organisms were characterized by a long-lived MRI signal (τ(1/2) > 14 min), indicating a high oxidative activity. The tissues of healthy organisms were characterized by a short-lived MRI signal (τ(1/2) = 1-3 min), indicating a high reducing activity. In the case of carbamoyl-PROXYL and carboxy-PROXYL, there was no difference in the histograms between control and cancer-bearing mice. The data show that the penetration of nitroxide in cells and tissues is obligatory for imaging of cancer, based on its redox activity. The principle of the method is applicable also to biopsy specimens, using MRI or EPR spectroscopy. We provide direct evidence that the nitroxide redox cycle could be used as a sensing platform for functional imaging of different pathologies, based on changes in cellular and tissue redox activity, as in the case of cancer.

Radical scavenging and radiomodulatory effects of Psoralea corylifolia Linn. substantiated by in vitro assays and EPR spectroscopy.

The present study is the first report of the radiomodulatory effects of Psoralea corylifolia Linn. The extract (IBG-RA-26) prepared from P. corylifolia was chemically analysed by HPLC, LC-MS/MS and NMR. The total polyphenolic content of IBG-RA-26 was 0.287 mg/ml of quercetin equivalents. IBG-RA-26 exhibited a dose-dependent increase in 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity. It exhibited comparable (> 50%) site-specific and non-site-specific hydroxyl radical scavenging activity in higher concentration ranges (500-1000 microg/ml), while at lower concentrations (5-50 microg/ml) it exhibited significantly (p < 0.05) higher non-site-specific scavenging ability compared to site-specific activity. Nitric oxide scavenging activity of IBG-RA-26 (5-1000 microg/ml) increased in a concentration-dependent manner, while maximum superoxide ion scavenging ability (79%) was observed at 50 microg/ml. The electron donation potential of IBG-RA-26 was found to be higher than that of ascorbic acid at lower concentrations (up to 5 microg/ml). Analysis of the ability of IBG-RA-26 to protect membranes against gamma-radiation, utilizing an artificial membrane system (liposome), revealed a significant (p < 0.05) decrease in the formation of malondialdehyde (MDA) as a function of the concentration of IBG-RA-26. Radiation-induced lysis of human erythrocytes was monitored and efficacy of IBG-RA-26 was tested in the concentration range 25-1000 microg/ml, with significant protective efficacy observed in the range 25-50 microg/ml. IBG-RA-26 rendered significant (p < 0.05) protection against radiation (0.25 kGy)-induced DNA damage. EPR spectroscopy was used to investigate the DPPH radical scavenging capacity of IBG-RA-26. IBG-RA-26 exhibited a good DPPH radical scavenging capacity in a concentration-dependent manner. By direct EPR spectroscopy we have also demonstrated the possible formation of free radical species in a solution of IBG-RA-26. The wide spectrum of radioprotective and antioxidant properties exhibited by IBG-RA-26 indicate that P. corylifolia has potential as a radiomodulatory agent.

Induction of immunostimulatory cytokine genes expression in human PBMCs by a novel semiquinone glucoside derivative (SQGD) isolated from a Bacillus sp. INM-1.

In the present study, a semiquinone glucoside derivative (SQGD) isolated from a radioresistant bacterium Bacillus sp. INM-1 was evaluated for its immunostimulatory activities. Human peripheral blood mononuclear cells (PBMCs) were stimulated by different doses (30-90 microg/ml) of SQGD for different time (3-12h) intervals at 37°C, and IL-12p40, IL-23p19, IL-10, RelA and c-Jun gene expression analysis was carried out by qRT-PCR method. SQGD dose dependent cytokines protein expression kinetic analysis was carried out using western blotting. As the results of SQGD (30µg/ml) stimulation for 3h at 37°C, significant induction in IL-12p40, IL-23p19 and RelA gene expression was observed in PBMCs compared to unstimulated control cells. However, no such induction in IL-10 and c-Jun gene expression was observed. Time dependent protein expression study indicated significant increase in IL-12p40, IL-12p35, IL-23p19 and RelA protein expression at 3-6h, which was found decrease at 12h upon SQGD treatment. In contrast, IL-10 protein expression was found to enhance significantly at 12h after SQGD treatment to the PBMCs. SQGD dose dependent study showed approximately similar level of induction in IL-12p40, IL-12p35, IL-23p19 and RelA proteins expression at all tested concentration (30-90 microg/ml) compared to control. However, no significant change in the IL-10 and c-Jun protein expression was observed at any SQGD concentration. SQGD treatment (0.25mg/kgbwt.) was also found to enhance anti-keyhole Limpet Hemocynin (KLH) IgM antibodies significantly in the mice immunized by KLH. Thus, SQGD fraction stimulates cellular immunity by inducing immunostimulatory cytokines and humoral immunity by enhancing IgM antibodies and could be a promising immunostimulant. Further studies related to molecular mechanisms offering immunostimulation is underway, will certainly helpful to unravel its mode of action in the biological system.

Imaging of cancer by redox-mediated mechanism: a radical diagnostic approach.

The present study describes a new diagnostic approach for carcinogenesis based on the different tissue redox activity of normal and cancer-bearing mammalians and its visualization and estimation by cell permeable and DNA annealing probe (nitroxide-labeled nitrosourea) and magnetic resonance imaging.

Evaluation of silymarin as a promising radioprotector.

