Superoxide Dismutase Sod Research Articles

Change in the Physiological and Biochemical Aspects of Tomato Caused by Infestation by Cryptic Species of Bemisia tabaci MED and MEAM1.

Simple SummaryBemisia tabaci is a polyphagous pest with cryptic species that infest various agricultural crops worldwide. Among the cryptic species, MED and MEAM1 are the most invasive, causing large production losses due to the feeding and transmission of the virus. We aimed to analyze the influence of these insects on the physiology and biochemistry of tomato. We found that the cryptic species MED caused considerable reduction in CO2 assimilation rates, stomatal conductance, and instantaneous carboxylation efficiency. Furthermore, plants infested with MED showed high activity of the enzymes peroxidase and polyphenol oxidase, which are considered expressions of plant defense. In contrast, plants infested with MEAM1 showed low peroxidase activity, which may result in a less lignified feeding place.Infestation by Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) causes damage to tomatoes with production losses of up to 100%, affecting the physiological and biochemical aspects of host plants. The objective of this study was to analyze the influence of infestation of cryptic species of B. tabaci MED and MEAM1 on the physiological and biochemical aspects of tomato. Tomato plants ‘Santa Adélia Super’ infested with B. tabaci (MED and MEAM1), and non-infested plants were evaluated for differences in gas exchange, chlorophyll - a fluorescence of photosystem II (PSII), and biochemical factors (total phenols, total flavonoids, superoxide dismutase—SOD, peroxidase—POD, and polyphenol oxidase—PPO). Plants infested with B. tabaci MED showed low rates of CO2 assimilation and stomatal conductance of 55% and 52%, respectively. The instantaneous carboxylation efficiency was reduced by 40% in MED and by 60% in MEAM1 compared to the control. Regarding biochemical aspects, plants infested by MED cryptic species showed high activity of POD and PPO enzymes and total phenol content during the second and third instars when compared to control plants. Our results indicate that B. tabaci MED infestation in tomato plants had a greater influence than B. tabaci MEAM1 infestation on physiological parameters (CO2 assimilation rate (A), stomatal conductance (gs), and apparent carboxylation efficiency (A/Ci)) and caused increased activity of POD and PPO enzymes, indicating plant resistance to attack. In contrast, B. tabaci MEAM1 caused a reduction in POD enzyme activity, favoring offspring performance.

Lethal and sublethal effects of the emerging contaminant oxytetracycline on the embryo-larval development of Rhinella arenarum

The antibiotic oxytetracycline (OTC) is commonly used in animal production and can enter aquatic ecosystems, causing adverse effects on non-target species. The aim of this work was to evaluate the lethal and sublethal effects of OTC on the embryonic and larval period of Rhinella arenarum, through standardized bioassays and oxidative stress (catalase-CAT-, superoxide dismutase-SOD-, glutathione S-transferase-GST-, reduced glutathione-GSH- and lipid peroxidation-TBARS-), neurotoxicity (acetylcholinesterase-AChE- and butyrylcholinesterase-BChE-) and genotoxicity (micronuclei test) biomarkers. Mortality was time and stage dependent, being the embryos (504 h-LC50 = 64.04 mg/L) more sensitive than the larvae (504 h-LC50 = 97.74 mg/L). Alterations in the oxidative stress biomarkers were observed mainly in larvae: CAT, SOD and GST decreased and GSH increased significantly. In embryos, only GST decreased significantly. Also, OTC increased the AChE and BChE activities but did not increase the micronuclei frequency. This study shows evidence that the presence of OTC in the environment may have negative effects on amphibians.

Ecotoxicity of herbicide carfentrazone-ethyl towards earthworm Eisenia fetida in soil

Herbicides pose a potential threat to the soil biodiversity and health. Carfentrazone-ethyl (CE), a triazolinones herbicide, is increasingly used in agricultural production. Its non-target toxic effects on soil microorganisms and soil enzymes are reported recently. However, the sublethal toxicity of CE on soil invertebrates like earthworms is not yet known. Therefore, in this work, the sublethal toxic effects of CE (0.05, 0.5, and 5.0 μg/g in soil) on the soil earthworm (Eisenia fetida) were evaluated using a battery of biomarkers including reactive oxygen species (ROS), enzyme (superoxide dismutase-SOD, catalase-CAT, peroxidase-POD, and glutathione S-transferase-GST) activities, malondialdehyde (MDA) contents, histopathological and DNA damage. Results indicated that CE increased ROS contents, enzyme activities, and MDA contents in the short-time (14 d), thus, causing a slight oxidative stress to E. fetida. However, the toxic effects of CE on earthworms gradually disappeared after 14 days. The CE did not cause histopathological and DNA damage in earthworms. Integrated Biological Response index (IBR) indicated that both concentration and exposure time of CE regulated its sublethal toxicity on earthworms. In conclusion, herbicide CE is safe to soil invertebrate earthworms when applied at the recommended doses. Our results contribute to the current understanding of CE effects on soil earthworms, and can be useful in developing soil health strategies under agrochemical use.

