Detection Of Superoxide Dismutase Research Articles

Alcohol- and Low-Iron Induced Changes in Antioxidant and Energy Metabolism Associated with Protein Lys Acetylation.

Understanding the role of iron in ethanol-derived hepatic stress could help elucidate the efficacy of dietary or clinical interventions designed to minimize liver damage from chronic alcohol consumption. We hypothesized that normal levels of iron are involved in ethanol-derived liver damage and reduced dietary iron intake would lower the damage caused by ethanol. We used a pair-fed mouse model utilizing basal Lieber-DeCarli liquid diets for 22 weeks to test this hypothesis. In our mouse model, chronic ethanol exposure led to mild hepatic stress possibly characteristic of early-stage alcoholic liver disease, seen as increases in liver-to-body weight ratios. Dietary iron restriction caused a slight decrease in non-heme iron and ferritin (FeRL) expression while it increased transferrin receptor 1 (TfR1) expression without changing ferroportin 1 (FPN1) expression. It also elevated protein lysine acetylation to a more significant level than in ethanol-fed mice under normal dietary iron conditions. Interestingly, iron restriction led to an additional reduction in nicotinamide adenine dinucleotide (NAD+) and NADH levels. Consistent with this observation, the major mitochondrial NAD+-dependent deacetylase, NAD-dependent deacetylase sirtuin-3 (SIRT3), expression was significantly reduced causing increased protein lysine acetylation in ethanol-fed mice at normal and low-iron conditions. In addition, the detection of superoxide dismutase 1 and 2 levels (SOD1 and SOD2) and oxidative phosphorylation (OXPHOS) complex activities allowed us to evaluate the changes in antioxidant and energy metabolism regulated by ethanol consumption at normal and low-iron conditions. We observed that the ethanol-fed mice had mild liver damage associated with reduced energy and antioxidant metabolism. On the other hand, iron restriction may exacerbate certain activities of ethanol further, such as increased protein lysine acetylation and reduced antioxidant metabolism. This metabolic change may prove a barrier to the effectiveness of dietary reduction of iron intake as a preventative measure in chronic alcohol consumption.

Superoxide Dismutase Detection on Silver Nanostructured Substrates through Surface-Enhanced Spectroscopic Techniques

Oxidative stress refers to the overproduction of reactive oxygen species and is often associated with numerous pathological conditions. Superoxide dismutase (SOD) is a widely used enzyme for evaluating oxidative stress, with numerous methods being developed for its detection in biological specimens like blood, urine, and saliva. In this study, a simple metal-assisted chemical etching method was employed for the fabrication of nanostructured silicon surfaces decorated with either silver dendrites or silver aggregates. Those surfaces were used as substrates for the immunochemical determination of SOD in synthetic saliva through surface-enhanced Raman spectroscopy (SERS) and surface-enhanced fluorescence (SEF). The immunoassay was based on a 3-step competitive assay format, which included, after the immunoreaction with the specific anti-SOD antibody, a reaction with a biotinylated secondary antibody and streptavidin. Streptavidin labeled with peroxidase was used in combination with a precipitating tetramethylbenzidine substrate for detection through SERS, whereas for SEF measurements, streptavidin labeled with the fluorescent dye Rhodamine Red-X was utilized. Both immunoassays were sensitive, with a detection limit of 0.01 μg/mL and a linear dynamic range from 0.03 to 3.3 μg/mL, enabling the evaluation of the oxidative stress status of an organism.

Immunosuppressant Tacrolimus Treatment Delays Acute Seizure Occurrence, Reduces Elevated Oxidative Stress, and Reverses PGF2α Burst in the Brain of PTZ-Treated Rats.

It is still an urgent need to find alternative and effective therapies to combat epileptic seizures. Tacrolimus as a potent immunosuppressant and calcineurin inhibitor is emerging as promising drug to suppress seizures. However, there are few reports applying tacrolimus to epilepsy and providing data for its antiseizure properties. In this study, we investigated the antiseizure effects of 5 and 10mg/kg doses of tacrolimus treatment priorly to pentylenetetrazol (PTZ) induction of seizures in rats. As an experimental design, we establish two independent rat groups where we observe convulsive seizures following 70mg/kg PTZ and sub-convulsive seizures detected by electroencephalography (EEG) following 35mg/kg PTZ. Thereafter, we proceed with biochemical analyses of the brain including assessment of malondialdehyde level as an indicator of lipid peroxidation and detection of superoxide dismutase (SOD) enzyme activity and PGF2α. Tacrolimus pre-treatment dose-dependently resulted in lesser seizure severity according to Racine's scale, delayed start-up latency of the first myoclonic jerk and attenuated the spike percentages detected by EEG in seizure-induced rats. However, only the higher dose of tacrolimus was effective to restore lipid peroxidation. An increase in SOD activity was observed in the PTZ group, mediated by seizure activity per se, however, it was greater in the groups that received treatment with 5 and 10mg/kg of Tacrolimus. PGF2α bursts following PTZ induction of seizures were reversed by tacrolimus pre-treatment in a dose-dependent manner as well. We report that the well-known immunosuppressant tacrolimus is a promising agent to suppress seizures. Comparative studies are necessary to determine the possible utilization of tacrolimus in clinical cases.