Silymarin, a purified extract of seeds of Silybum marianum L. and well known for its hepatoprotective abilities, has been evaluated for inherent utility as a radioprotective agent. A fraction (INM-7035) was authenticated by characterizing the percentage composition of silybin A and B (39.9% and 57.4%). Free radical scavenging activities of INM-7035 against superoxide radicals (>68%), hydroxyl radicals (>33.75%), DPPH (67.2%), and ABTS (32.4%) were also evaluated. The fraction chelated (>30%) ferrous ions, thereby able to restrict amplification. INM-7035 exhibited >50% peroxyl radical scavenging activity in the lipid phase along with dose-dependent (R2 = 0.990) reducing power in the aqueous phase. Radiation-induced free radical flux can lead to disruption of biomolecules like membrane lipids. INM-7035 completely inhibited lipid peroxidative stress in case of membranes against supralethal radiation stress in the liposomal system. The ability of INM-7035 to modulate the levels of NF-kappaB, indicated its inherent potential as a radioprotective bioactive constituent.

Electron paramagnetic resonance spectroscopy in radiation research: Current status and perspectives.

Exposure to radiation leads to a number of health-related malfunctions. Ionizing radiation is more harmful than non-ionizing radiation, as it causes both direct and indirect effects. Irradiation with ionizing radiation results in free radical-induced oxidative stress. Free radical-mediated oxidative stress has been implicated in a plethora of diseased states, including cancer, arthritis, aging, Parkinson's disease, and so on. Electron Paramagnetic Resonance (EPR) spectroscopy has various applications to measure free radicals, in radiation research. Free radicals disintegrate immediately in aqueous environment. Free radicals can be detected indirectly by the EPR spin trapping technique in which these forms stabilize the radical adduct and produce characteristic EPR spectra for specific radicals. Ionizing radiation-induced free radicals in calcified tissues, for example, teeth, bone, and fingernail, can be detected directly by EPR spectroscopy, due to their extended stability. Various applications of EPR in radiation research studies are discussed in this review.

Nitroxyl radicals for labeling of conventional therapeutics and noninvasive magnetic resonance imaging of their permeability for blood-brain barrier: relationship between structure, blood clearance, and MRI signal dynamic in the brain.

The present study describes a novel nonradioactive methodology for in vivo noninvasive, real-time imaging of blood-brain barrier (BBB) permeability for conventional drugs, using nitroxyl radicals as spin-labels and magnetic resonance imaging (MRI). Two TEMPO-labeled analogues (SLENU and SLCNUgly) of the anticancer drug lomustine [1-(2-chloroethyl)-3-cyclohexyl-1-nitrosourea] were synthesized, using a substitution of the cyclohexyl part with nitroxyl radical. Nonmodified nitroxyl radical TEMPOL was used for comparison. The nitroxyl derivatives were injected intravenously in healthy mice via the tail vein, and MR imaging of the brain was performed on a 7.0 T MRI. The MRI signal dynamic of SLENU and SLCNUgly followed the same kinetics as nonmodified TEMPO radical. SLENU and SLCNUgly were rapidly transported and randomly distributed in the brain tissue, which indicated that the exchange of cyclohexyl part of lomustine with TEMPO radical did not suppress the permeability of the anticancer drug for BBB. The selected nitroxyl derivatives possessed different hydrophobicity, cell permeabilization ability, and blood clearance. Based on these differences, we investigated the relationship betweenthe structure of nitroxyl derivatives, their half-life in the circulation, and their MRI signal dynamic in the brain. This information was important for estimation of the merits and demerits of the described methodology and finding pathways for overcoming the restrictions.

EPR signal reduction kinetic of several nitroxyl derivatives in blood in vitro and in vivo.

The present study is focused on the mechanism(s) of electron-paramagnetic resonance (EPR) signal reduction kinetic of several nitroxyl radicals and nitroxyl-labeled anticancer drugs in physiological solutions in the context of their application for evaluation of oxidation/reduction status of blood and tissues--an important step in biomedical diagnostics and planning of therapy of many diseases. The nitroxyl derivatives were characterized with different size and water-solubility. Some of them are originally synthesized. In buffer, in the absence of reducing and oxidizing equivalents, the EPR signal intensity of all nitroxyls was constant with the time. In serum and cell cultured medium, in an absence of cells and in a negligible amount of reducing and oxidizing equivalents, there was no significant EPR signal reduction, too. In vitro (in freshly isolated blood samples), the EPR signal intensity was characterized with slow decrease within 30 min, presumably as a result of interaction between the nitroxyl derivative and blood cells. The EPR spectrum of hydrophobic nitroxyls showed a slight anisotropy in cell-containing solutions and it did not changed in non-cell physiological solutions. This suggests for a limited motion of more hydrophobic nitroxyls through their preferable location in cell membranes. In vivo (in the bloodstream of mice under anesthesia), the EPR signal reduction kinetic was characterized by two phases: i) a rapid enhancement within 30 s as a result of increasing of nitroxyl concentration in the bloodstream after its intravenous injection, followed by ii) a rapid decrease (approximately 80-100%) within 2-5 min, presumably as a result of transportation of nitroxyl in the tissues. The hydrophobic nitroxyls were characterized with stronger and faster decrease in EPR signal intensity in the blood in vivo, as a result of their higher cell permeability, rapid clearance from the bloodstream and/or transportation in the surrounding tissues. The hydrophilic nitroxyls persist in the bloodstream (in their radical form) for a comparatively long time. The data suggest that the hydrophobic cell-permeable nitroxyl derivatives are most appropriate for evaluation of cell and tissue oxidation/reduction status, while the hydrophilic nitroxyls (impermeable for cell membranes or with very slow cell permeability) are most appropriate for evaluation of oxidation/reduction status of blood using EPR imaging.

Nitroxyl radicals as low toxic spin-labels for non-invasive magnetic resonance imaging of blood-brain barrier permeability for conventional therapeutics.

The present study describes a novel non-radioactive methodology for in vivo non-invasive, real-time imaging of blood-brain barrier (BBB) permeability for conventional drugs, using nitroxyl radicals as spin-labels and magnetic resonance imaging (MRI).