Effects of transportation stress on antioxidation, immunity capacity and hypoxia tolerance of rainbow trout (Oncorhynchus mykiss)

Transportation is common and necessary in aquaculture practice. A series of physiological changes in fish are caused by transportation stress. The present study investigated the effects of transportation of rainbow trout Onchorhynchus mykiss. Juvenile rainbow trout were placed in plastic bags, and sampled before transportation (P1), two hours after transportation (P2) and 48 h after transportation (P3). The water temperature, dissolved oxygen (DO), ammonia nitrogen (NH4+-N), nitrate nitrogen (NO3--N) and nitrite nitrogen (NO2–-N) were analyzed. The relative expression of genes involved in stress and immunity (IL-1β, IL-8, TNF-α, LYZⅡ, HSP70 and HSP90), hypoxia tolerance (hif-1α, GLUT-1) and NF-kB pathway related (NF-kB, Ikb and Ikb2) were analyzed by RT-QPCR. In addition, oxidative damage, antioxidant and immune parameters (the activity of malondialdehyde MDA, superoxide dismutase SOD, catalase CAT and total antioxidant capacity T-AOC, acid phosphatase ACP, alkaline phosphatase AKP and total nitric oxide synthase NOS) were determined. The level of DO at P2 point was significantly decreased compared with P1 and P3 point. The NH4+-N concentration was significantly higher than that at P1 and P3. Further, there was a significant increase in HSP70 expression in gill and muscle during transportation. The expression of TNF-α, IL-1 β and IL-8 in gill and IL-1 β and IL-8 in muscle were significantly up regulated during transportation. The HSP70/90, TNF-α, IL-1 β and IL-8 genes can be used as bio-indicators to monitor stress. Moreover, ACP enzyme activity in liver and muscle, AKP in gill and MDA contents in gill and muscle were significantly higher at P2. The activity of NOS and T-AOC in gill was significantly higher at P3, and showed an increasing trend. In general, this study provides basial information of transportation, and will be helpful to choose more suitable conditions for successful transportation of juvenile rainbow trout.

Memantine Modulates Oxidative Stress in the Rat Brain following Experimental Autoimmune Encephalomyelitis.

Experimental autoimmune encephalomyelitis (EAE) is an animal model most commonly used in research on the pathomechanisms of multiple sclerosis (MS). The inflammatory processes, glutamate excitotoxicity, and oxidative stress have been proposed as determinants accompanying demyelination and neuronal degeneration during the course of MS/EAE. The aim of the current study was to characterize the role of NMDA receptors in the induction of oxidative stress during the course of EAE. The effect of memantine, the uncompetitive NMDA receptor antagonist, on modulation of neurological deficits and oxidative stress in EAE rats was analyzed using several experimental approaches. We demonstrated that the expression of antioxidative enzymes (superoxide dismutases SOD1 and SOD2) were elevated in EAE rat brains. Under the same experimental conditions, we observed alterations in oxidative stress markers such as increased levels of malondialdehyde (MDA) and decreased levels of sulfhydryl (-SH) groups, both protein and non-protein (indicating protein damage), and a decline in reduced glutathione. Importantly, pharmacological inhibition of ionotropic NMDA glutamate receptors by their antagonist memantine improved the physical activity of EAE rats, alleviated neurological deficits such as paralysis of tail and hind limbs, and modulated oxidative stress parameters (MDA, -SH groups, SOD’s). Furthermore, the current therapy aiming to suppress NMDAR-induced oxidative stress was partially effective when NMDAR’s antagonist was administered at an early (asymptomatic) stage of EAE.

Genotoxicity and behavioral alterations induced by retene in adult zebrafish

Retene (1-methyl-7-isopropylphenanthrene; RET), a non-priority polycyclic aromatic hydrocarbon (PAH), is one of the most widely produced PAHs following cellulose burning. Despite RET toxicity observed in various aquatic organisms, the underlying mechanisms remain unclear. In this study, the genotoxic and neurotoxic effects of RET were investigated using zebrafish as experimental model. Adult zebrafish were exposed to 250, 500, 750, and 1000 µg/L RET for 96 h. RET increased micronuclei frequency, demonstrating its genotoxic potential. After RET exposure, adult zebrafish exhibited alterations in locomotion and exploration endpoints. RET also increased the levels of the reactive oxygen species (ROS) in the zebrafish brains, indicating an imbalance in the redox status. Moreover, in its initial concentration, the mRNA levels of superoxide dismutase (sod1 and sod2), glutathione-disulfide reductase (gsr), glutathione peroxidase 1 A (gpx1a) were up-regulated; however, in the two highest RET concentrations, the transcriptional level of nuclear factor erythroid 2 like 2 (nfe2l2) was down-regulated, leading to the reduction of mRNA levels of all the antioxidant defense genes investigated. Furthermore, RET decreased the mRNA levels of genes encoding components of the cholinergic, serotoninergic, and gamma-amino butyric acid (GABA) neurotransmitter systems. Taken together, these results demonstrate that RET affects DNA generating micronuclei and provide new evidence suggesting that the behavioral alterations in zebrafish could be due to changes in oxidative stress and the mRNA expression of the neurotransmitter systems. These results reinforce the risk posed by environmental pollutants to aquatic ecosystems, especially those whose toxic potentials remain underestimated.

