Nanobiotechnological basis of an oxygen carrier with enhanced carbonic anhydrase for CO2 transport and enhanced catalase and superoxide dismutase for antioxidant function.

This is a mini review on the biotechnological aspects of the most extensively developed hemoglobin-based oxygen carriers The emphasis is on the most recent Polyhemoglobin-catalase-superoxide dismutase-carbonic anhydrase (PolyHb-CAT-SOD-CA), which is a nanobiotechnological complex that is being investigated and scaled up with the potential for clinical use as nanobiotherapeutics. Hemoglobin, a tetramer, is an excellent oxygen carrier. However, in the body it is converted into toxic dimers. Diacid or glutaraldehyde can crosslink hemoglobin into polyhemoglobin (PolyHb) and prevent its breakdown into toxic dimers. This has been developed and tested in clinical trials. A bovine polyhemoglobin has been approved for routine clinical use for surgical procedures in South Africa and Russia. Clinical trials with human PolyHb in hemorrhagic shock were effective but with a very slight increase in non-fatal myocardial ischemia. This could be due to a number of reasons. For those conditions with ischemia-reperfusion, one would need an oxygen carrier with antioxidant properties. One approach to remedy this is with prepared polyhemoglobin-catalase-superoxide dismutase (PolyHb-CAT-SOD). Another reason is an increase in intracellular pCO2. We therefore added an enhanced level of carbonic anhydrase to prepare a PolyHb-CAT-SOD-CA. The result is an oxygen carrier with enhanced Carbonic Anhydrase for CO2 transport and enhanced Catalase and Superoxide Dismutase for antioxidant functions. Detailed efficacy and safety studies have led to the industrial scale up towards clinical trial. In the meantime, oxygen carriers are being investigated around the world for use in ex vivo biotechnological fluid for organ preservation for transplantation, with one already approved in France.

Assessment of heavy metals and associated oxidative stress in occupationally exposed workers from Bannu and Karak Districts in Pakistan.

Heavy metals (HMs) are extensively found in occupationally exposed miners and industrial workers, which may cause serious health-related problems to the large workforce. In order to evaluate the impact of these toxic pollutants, we have investigated the effect of cadmium (Cd), chromium (Cr), copper (Cu), and lead (Pb) concentration on exposed workers of mining, and woolen textile mill and compared the findings with unexposed individuals. From each category like exposed workers (mining, and woolen mill textile site) and unexposed individuals, 50 blood samples were taken. The occurrence of HMs in a sample was investigated through atomic absorption spectrometry while the oxidative stress marker malondialdehyde (MDA) and antioxidant enzyme statuses such as superoxide dismutase (SOD) and catalase (CAT) were analyzed in exposed and control samples. The results showed significant (p < 0.05) variation in Cd, Cr, Cu, and Pb levels in exposed and control samples. The concentration of Cd in the blood of WMWs, KMWs, and control group was 5.75, 3.89, and 0.42 µg/dL, respectively. On the other hand, the concentration of Pb in the blood of WMWs, MWs, and control was 32.34, 24.39, and 0.39 µg/dL while the concentrations of Cr and Cu in the blood of WMWs, MWs, and control group were 11.61 and 104.14 µg/dL, 4.21 and 113.21 µg/dL, 0.32 and 65.53 µg/dL, respectively. An increase in MDA was recorded in the exposed workers' group as compared to control subjects, whereas SOD and CAT activities decreased. Meanwhile, MDA was significantly and positively (p < 0.01) correlated with HMs, while negative significant correlations were found among HMs with SOD and CAT.

Rise and fall of peroxisomes during Alzheimer´s disease: a pilot study in human brains.