APPL1 ameliorates myocardial ischemia-reperfusion injury by regulating the AMPK signaling pathway.

Myocardial ischemia-reperfusion injury results in elevated reactive oxygen species (ROS) production and causes oxidative stress damage. Therefore, the current study aimed to investigate whether adaptor protein phosphotyrosine interacting with PH domain and leucine zipper 1 (APPL1) could induce the expression of antioxidant enzymes through AMP-activated protein kinase (AMPK) signaling in order to alleviate the injury caused by ischemia/hypoxia-reperfusion. Following induction of hypoxia-reoxygenation (H/R) injury in H9c2 cells, the liver kinase B1 (LKB1)/AMPK/acetyl-CoA carboxylase α (ACC) signaling pathway was investigated using western blot analysis, along with the detection of superoxide dismutase (SOD)2 and SOD3 expression. Additionally, cell viability was detected using a Cell Counting Kit-8 assay and ROS production was analyzed using ROS staining, whereas the expression levels of inflammatory mediators (TNF-α, monocyte chemoattractant protein 1 and IL-1β), apoptosis mediators [cleaved caspase-3, cleaved poly (ADP-ribose) polymerase and Bcl-2] and nuclear factor erythroid 2-related factor 2 signaling pathway-related proteins were detected via western blot analysis following overexpression of APPL1 alone or in combination with compound C treatment (an AMPK inhibitor). The results indicated that H/R induction upregulated the phosphorylation levels of LKB1, AMPK and ACC, and decreased the expression levels of APPL1 and SOD enzyme activities. APPL1 overexpression increased the phosphorylation levels of LKB1, AMPK and ACC, SOD enzyme activity and cell viability whereas the expression levels of proinflammatory mediators and proapoptotic mediators, and the levels of ROS production were markedly decreased when compared with H/R group with empty plasmid transfection. APPL overexpression-mediated effects were significantly abrogated by compound C. Taken together, the data indicated that APPL1 inhibited ROS production and H/R-induced myocardial injury via the AMPK signaling pathway. Therefore, APPL1 may serve as a potential therapeutic target for myocardial H/R injury.

A new insight to characterize immunomodulation based on hepatopancreatic transcriptome and humoral immune factor analysis of the Cherax quadricarinatus infected with Aeromonas veronii

Cherax quadricarinatus is a type of large freshwater crayfish that is characterized by rapid growth and formidable adaptability. It has also been widely cultured and studied as a model organism. Aeromonas veronii is the dominant pathogen in aquatic environments and the primary threat to aquaculture’s economic stability. To better understand the interactions between C. quadricarinatus and A. veronii, high-throughput RNA sequencing of the C. quadricarinatus hepatopancreas was carried out on a control group, susceptible group (6 h after infection), and resistant group (48 h after infection). A total of 65,850,929 genes were obtained. Compared with the control group, 2616 genes were up-regulated and 1551 genes were down-regulated in the susceptible group; while 1488 genes were up-regulated and 1712 genes were down-regulated in the resistant group. GO and KEGG analysis showed that these differentially expressed genes (DEGs) were associated with multiple immune pathways, including Toll-like receptors (TLRs), antigen processing and presentation, NOD-like receptor signaling pathway, phagosome, lysosome, JAK-STAT signaling pathway. qRT-PCR showed that infection by A. veronii changed the expression pattern of the serine proteinase inhibitor (SPI), crustacean hyperglycemic hormone (CHH), anti-lipopolysaccharide factor (ALF), and extracellular copper/zinc superoxide dismutase (SOD1), all of which were significantly higher than in the control group up to 48 h after infection. In addition, detection of superoxide dismutase (SOD), catalase (CAT), lysozyme (LZM), and phenoloxidase (PO) activity, as well as ceruloplasmin (CP) concentration at different times after infection showed diverse trends. Furthermore, pathological sections obtained 24 h after infection show lesions on the hepatopancreas and intestinal tissues caused by A. veronii. The results of this study provide a foundation for analyzing the immune mechanism of C. quadricarinatus infected with A. veronii at the transcriptional level and a theoretical basis for screening disease-resistant individuals to ensure healthy economic development of the aquatic industry.