Efficacy of Ulinastatin and Sulforaphane Alone or in Combination in Rat Model of Streptozotocin Diabetes Induced Vascular Dementia.

ObjectiveVascular Dementia (VaD), is associated with metabolic conditions. Diabetes is a major risk factor for the development of VaD. This study investigates the efficacy of ulinastatin (UTI) and sulforaphane (SUL) in streptozotocin (STZ)-diabetes induced vascular endothelium dysfunction and related dementia.MethodsSingle dose STZ (50 mg/kg i.p.) was administered to Albino Wistar rats (male, 200−250 g). Morris water maze and attentional set shifting tests were used to assess the spatial learning, memory, reversal learning, and executive functioning in animals. Body weight, serum glucose, serum nitrite/nitrate, vascular endothelial function, aortic superoxide anion, brains’ oxidative markers (thiobarbituric acid reactive species-TBARS, reduced glutathione-GSH, superoxide dismutase-SOD, and catalase-CAT), inflammatory markers (IL-6, IL-10, TNF-a, and myeloperoxidase-MPO), acetylcholinesterase activity-AChE, blood brain barrier (BBB) permeability and histopathological changes were also assessed. UTI (10,000 U/kg) and SUL (25 mg/kg) were used alone as well as in combination, as the treatment drugs. Donepezil (0.5 mg/kg) was used as a positive control.ResultsSTZ-administered rats showed reduction in body weight, learning, memory, reversal learning, executive functioning, impairment in endothelial function, BBB permeability, increase in serum glucose, brains’ oxidative stress, inflammation, AChE-activity, BBB permeability and histopathological changes. Administration of UTI and SUL alone as well as in combination, significantly and dose dependently attenuated the STZ-diabetes-induced impairments in the behavioral, endothelial, and biochemical parameters.ConclusionSTZ administration caused diabetes and VaD which was attenuated by the administration of UTI and SUL. Therefore, these agents may be studied further for the assessment of their full potential in diabetes induced VaD.

Effects of methanolic leaf extract of Kigelia africana on hemodynamic parameters and redox status of normotensive wistar rats

Kigelia Africana has a rich history of ethnobotanical usage in the treatment of a wide range of illnesses andcomplications like hypertension, cancer and microbial infections in south western Nigeria. The present work sought to investigate the effects of methanolic leaf extract of Kigelia Africana on the blood pressure and biochemical indices of normotensive albino rats. Twenty five male albino Wistar rats, weighing between 180-200g, and divided into five groups (I-V) of five animals each were used for the experiment. GroupsII – V were administered 1mg/kg b.w ramipril, 25-, 50- and 100mg/kg b.wKigelia Africana respectively by oral gavage for 14 consecutive days. Group I received vehicle (1 mg/kg distilled water) only throughout the duration of the experiment and served as control. Twenty-four hours after the last administration, the blood pressure was determined before sacrificing the animals following anesthesia. The activities of serum biomarkers (aspartate aminotransferase AST, total protein TP, alanine aminotransferase ALT, alkaline phosphatase ALP), serum lipid profile; (cholesterol TC, low/very low density lipoprotein LDL-c/VLDL-c, high density lipoprotein HDL and triglyceride TG) and cardiac antioxidant indices (catalase CAT, superoxide dismutase SOD and glutathione GSH) were determined. Administration of ramipril (1mg/kg b.w) and all dosages of Kigelia africana (25, 50 and 100mg/kg b.w) caused significant (P<0.05) decrease in the blood pressure of the animals when compared to the control. The values obtained further showed that ramipril and the extract (at all dosages) caused significant reduction in TC, TG, LDL-c and VLDL-c and coronary risk index, CRI while there was significant increase in the level of HDL when compared to the control. All dosages of Kigelia africana extract resulted in significant (P<0.05) decrease in serum activities of AST, ALT and ALP when compared with the control. The significant (P<0.05) increases in cardiac catalase activity and GSH concentration recorded in rats treated with Kigelia africana (25, 50 and 100mg/kg b.w.) were comparable with those administered the reference drug, ramipril (1 mg/kg). Cardiac SOD activity was however decreased in rats administered extract (25, 50 or 100 mg/kg b.wt) and ramipril (1mg/kg bw). The results suggest the hypolipidemic, hypotensive and antioxidant properties and of methanolic extract of Kigelia africana leaf and lend support to the ethnobotanical usage of the leaf in the treatment of hypertension.