Peroxisomes are eukaryotic organelles that rapidly change in number depending on the metabolic requirement of distinct cell types and tissues. In the brain, these organelles are essential for neuronal migration and myelination during development and their dysfunction is associated with age-related neurodegenerative diseases. Except for one study analysing ABCD3-positive peroxisomes in neurons of the frontal neocortex of Alzheimer disease (AD) patients, no data on other brain regions or peroxisomal proteins are available. In the present morphometric study, we quantified peroxisomes labelled with PEX14, a metabolism-independent peroxisome marker, in 13 different brain areas of 8 patients each either with low, intermediate or high AD neuropathological changes compared to 10 control patients. Classification of patient samples was based on the official ABC score. During AD-stage progression, the peroxisome density decreased in the area entorhinalis, parietal/occipital neocortex and cerebellum, it increased and in later AD-stage patients decreased in the subiculum and hippocampal CA3 region, frontal neocortex and pontine gray and it remained unchanged in the gyrus dentatus, temporal neocortex, striatum and inferior olive. Moreover, we investigated the density of catalase-positive peroxisomes in a subset of patients (> 80 years), focussing on regions with significant alterations of PEX14-positive peroxisomes. In hippocampal neurons, only one third of all peroxisomes contained detectable levels of catalase exhibiting constant density at all AD stages. Whereas the density of all peroxisomes in neocortical neurons was only half of the one of the hippocampus, two thirds of them were catalase-positive exhibiting increased levels at higher ABC scores. In conclusion, we observed spatiotemporal differences in the response of peroxisomes to different stages of AD-associated pathologies.

Enhanced enzymatic activity and conformational stability of catalase in presence of tetrahedral DNA nanostructures: A biophysical and kinetic study.

The emergence of DNA nanotechnology has shown enormous potential in a vast array of applications, particularly in the medicinal and theranostics fields. Nevertheless, the knowledge on the biocompatibility between DNA nanostructures and cellular proteins is largely unknown. Herein, we report the biophysical interaction between proteins (circulatory protein bovine serum albumin, BSA, and the cellular enzyme bovine liver catalase, BLC) and tetrahedral DNA (tDNA), which are well-known nanocarriers for therapeutics. Interestingly, the secondary conformation of BSA or BLC was unaltered in the presence of tDNAs which supports the biocompatible property of tDNA. In addition, thermodynamic studies showed that the binding of tDNAs with BLC has a stable non-covalent interaction via hydrogen bond and van der Waals contact, which is indicative of a spontaneous reaction. Furthermore, the catalytic activity of BLC was increased in the presence of tDNAs after 24 h of incubation. These findings indicate that the presence of tDNA nanostructures not only ensures a steady secondary conformation of proteins, but also stabilize the intracellular proteins like BLC. Surprisingly, our investigation discovered that tDNAs have no effect on albumin proteins, either by interfering or by adhering to the extracellular proteins. These findings will aid in the design of future DNA nanostructures for biomedical applications by increasing the knowledge on the biocompatible interaction of tDNAs with biomacromolecules.

P19-derived neuronal cells express H1, H2, and H3 histamine receptors: a biopharmaceutical approach to evaluate antihistamine agents.

Histamine is a biogenic amine implicated in various biological and pathological processes. Convenient cellular models are needed to screen and develop new antihistamine agents. This report aimed to characterize the response of neurons differentiated from mouse P19 embryonal carcinoma cells to histamine treatment, and to investigate the modulation of this response by antihistamine drugs, vegetal diamine oxidase, and catalase. The exposure of P19 neurons to histamine reduced cell viability to 65% maximally. This effect involves specific histamine receptors, since it was prevented by treatment with desloratadine and cimetidine, respectively, H1 and H2 antagonists, but not by the H3 antagonist ciproxifan. RT-PCR analysis showed that P19 neurons express H1 and H2 receptors, and the H3 receptor, although it seemed not involved in the histamine effect on these cells. The H4 receptor was not expressed. H1 and H2 antagonists as well as vegetal diamine oxidase diminished the intracellular Ca2+ mobilization triggered by histamine. The treatment with vegetal diamine oxidase or catalase protected against mortality and a significant reduction of H2O2 level, generated from the cells under the histamine action, was found upon treatments with desloratadine, cimetidine, vegetal diamine oxidase, or catalase. Overall, the results indicate the expression of functional histamine receptors and open the possibility of using P19 neurons as model system to study the roles of histamine and related drugs in neuronal pathogenesis. This model is less expensive to operate and can be easily implemented by current laboratories of analysis and by Contract Research Organizations.

Cellular protection from H2O2 toxicity by Fv-Hsp70: protection via catalase and gamma-glutamyl-cysteine synthase.