Detection of superoxide dismutase (Cu–Zn) isoenzymes in leaves and pseudobulbs of Bulbophyllum morphologlorum Kraenzl orchid by comparative proteomic analysis

Typically, biological systems are protected from the toxic effect of free radicals by antioxidant defense. Extracts from orchids have been reported to show high levels of exogenous antioxidant activity including Bulbophyllum orchids but so far, there have been no reports on antioxidant enzymes. Therefore, differences in protein expression from leaves and pseudobulbs of Bulbophyllum morphologlorum Kraenzl and Dendrobium Sonia Earsakul were studied using two-dimensional gel electrophoresis and mass spectrometry (LC/MS/MS). Interestingly, the largest group of these stress response proteins were associated with antioxidant defense and temperature stress, including superoxide dismutase (Cu–Zn) and heat shock protein 70. The high expression of this antioxidant enzyme from Bulbophyllum morphologlorum Kraenzl was confirmed by activity staining on native-PAGE, and the two Cu/Zn-SODs isoenzymes were identified as Cu/Zn-SOD 1 and Cu/Zn-SOD 2 by LC/MS/MS. The results suggested that Bulbophyllum orchid can be a potential plant source for medicines and natural antioxidant supplements.

Ameliorative effect of Magnesium Isoglycyrrhizinate on hepatic encephalopathy by Epirubicin

BackgroundThe purpose of the present study was to evaluate the protective effect of Magnesium Isoglycyrrhizinate (MI) on Epirubicin (EPI)-induced hepatic encephalopathy (HE) and explore its underlying mechanism. MethodsMice were divided randomly into groups for treatments as follows: control group, EPI group (Model group), EPI + MI (25, 50 mg/kg) group. Morris water maze test were conducted to evaluate the spatial learning and memory ability. The serum and hippocampus levels of oxidative stress or inflammation were uncovered with the detection of superoxide dismutase (SOD), malondialdehyde (MDA), and pro-inflammatory cytokines (IL-1β, IL-6, TNF-α). ResultsAs a result, treatment with MI effectively ameliorated the EPI-induced decline in the ability of spatial learning and memory. MI also significantly relieved the severity of oxidative stress or inflammation in serum and hippocampus, which was accompanied with regulating liver functional parameters. Western blot data demonstrated that administration of MI could regulate the redox-related expressions of Txnip, Trx, Nrf2, HO-1, p-IκB-α, p-NF-κB, Caspase-3, Caspase-9, Bax and Bcl-2 in EPI-stimulated hepatic encephalopathy (HE). And the potency of MI treatments on Nrf2, NF-κB expression was also confirmed with immunohistochemical analysis. ConclusionsTaken together, the protective effect of Magnesium Isoglycyrrhizinate on EPI-induced hepatic encephalopathy might be mediated via the Txnip/Nrf2/NF-κB signaling pathway.

Effects of Lyciumbarbarum polysaccharides on the damage to human endometrial stromal cells induced by hydrogen peroxide.

Previous studies have shown that Lyciumbarbarum polysaccharides (LBPs) serve an important role in antioxidant activity to protect the cells and tissues. However, the specific mechanism of LBPs in the prevention of endometrial damage remains to be elucidated. Using morphological observation, cell proliferation assay, the detection of superoxide dismutase (SOD) activity and the content of malondialdehyde (MDA) in cell culture supernatant fluid, the detection by western blot analysis and reverse transcription-quantitative polymerase chain reaction of the mRNA and protein expression levels of caspase‑3 and Bcl‑2 in endometrial stromal cells (ESCs), it was demonstrated that, invitro, hydrogen peroxide (H2O2)-induced death of ESCs, increased the content of MDA and decreased the activity of SOD, and decreased the expression of Bcl-2 and increased the expression of caspase‑3. LBPs can inhibit H2O2‑induced cell death of ESCs, decrease the content of MDA in ESCs and increase the activity of SOD, as well as increasing the expression of Bcl‑2 and decreasing the expression levels of caspase‑3. These findings suggested that LBPs can inhibit H2O2‑induced apoptosis of EECs and that LBPs are able to offer a significant protection against oxidative stress to ESCs.