1-Methylcyclopropene Preserves the Quality of Chive (Allium schoenoprasum L.) by Enhancing Its Antioxidant Capacities and Organosulfur Profile during Storage.

The quality, antioxidant capacities, and organosulfur profile of chives (Allium schoenoprasum L.) treated with 1-methylcyclopropene (1-MCP) during storage were investigated in this study. The 1-MCP treatment (100 μL/L, fumigation 12 h at 20 °C) effectively inhibited tissue respiration and H2O2 production, enhanced the ascorbic acid (ASA) and glutathione (GSH) content, and promoted the activity of antioxidant enzymes (superoxide dismutase SOD, Catalase CAT, and ascorbic peroxidase APX) during the 5-day storage period at 20 °C. The result further showed that the 1-MCP treatment inhibited chlorophyll degradation, alleviated cell membrane damage, and delayed the chive senescence, with the yellowing rate being reduced by 67.8% and 34.5% in the 1-MCP treated chives on days 4 and 5 of storage at 20 °C, respectively. The free amino acid content of the chive was not affected by the 1-MCP treatment at 20 °C. However, the senescence rate of the chive was not reduced by the 1-MCP treatment when stored at 3 °C. The liquid chromatography data further showed that the 1-MCP treatment induced a 15.3% and 13.9% increase in the isoalliin and total S-alk(en)ylcysteine sulfoxides (ACSOs) content of the chive on day 2 at 20 °C, respectively. Furthermore, there was a strong positive correlation between ACSOs content and CAT/APX activity, indicating that ACSOs probably played a key role in enhancing the antioxidant capacities of the chive during storage at 20 °C. Thus the study efficiently demonstrates that 1-methylcyclopropene preserves the quality of chive (Allium schoenoprasum L.) by enhancing its antioxidant capacities and organosulfur profile during storage.

Heat stress-mediated effects on the morphophysiological, biochemical, and ultrastructural parameters of germinating Melanoxylon brauna Schott. seeds.

The present study showed that the heat stress (40°C) caused changes in morphophysiological, biochemical, and ultrastructural parameters to the seeds Melanoxylon brauna, ultimately leading to loss of germination capacity. Temperature is an abiotic factor that influences seed germination. In the present study, we investigated morphophysiological, biochemical, and ultrastructural changes during the germination of Melanoxylon brauna seeds under heat stress. Seed germination was evaluated at constant temperatures of 25 and 40°C. The samples consisted of seeds soaked in distilled and ionized water for 48 and 96h at both temperatures. For the evaluation of internal morphology, the seeds were radiographed. Ultrastructural parameters were assessed using transmission electron microscopy (TEM). The production of reactive oxygen species (ROS), content of malondialdehyde (MDA) and glucose, carbonylated proteins, and activity of the enzymes (superoxide dismutase-SOD, ascorbate peroxidase-APX, catalase-CAT, peroxidase-POX, glucose-6-phosphate dehydrogenase-G6PDH, lipase, α- and β-amylase, and protease) were measured by spectrophotometric analysis. An 82% reduction in the germination of M. brauna seeds was observed at 25°C, and 0% at 40°C. TEM showed that seeds submitted to heat stress (40°C) had poorly developed mitochondria and significantly reduced respiration rates. The content of ROS and protein carbonylation in seeds subjected to 40°C increased compared to that at 25°C. The activity of antioxidant enzymes, namely SOD, APX, CAT, and POX, was significantly reduced in seeds subjected to heat stress. Glucose content, G6PDH, and lipase activity also decreased when the seeds were exposed to heat stress. Conversely, α- and β-amylase enzymes and the protease increased due to the increase in temperature. Our data showed that the increase in temperature caused an accumulation of ROS, increasing the oxidative damage to the seeds, which led to mitochondrial dysfunction, ultimately leading to loss of germination.

The Copper Chaperone CcsA, Coupled with Superoxide Dismutase SodA, Mediates the Oxidative Stress Response in Aspergillus fumigatus.