Heat shock proteins (HSPs), especially Hsp70 (HSPA1), have been associated with cellular protection from various cellular stresses including heat, hypoxia-ischemia, neurodegeneration, toxins, and trauma. Endogenous HSPs are often synthesized in direct response to these stresses but in many situations are inadequate in protecting cells. The present study addresses the transduction of Hsp70 into cells providing protection from acute oxidative stress by H2O2. The recombinant Fv-Hsp70 protein and two mutant Fv-Hsp70 proteins minus the ATPase domain and minus the ATPase and terminal lid domains were tested at 0.5 and 1.0 µM concentrations after two different concentrations of H2O2 treatment. All three recombinant proteins protected SH-SY5Y cells from acute H2O2 toxicity. This data indicated that the protein binding domain was responsible for cellular protection. In addition, experiments pretreating cells with inhibitors of antioxidant proteins catalase and gamma-glutamylcysteine synthase (GGCS) before H2O2 resulted in cell death despite treatment with Fv-Hsp70, implying that both enzymes were protected from acute oxidative stress after treatment with Fv-Hsp70. This study demonstrates that Fv-Hsp70 is protective in our experiments primarily by the protein-binding domain. The Hsp70 terminal lid domain was also not necessary for protection.

Biochemical and phytochemical responses of Ammi visnaga L. (Apiaceae) callus culture elicited by SiO2 and graphene Oxide-SiO2 nanoparticles.

Ammi visnaga L. is an enriched medicinal plant with medicinally important compounds. Two types of nanoparticles (NPs) including silica (SiO2) and graphene oxide bound with SiO2 (GO-SiO2) NPs at different concentrations (0, 15, 25 mg L-1) were used as elicitors to investigate their effects on callus morphology, H2O2 content, total phenolic content (TPC), total flavonoids content (TFC), ferric reducing/antioxidant power (FRAP), and few antioxidant enzymes such as catalase (CAT), guaiacol peroxidase (GPX), superoxide dismutase (SOD), ascorbate peroxidase (APX), and polyphenol oxidase (PPO) in the callus cultures of A. visnaga. The effects of elicitation of both NPs on calli were observed using a scanning electron microscope (SEM). The 15 mg L-1 concentration of GO-SiO2 NPs produced the highest TPC (193.3 mg GAE g-1 FW), CAT (13.1 U mg-1 Protein), GPX (0.0089 U mg-1 Protein), and APX (0.079 U mg-1 Protein). Whereas, the maximum content of H2O2 (0.68 µmol g-1 FW), FRAP (0.0092 µmol mg-1), and TFC (62.27 mg QE g-1 FW) was observed at 25 mg L-1 and 15 mg L-1 of SiO2 NPs, respectively. Conclusively, in the callus culture of A. visnaga, the 15 mg L-1 concentration of GO-SiO2 NPs was the most suitable dosage for enhancing the enzymatic antioxidant activities (CAT, GPX, APX) and TPC, rather than SiO2 NPs.

A Direct Catalytic Ethanol Fuel Cell (DCEFC) Modified by LDHs, or by Catalase-LDHs, and Improvement in Its Kinetic Performance: Applications for Human Saliva and Disinfectant Products for COVID-19.

In this work, it has been experimentally proven that the kinetic performance of a common Direct Catalytic Ethanol Fuel Cell (DCEFC) can be increased by introducing nanostructured (ZnII,AlIII(OH)2)+NO3-·H2O Layered Double Hydroxides (LDHs) into the anode compartment. Carrying out the measurements with the open-circuit voltage method and using a kinetic format, it has been shown that the introduction of LDHs in the anodic compartment implies a 1.3-fold increase in the calibration sensitivity of the method. This improvement becomes even greater in the presence of hydrogen peroxide in a solution. Furthermore, we show that the calibration sensitivity increased by 8-times, when the fuel cell is modified by the enzyme catalase, crosslinked on LDHs and in the presence of hydrogen peroxide. The fuel cell, thus modified (with or without enzyme), has been used for analytical applications on real samples, such as biological (human saliva) and hand disinfectant samples, commonly used for the prevention of COVID-19, obtaining very positive results from both analytical and kinetic points of view on ethanol detection. Moreover, if the increase in the calibration sensitivity is of great importance from the point of view of analytical applications, it must be remarked that the increase in the speed of the ethanol oxidation process in the fuel cell can also be extremely useful for the purposes of improving the energy performance of a DCEFC.

Nanozymes towards Personalized Diagnostics: A Recent Progress in Biosensing.