GANRA-5 protects both cultured cells and mice from various radiation types by functioning as a free radical scavenger

The radio-protective effects of the oxazolone derivative chemical compound 4-(4-methoxy-3-methoxyphenyl-methyl)-2-phenyl- 5(4H)-oxazolone (GANRA-5) against different types of radiation including X-rays, carbon ion beams, microwaves and ultraviolet light (UV) were studied. Cell proliferation/cytotoxicity assay and colony-forming assay were conducted to evaluate the toxicity of GANRA-5. To test its influence on the induction of double-stranded break (DSB) formation and genomic instability, γH2AX focus-forming assay as well as cytokinesis-block micronucleus assay was utilized. Our results indicate that GANRA-5 exhibits low toxicity, while providing high radio-protective effects for MRC-5 cells against different types of radiation. We also found that GANRA-5 acts as a free radical scavenger. Our animal studies provided evidence that GANRA-5 significantly increases the survival rate of mice after X-ray irradiation. Analyses of hemogram, visceral index and detection of superoxide dismutase (SOD) and malondialdehyde (MDA) in the viscera indicate both low toxicity of GANRA-5, combined with its ability to shield radiation risk. In conclusion, our results suggest that GANRA-5 has the potential to be used as a safe and efficient radio-protectant.

Antioxidant activities of saponins extracted from Radix Trichosanthis: an in vivo and in vitro evaluation

BackgroundRadix Trichosanthis (RT), the dry root tuber of Trichosanthis kirilowii Maxim (Cucurbitaceae), is a traditional Chinese medicine. Although a wide range of saponin pharmacological properties has been identified, to our knowledge, this may be the first report to investigate the crude saponins from RT. The purpose of this study was to delineate the antioxidant activity both in vitro and in vivo by using ethyl acetate (EtOAc), n-butanol, and the mixture of n-butanol and EtOAc fractions.MethodsIn vitro antioxidant activity was detected by using DPPH free radical, hydrogen peroxide scavenging, and reducing power assays. After pretreatment with different fractions saponins at 2 mg/kg/d and 3 mg/kg/d of crude drug, respectively, an established CCl4 induced acute cytotoxicity model was used to evaluate the in vivo antioxidant potential by detection of superoxide dismutase (SOD), malonaldehyde (MDA), lactate dehydrogenase (LDH), and total antioxidant capacity (T-AOC) levels.ResultsThe in vitro assay showed that the antioxidant activity of all the three fractions was promising. The reducing power of the EtOAc and the mixture of n-butanol and EtOAc extracts increased in a dose dependent manner. However, both the n-butanol and the mixture of n-butanol and EtOAc fractions in low dose exhibited in a time dependent manner with prolonged reaction time. As for hydrogen peroxide scavenging capability, the n-butanol fraction mainly demonstrated a time dependent manner, whereas EtOAc fraction showed a dose dependent manner. However, in case of in vivo assay, an increase of SOD and T-AOC and decrease of MDA and LDH levels were only observed in n-butanol (2 mg/kg/d of crude drug) extracts pretreatment group.ConclusionsRT saponins in n-butanol fraction might be a potential antioxidant candidate, as CCl4-induced oxidative stress has been found to be alleviated, which may be associated with the time dependent manner of n-butanol saponins in a low dose. Further studies will be needed to investigate the active individual components in n-butanol extract, in vivo antioxidant activities and antioxidant mechanisms.

An improved photochemical method for the rapid spectrophotometric detection of superoxide dismutase

A sensitive and convenient method is described for estimating superoxide dismutase activity using a photochemical augmentation procedure. This method is applicable to both liquid assays and polyacrylamide gel electropherograms. The flux of superoxide is generated by illuminating a reaction mixture containing dianisidine and riboflavin by either a laser source or light from a fluorescent lamp. The oxidation of dianisidine, as sensitized by riboflavin, is enhanced by superoxide dismutase. The increase is linearly dependent on superoxide dismutase concentration. The photochemical reaction is allowed to proceed uninterrupted for a standardized optimum time and intensity of illumination and then terminated by addition of a buffer, ‘finibuf’, which stabilizes the chromophoric complex formed. This permits the spectrophotometric absorbance measurements of a number of samples collectively and also eliminates the interruption of illumination with the concomitant requirement of a spectrophotometer for constant recording of the absorbance. This method is of utility to both biochemists and clinicians.