Superoxide dismutases (SODs) are important metalloenzymes that protect fungal pathogens against the toxic effects of reactive oxygen species (ROS) generated by host defense mechanisms during the infection process. The activation of Cu/Zn-SOD1 is found to be dependent on copper chaperone for SOD1 (Ccs1). However, the role of the Ccs1 ortholog in the human pathogen Aspergillus fumigatus and how these SODs coordinate to mediate oxidative stress response remain elusive. Here, we demonstrated that A. fumigatus CcsA, a Saccharomyces cerevisiae Ccs1 ortholog, is required for cells in response to oxidative response and the activation of Sod1. Deletion of ccsA resulted in increased ROS accumulation and enhanced sensitivity to oxidative stress due to the loss of SodA activity. Molecular characterization of CcsA revealed that the conserved CXC motif is required not only for the physical interaction with SodA but also for the oxidative stress adaption. Notably, addition of Mn2+ or overexpression of cytoplasmic Mn-SodC could rescue the defects of the ccsA or sodA deletion mutant, indicating the important role of Mn2+ and Mn-SodC in ROS detoxification; however, deletion of the CcsA-SodA complex could not affect A. fumigatus virulence. Collectively, our findings demonstrate that CcsA functions as a Cu/Zn-Sod1 chaperone that participates in the adaptation to oxidative stress in A. fumigatus and provide a better understanding of the CcsA-SodA complex-mediated oxidative stress response in filamentous fungi. IMPORTANCE Reactive oxygen species (ROS) produced by phagocytes have been reported to participate in the killing of fungal pathogens. Superoxide dismutases (SODs) are considered to be the first line of defense against superoxide anions. Characterizing the regulatory mechanisms of SOD activation is important for understanding how fungi adapt to oxidative stress in hosts. Our findings demonstrated that CcsA functions as a SodA chaperone in A. fumigatus and that the conserved CXC motif within CcsA is required for its interaction with SodA and the CcsA-SodA-mediated oxidative response. These data may provide new insights into how fungal pathogens adapt to oxidative stress via the CcsA-SodA complex.

Behavioral and biochemical investigations to explore the efficacy of quercetin and folacin in experimental diabetes induced vascular endothelium dysfunction and associated dementia in rats.

Vascular dementia (VaD), being strongly associated with metabolic conditions is a major health concern around the world. Diabetes is a major risk factor for the development of VaD. This study investigates the efficacy of quercetin and folacin in diabetes induced vascular endothelium dysfunction and related dementia. Single dose streptozotocin (STZ) (50mg/kg i.p) was administered to albino Wistar rats (male, 200-250g) by dissolving in citrate buffer. Morris water maze (MWM) and attentional set shifting tests were used to assess the spatial learning, memory, reversal learning, and executive functioning in animals. Body weight, serum glucose, serum nitrite/nitrate, vascular endothelial function, aortic superoxide anion, brains' oxidative markers (thiobarbituric acid reactive species-TBARS, reduced glutathione-GSH, superoxide dismutase-SOD, and catalase-CAT), mitochondrial enzyme complex (I, II, and IV), inflammatory markers (interleukin-IL-6, IL-10, tumor necrosis factor-TNF-α, and myeloperoxidase-MPO), and acetylcholinesterase activity-AChE were also assessed. Quercetin (30mgkg-1/60mgkg-1) and folacin (30mgkg-1/60mgkg-1) were used as the treatment drugs. Donepezil (0.5mgkg-1) was used as a positive control. STZ administered rats showed reduction in learning, memory, reversal learning, executive functioning, impairment in endothelial function, increase in brains' oxidative stress; inflammation; AChE activity, and decrease in mitochondrial complex (I, II, and IV) activity. Administration of quercetin and folacin in two different doses, significantly attenuated the STZ induced diabetes induced impairments in the behavioral, endothelial, and biochemical parameters. STZ administration caused diabetes and VaD which was attenuated by the administration of quercetin and folacin. Therefore, these agents may be studied further for the assessment of their full potential in diabetes induced VaD conditions.

Inhibitory effect of nano selenium on the recurrence of Aeromonas hydrophila bacteria in Cyprinus carpio

The recurrence of bacterial infection in fish is related to several factors, including the immune status and oxidative state. A total number of 240 Cyprinus carpio was divided into four treatments; control (T1) fed on a free diet and three treatments (T2–4) fed on supplemented diet with nanoparticles of selenium Se NPs (1 mg/kg fish feed) for 10, 20, and 30 days, respectively. Supplemented fish with Se NPs for 30 days (T4) had significantly higher total protein and globulin content (5.83 and 3.1 g/dl, respectively) compared to the control indicating an enhanced immune status. Fish fed on Se NPs supplementation for 30 days (T4) recorded significantly higher anti-inflammatory cytokine “interleukin (IL)-10”, while the pro-inflammatory cytokines “tumor necrosis factor (TNF)–α, IL-1β, and IL-8” reduced significantly compared to the control. Antioxidant enzymes (glutathione peroxidase GPx, catalase CAT, and superoxide dismutase SOD) were gradually and significantly enhanced in a time-dependent manner. Genotoxicity (DNA damages, DNA fragmentation, and presence of micronuclei) was low in supplemented fish while challenged fish performed in an opposite manner. However, Se NPs supplementation had overcome those withdraws causing a decrease in the signs of genotoxicity. Se NPs improved the immunity, antioxidant, and histopathological status of C. carpio showing enhancements in disease resistance, as reducing the chance of recurrence of Aeromonas hydrophila infection and genotoxicity.