This review highlights the recent advancements in the field of nanozymes and their applications in the development of point-of-care biosensors. The use of nanozymes as enzyme-mimicking components in biosensing systems has led to improved performance and miniaturization of these sensors. The unique properties of nanozymes, such as high stability, robustness, and surface tunability, make them an attractive alternative to traditional enzymes in biosensing applications. Researchers have explored a wide range of nanomaterials, including metals, metal oxides, and metal-organic frameworks, for the development of nanozyme-based biosensors. Different sensing strategies, such as colorimetric, fluorescent, electrochemical and SERS, have been implemented using nanozymes as signal-producing components. Despite the numerous advantages, there are also challenges associated with nanozyme-based biosensors, including stability and specificity, which need to be addressed for their wider applications. The future of nanozyme-based biosensors looks promising, with the potential to bring a paradigm shift in biomolecular sensing. The development of highly specific, multi-enzyme mimicking nanozymes could lead to the creation of highly sensitive and low-biofouling biosensors. Integration of nanozymes into point-of-care diagnostics promises to revolutionize healthcare by improving patient outcomes and reducing costs while enhancing the accuracy and sensitivity of diagnostic tools.

Different Directions of Effects of Polyclonal IgG Antibodies from Patients with Schizophrenia and Healthy Individuals on Cell Death In Vitro: A Pilot Study.

Numerous studies indicate the involvemen of oxidative stress in the pathogenesis of schizophrenia. It has been shown that the serum pool of antibodies in patients with schizophrenia contains catalytically active antibodies (abzymes) that have a wide range of activities, including redox properties. In the present work, the effects of IgGs-having oxidoreductase activities-isolated from the serum of patients with schizophrenia and healthy individuals were studied in vitro. The IgGs were purified by affinity chromatography followed by an SDS-PAGE analysis of homogeneity in a 4-18% gradient gel. The catalase and superoxide dismutase (SOD) activities of the IgGs were measured spectrophotometrically using a kinetic module. Human neuroblastoma SH-SY5Y cells were cultured with IgG at a final concentration of 0.2 mg/mL for 24 h. In a parallel experiment, tert-butyl hydroperoxide was used as an oxidative stressor. The number of dead cells after incubation was determined with fluorescent dyes, propidium iodide and Hoechst, by high-throughput screening on the CellInsight CX7 platform. A cytotoxic effect of the IgG from the schizophrenia patients on SH-SY5Y cells was detected after 24 h incubation. A correlation was found between the SOD activity of the IgGs and IgG-induced cell death. Under the induced oxidative stress, the cytotoxic effect of the IgG from the patients with schizophrenia on the SH-SY5Y cell line was five times stronger. Meanwhile, the IgG from the healthy individuals exerted a cytoprotective effect on the cultured cells, accompanied by high catalase activity. Thus, the observed influence on cell viability depends on the catalytic properties of the abzymes.

Peroxisomes during postnatal development of mouse endocrine and exocrine pancreas display cell-type- and stage-specific protein composition.

Peroxisomal dysfunction unhinges cellular metabolism by causing the accumulation of toxic metabolic intermediates (e.g. reactive oxygen species, very -chain fatty acids, phytanic acid or eicosanoids) and the depletion of important lipid products (e.g. plasmalogens, polyunsaturated fatty acids), leading to various proinflammatory and devastating pathophysiological conditions like metabolic syndrome and age-related diseases including diabetes. Because the peroxisomal antioxidative marker enzyme catalase is low abundant in Langerhans islet cells, peroxisomes were considered scarcely present in the endocrine pancreas. Recently, studies demonstrated that the peroxisomal metabolism is relevant for pancreatic cell functionality. During the postnatal period, significant changes occur in the cell structure and the metabolism to trigger the final maturation of the pancreas, including cell proliferation, regulation of energy metabolism, and activation of signalling pathways. Our aim in this study was to (i) morphometrically analyse the density of peroxisomes in mouse endocrine versus exocrine pancreas and (ii) investigate how the distribution and the abundance of peroxisomal proteins involved in biogenesis, antioxidative defence and fatty acid metabolism change during pancreatic maturation in the postnatal period. Our results prove that endocrine and exocrine pancreatic cells contain high amounts of peroxisomes with heterogeneous protein content indicating that distinct endocrine and exocrine cell types require a specific set of peroxisomal proteins depending on their individual physiological functions. We further show that significant postnatal changes occur in the peroxisomal compartment of different pancreatic cells that are most probably relevant for the metabolic maturation and differentiation of the pancreas during the development from birth to adulthood.