Sex Differences in the Relation Between Frailty and Endothelial Function in Old Mice

Frailty is an accumulation of deficits that stem from impairments in multiple organ systems, potentially including the vasculature. Older women typically have higher frailty scores than older men, but it is unknown if there are sex differences in the relation of frailty to physiological function. Endothelial function declines with age on average, but there is a high variability in the amount of decline. Thus, we sought to determine sex differences in the relation of frailty to endothelial dysfunction among old mice. In young male (n= 24, 8.5±0.27 mo), young female (n=20, 4.9±0.01 mo), old male (n=32, 28±0.56 mo) and old female (n= 17, 25.8±0.47 mo) C57BL/6 mice, we evaluated frailty by a 31-item frailty index (FI) based on established signs of deterioration. We assessed endothelial function by endothelium-dependent dilation (EDD, maximal response to acetylcholine) ex vivo in isolated, pressurized mesentery arteries and middle cerebral arteries (MCAs). We also assessed EDD after incubation with the superoxide scavenger TEMPOL and endothelium-independent dilation (EID, maximal response to sodium nitroprusside). We quantified gene expression for pro-inflammatory interleukin-1β (Il1b), pro-oxidant NADPH oxidase 2 (Nox2), and antioxidants superoxide dismutases (Sod1, Sod2, and Sod3) in mesentery arteries. Old mice had a greater FI compared with young mice (old: 5.53±2.3, young: 1.01±0.67, p=0.003), but FI did not differ between old male and female mice (p=0.41). When specific FI components were examined, old female mice had higher scores for alopecia, rectal prolapse, and cataracts but lower scores for vaginal/penile prolapse and grimace scale compared with old male mice (p<0.05). Old mice had a 25% impairment in MCA EDD and 16% impairment in mesentery artery EDD compared with young mice (p<0.05). While EDD did not differ between arteries from old male and female mice (p>0.05), there were sex differences in the relations between FI and EDD. For the MCA, FI was inversely related to EDD among old female (r=-0.49, p=0.02), but not old male mice (r=0.15, p=0.24). In contrast, for the mesentery artery, FI was inversely related to EDD among old males (r=-0.42, p=0.01), but not old female mice (r=0.37, p=0.07). EID did not differ with sex or age group (p>0.05). A higher FI was related to a greater improvement in EDD with TEMPOL in the MCA in only old female mice and in the mesentery arteries in only old male mice (p<0.05). In mesentery arteries from old compared with young mice, gene expression was 42% higher for Il1b, 34% lower for Sod2 and 54% lower for Sod3 (p<0.05), but not different between ages for Sod1 or Nox2 (p>0.05). Among old mice, Il1b expression was23% higher in mesenteries from old male compared with female mice (p=0.001), but sex differences were not present for the other genes (p>0.05). In summary, we found no sex differences for composite FI scores or endothelial impairment among old mice. However, there are sex differences in the individual components of FI and in the relations of frailty to endothelial function, and these may be mediated by sex differences in arterial oxidative stress and pro-inflammatory signaling.

Reactive oxygen species production and the influence of exogenous copper in yeast mitochondria

Mitochondria are essential organelles within eukaryotic cells due to their role in metabolism, ATP production via the oxidative phosphorylation (OXPHOS) pathway, and other regulatory cellular processes. Our aim is to understand how copper in the mitochondria of yeast affects the production of reactive oxygen species due to its influence on the electron transport chain (ETC) and the localization of reactive oxygen species production during yeast lifespan. Reactive oxygen species (ROS) provide signaling in the cells of organisms which is beneficial for cells to maintain homeostasis. However, when there is a surplus of ROS this can lead to harmful and/or lethal results for the cell. To detect ROS, yeast cultures were grown for one to three days and treated with the fluorescent ROS indicator dihydroethidium (DHE), as well as the fluorescent indicators SYTO and DAPI which stain mitochondrial DNA and adenine-thymine rich regions in the DNA respectively. The ROS generated from the yeast was then analyzed via a fluorescence plate reader, fluorescent activated cell sorting (FACS), and fluorescent microscopy to compare methods of determining ROS in the samples. To investigate mitochondrial ROS production, cells were grown in the presence of antimycin A (a Complex III inhibitor), oligomycin (a Complex V inhibitor), or menadione (a futile redox cycling molecule). Detectable increases in ROS were observed through the inhibition of Complex III and in the presence of menadione. We are now using these methods in yeast with a superoxide dismutase 1 (SOD1) gene deletion, to understand the influence of exogenous copper on the generation of ROS. Copper is worth investigating in conjunction with ROS because the mitochondria in mammals and yeast cells contain a labile copper pool located in the matrix, which is used in the metalation of the copper containing enzymes Complex IV, Cytochrome C Oxidase(CcO), and superoxide dismutase (Sod1). It has been established that supplementing cultures with copper increases CcO and Sod1 activity, supporting previous results that exogenous copper contributes to a labile copper pool within the mitochondria to increase enzyme activity. Our results show that small amounts exogenous copper (0.25 mM copper sulfate in restricted nutrient media) extend yeast chronological lifespan, but a dramatic reduction in lifespan is observed when copper levels are increased to 2.0 mM copper sulfate or higher. These data indicates that low levels of exogenous copper in the media is beneficial for yeast in restricted media conditions. Our current work assesses how exogenous copper changes ETC protein complex expression, specifically looking at CcO subunits, and the production of ROS under these conditions.

SOD3 Is a Non-Mutagenic Growth Regulator Affecting Cell Migration and Proliferation Signal Transduction.

Superoxide dismutase (SOD) family isoenzymes, SOD1, SOD2, and SOD3, synthesize hydrogen peroxide (H2O2), which regulates the signal transduction. H2O2 is a second messenger able to enter into the cells through aquaporin 3 cell membrane channels and to modify protein tyrosine phosphatase activity. SOD3 has been shown to activate signaling pathways in tissue injuries, inflammation, and cancer models. Similar to the H2O2 response in the cells, the cellular response of SOD3 is dose-dependent; even a short supraphysiological concentration reduces the cell survival and activates the growth arrest and apoptotic signaling, whereas the physiological SOD3 levels support its growth and survival. In the current work, we studied the signaling networks stimulated by SOD3 overexpression demonstrating a high diversity in the activation of signaling cascades. The results obtained suggest that SOD3, although inducing cell growth and affecting various biological processes, does not cause detectable long-term DNA aberrations. Therefore, according to the present data, SOD3 is not a mutagen. Additionally, we compared SOD3-driven immortalized mouse embryonic fibroblasts to SV40 immortalized NIH3T3 cells, demonstrating a marked difference in the activation of cellular kinases. The data presented may contain important druggable targets to abrogate unwanted cell growth.

Functional genomics of a Spiroplasma associated with the carmine cochineals Dactylopius coccus and Dactylopius opuntiae

BackgroundSpiroplasma is a widely distributed endosymbiont of insects, arthropods, and plants. In insects, Spiroplasma colonizes the gut, hemolymph, and reproductive organs of the host. Previous metagenomic surveys of the domesticated carmine cochineal Dactylopius coccus and the wild cochineal D. opuntiae reported sequences of Spiroplasma associated with these insects. However, there is no analysis of the genomic capabilities and the interaction of this Spiroplasma with Dactylopius.ResultsHere we present three Spiroplasma genomes independently recovered from metagenomes of adult males and females of D. coccus, from two different populations, as well as from adult females of D. opuntiae. Single-copy gene analysis showed that these genomes were > 92% complete. Phylogenomic analyses classified these genomes as new members of Spiroplasma ixodetis.Comparative genome analysis indicated that they exhibit fewer genes involved in amino acid and carbon catabolism compared to other spiroplasmas. Moreover, virulence factor-encoding genes (i.e., glpO, spaid and rip2) were found incomplete in these S. ixodetis genomes. We also detected an enrichment of genes encoding the type IV secretion system (T4SS) in S. ixodetis genomes of Dactylopius. A metratranscriptomic analysis of D. coccus showed that some of these T4SS genes (i.e., traG, virB4 and virD4) in addition to the superoxide dismutase sodA of S. ixodetis were overexpressed in the ovaries.ConclusionThe symbiont S. ixodetis is a new member of the bacterial community of D. coccus and D. opuntiae. The recovery of incomplete virulence factor-encoding genes in S. ixodetis of Dactylopius suggests that this bacterium is a non-pathogenic symbiont. A high number of genes encoding the T4SS, in the S. ixodetis genomes and the overexpression of these genes in the ovary and hemolymph of the host suggest that S. ixodetis use the T4SS to interact with the Dactylopius cells. Moreover, the transcriptional differences of S. ixodetis among the gut, hemolymph and ovary tissues of D. coccus indicate that this bacterium can respond and adapt to the different conditions (e.g., oxidative stress) present within the host. All this evidence proposes that there is a strong interaction and molecular signaling in the symbiosis between S. ixodetis and the carmine cochineal Dactylopius.