Role of Metabolism on Alcohol Preference, Addiction, and Treatment.

Studies presented in this chapter show that: (1) in the brain, ethanol is metabolized by catalase to acetaldehyde, which condenses with dopamine forming salsolinol; (2) acetaldehyde-derived salsolinol increases the release of dopamine mediating, via opioid receptors, the reinforcing effects of ethanol during the acquisition of ethanol consumption, while (3) brain acetaldehyde does not influence the maintenance of chronic ethanol intake, it is suggested that a learned cue-induced hyperglutamatergic system takes precedence over the dopaminergic system. However, (4) following a prolonged ethanol deprivation, the generation of acetaldehyde in the brain again plays a role, contributing to the increase in ethanol intake observed during ethanol re-access, called the alcohol deprivation effect (ADE), a model of relapse behavior; (5) naltrexone inhibits the high ethanol intake seen in the ADE condition, suggesting that acetaldehyde-derived salsolinol via opioid receptors also contributes to the relapse-like drinking behavior. The reader is referred to glutamate-mediated mechanisms that trigger the cue-associated alcohol-seeking and that also contribute to triggering relapse.

Measures of Oxidative Status Markers in Relation to Age, Sex, and Season in Sick and Healthy Captive Asian Elephants in Thailand.

Oxidative stress is a pathological condition that can have adverse effects on animal health, although little research has been conducted on wildlife species. In this study, blood was collected from captive Asian elephants for the assessment of five serum oxidative status markers (reactive oxygen species (ROS) concentrations; malondialdehyde, MDA; albumin; glutathione peroxidase, GPx; and catalase) in healthy (n = 137) and sick (n = 20) animals. Health problems consisted of weakness, puncture wounds, gastrointestinal distress, eye and musculoskeletal problems, and elephant endotheliotropic herpesvirus hemorrhagic disease (EEHV-HD). Fecal samples were also collected to assess glucocorticoid metabolites (fGCMs) as a measure of stress. All data were analyzed in relation to age, sex, sampling season, and their interactions using generalized linear models, and a correlation matrix was constructed. ROS and serum albumin concentrations exhibited the highest concentrations in aged elephants (>45 years). No sex differences were found for any biomarker. Interactions were observed for age groups and seasons for ROS and catalase, while GPx displayed a significant interaction between sex and season. In pairwise comparisons, significant increases in ROS and catalase were observed in summer, with higher ROS concentrations observed only in the adult female group. Lower catalase activity was exhibited in juvenile males, subadult males, adult females, and aged females compared to subadult and adult elephants (males and females) in winter and the rainy season. There was a positive association between catalase activity and fGCMs (r = 0.23, p < 0.05), and a number of red blood cell parameters were positively associated with several of these biomarkers, suggesting high oxidative and antioxidative activity covary in red cells (p < 0.05). According to health status, elephants with EEHV-HD showed the most significant changes in oxidative stress markers, with MDA, GPx, and catalase being higher and albumin being lower than in healthy elephants. This study provides an analysis of understudied health biomarkers in Asian elephants, which can be used as additional tools for assessing the health condition of this species and suggests age and season may be important factors in data interpretation.

A catalase inhibitor: Targeting the NADPH-binding site for castration-resistant prostate cancer therapy.

Catalase (CAT) is an important antioxidant enzyme that breaks down H2O2 into water and oxygen. Inhibitor-modulating CAT activity in cancer cells is emerging as a potential anticancer strategy. However, the discovery of CAT inhibitors towards the heme active center located at the bottom of long and narrow channel has made little progress. Therefore, targeting new binding site is of great importance for the development of efficient CAT inhibitors. Here, the first NADPH-binding site inhibitor of CAT, BT-Br, was designed and synthesized successfully. The cocrystal structure of BT-Br-bound CAT complex was determined with a resolution of 2.2 Å (PDB ID:8HID), which showed clearly that BT-Br bound at the NADPH-binding site. Furthermore, BT-Br was demonstrated to induce ferroptosis in castration-resistant prostate cancer (CRPC) DU145 cells and eventually reduce CRPC tumors in vivo effectively. The work indicates that CAT has potential as a novel target for CRPC therapy based on ferroptosis inducing.