ACELLULAR LIVER SCAFFOLD PROMOTES CELL RECRUITMENT AFTER HETEROTOPIC TRANSPLANTATION

<h3>Background</h3> Liver tissue engineering is a promising tool to solve the shortage of organs. In this context, acellular liver scaffolds (ALS) obtained by decellularization are an attractive substrate to organ transplantation. However, the cell recruitment proprieties after in vivo transplantation (Tx) remain unclear. <h3>Objective</h3> To address this issue, we aimed to investigate the ALS in vivo cell recruitment proprieties after heterotopic auxiliary liver transplantation in rats. <h3>Methods</h3> Donor Wistar rats (n=4) were submitted to a surgical procedure to promote liver procurement. For decellularization, the livers were perfused through the portal vein (PV) using an infusion pump at 3 ml/min with water for 2 hours followed by 1% Triton X-100 for 2 hours and SDS 1% for 18-24h. After decellularization, livers were washed with water for 2 days and submitted to DNA quantification and histology analysis (H&E and Sirius red). Then, to transplant ALS, recipient rats (n=3) were submitted to a surgical procedure to remove the left-side kidney. The PV and inferior vena cava of the ALS were anastomosed to the recipient rats' left arterial and renal vein respectively. Recipients rat serum biochemical analysis (ALB- albumin) was measured before and post-Tx. H&E, Periodic acid–Schiff, TUNEL and, immunohistochemistry (Ki67, Superoxide dismutase-SOD, Catalase-CAT, Glutathione-peroxidase-GPX) analyzes were performed to evaluate ALS post-Tx. Data were expressed as mean ± SD and all quantitative parameters were analyzed using Student's t-test. <h3>Results</h3> DNA quantification analysis showed a significant reduction (1813.8±125.6 to 19.2±8,03 ng DNA/mg dry weight) and H&E staining showed that the decellularization process removed the cells and preserves the structure and components of the ALS. Active blood flow within the ALS was observed indicating that the vessels were able to sustain the arterial blood pressure when the circulation was re-established. H&E analysis revealed that ALS was able to promote cell recruitment and to allow migration and distribution of Ki67, SOD, CAT and, GPX positive cells in whole ALS parenchyma 2 hours post-Tx. Also, no glycogen storage was observed into ALS 2h post-Tx. ALB serum analysis showed no adverse impact on recipient rat biochemical parameters (2.17±0.05 vs.1.8± 1.11). <h3>Conclusion</h3> In conclusion, ALS is a promising approach to liver transplantation able to promote various cell type recruitment, distribution, and whole recellularization in vivo.

Selection and validation of reference genes for gene expression studies in Xanthomonas arboricola pv. juglandis subjected to abiotic stress

AbstractThe selection and validation of reference genes for reverse‐transcription quantitative PCR (RT‐qPCR) is key in the normalization of data to evaluate the expression levels of target genes. In this study, the expression levels of nine candidate reference genes in Xanthomonas arboricola pv. juglandis (Xaj) undergoing three abiotic stresses, including thermal stress at 15 ± 1°C and 35 ± 1°C and UV light, were evaluated by RT‐qPCR. A ranked list of reference genes based on expression stability was assessed using BestKeeper, NormFinder, GeNorm, and ΔCt methods. In Xaj undergoing all tested abiotic stress conditions, two reference genes (RGs) were sufficient for effective normalization of RT‐qPCR data. ATP synthase subunit δ (atpD) and RNA polymerase δ factor (rpoD) were the most stably expressed RGs, while gyrase subunit A (gyrA) and gyrase subunit B (gyrB) were the least stable. A combination of the most and least stable RGs was used for the normalization of two isoforms of three target genes, catalase (cat1 and cat2), superoxide dismutase (sod1 and sod2), and glutathione peroxidase (gpx1 and gpx2), showing that the validation of RGs was needed in each experimental condition to avoid misinterpretation of the expression levels of target genes. These results can be useful in studies that seek to understand the biochemical and molecular complex pathways involved in stress tolerance mechanisms in Xaj and to evaluate the effects of stress conditions on plant–pathogen interactions.

Biochemical and histopathological effects of low dose vanadium in the healing of acetic acid-induced colitis in male wistar rats.

Vanadium has been reported to possess relevant therapeutic properties such as anti-diabetic and anti-tumoral. This study aimed at determining the effects of vanadium on experimentally induced colitis in rats. Forty-five male Wistar rats (103±3.90g, n=15) were used for this study and were divided into three groups. Group 1 (Untreated control) had nothing added to their drinking, while groups 2 and 3 received sodium metavanadate at a dose of 50 and 200mg/L respectively in their drinking water for 10weeks. Colitis was thereafter induced by intra colonic administration of 1.50mL of 6% acetic acid. Animals were sacrificed on day 0 (pre-induction), three- and seven-days post induction. Blood samples were collected for haematological variables and the distal 8cm of the colon was collected for macroscopic, histological and biochemical (malondialdehyde-MDA, superoxide dismutase-SOD, catalase-CAT, glutathione peroxidase- GPx and nitrite concentration- NO) assessment. Low dose vanadium proved beneficial in ameliorating acetic acid-induced colitis by improving both histopathological and haematological changes. Gross observation showed a faster healing rate in vanadium treated groups (50and 200mg/L) compared with untreated control at day 3 (40 and 26.20 vs. 2.50%) and day 7 (80 and 66.70 vs. 42%) respectively. Vanadium also appears to exert its beneficial effects on acetic acid-induced colitis via up regulation of antioxidant enzymes (SOD, CAT, GPx) and NO while decreasing the over production of MDA. Vanadium at small concentration functions as an essential trace element and may be able to promote healing process during ulcerative colitis.