Simple assay for quantifying xanthine oxidase activity.

This paper reports a sensitive method for assaying xanthine oxidase (XO) enzyme activity. XO produces hydrogen peroxide (H2O2) and superoxide anion radicals (O2•-), promoting the development of oxidative stress-related diseases, and is inhibited by various plant extracts. XO activity is quantified by incubating enzyme samples with an appropriate xanthine concentration as the substrate. The proposed method requires XO activity to be quantified based on H2O2 generation using a 3,3',5,5'-tetramethylbenzidine (TMB)-H2O2 system catalyzed by cupric ions. After a 30-min incubation at 37 °C, sufficient cupric ion and TMB amounts are added. The assay produces optical signals that can be visually recognized or detected with a UV-visible spectrometer. A direct correlation was found between XO activity and the absorbance at 450 nm of the resulting di-imine (dication) yellow product. The proposed method uses sodium azide to prevent catalase enzyme interference. The new assay's function was confirmed using the TMB-XO assay and a Bland-Altman plot. The resulting correlation coefficient was 0.9976. The innovative assay was relatively precise and comparable to the comparison protocols. In conclusion, the presented method is very efficient at measuring XO activity.

Alkaline ceramidase (ClAC) inhibition enhances heat stress response in Cyrtorhinus lividipennis (Reuter).

Ceramidases (CDases) are vital sphingolipid enzymes involved in organismal growth and development. They have been reported as key mediators of thermal stress response. However, whether and how CDase responds to heat stress in insects remain unclear. Herein, we identified two CDase genes, C. lividipennis alkaline ceramidase (ClAC) and neutral ceramidase (ClNC), by searching the transcriptome and genome databases of the mirid bug, Cyrtorhinus lividipennis, an important natural predator of planthoppers. Quantitative PCR (qPCR) analysis showed that both ClNC and ClAC were highly expressed in nymphs than in adults. ClAC was especially highly expressed in the head, thorax, and legs, while ClNC was widely expressed in the tested organs. Only the ClAC transcription was significantly affected by heat stress. Knocking down ClAC increased the C. lividipennis nymph survival rate under heat stress. The transcriptome and lipidomics data showed that the RNA interference-mediated suppression of ClAC significantly upregulated the transcription level of catalase (CAT) and the content of long-chain base ceramides, including C16-, C18-, C24-, and C31- ceramides. In C. lividipennis nymphs, ClAC played an important role in heat stress response, and the upregulation of nymph survival rate might be caused by variation in the ceramide levels and transcriptional changes in CDase downstream genes. This study improves our understanding of the physiological functions of insect CDase under heat stress and provides valuable insights into the nature enemy application.

Comparing the Role of ROS and RNS in the Thermal Stress Response of Two Cnidarian Models, Exaiptasia diaphana and Galaxea fascicularis.

Coral reefs are threatened by climate change, because it causes increasingly frequent and severe summer heatwaves, resulting in mass coral bleaching and mortality. Coral bleaching is believed to be driven by an excess production of reactive oxygen (ROS) and nitrogen species (RNS), yet their relative roles during thermal stress remain understudied. Here, we measured ROS and RNS net production, as well as activities of key enzymes involved in ROS scavenging (superoxide dismutase and catalase) and RNS synthesis (nitric oxide synthase) and linked these metrics to physiological measurements of cnidarian holobiont health during thermal stress. We did this for both an established cnidarian model, the sea anemone Exaiptasia diaphana, and an emerging scleractinian model, the coral Galaxea fascicularis, both from the Great Barrier Reef (GBR). Increased ROS production was observed during thermal stress in both species, but it was more apparent in G. fascicularis, which also showed higher levels of physiological stress. RNS did not change in thermally stressed G. fascicularis and decreased in E. diaphana. Our findings in combination with variable ROS levels in previous studies on GBR-sourced E. diaphana suggest G. fascicularis is a more suitable model to study the cellular mechanisms of coral bleaching.

Heat Acclimation under Drought Stress Induces Antioxidant Enzyme Activity in the Alpine Plant Primula minima.

Heat and drought stresses are increasingly relevant topics in the context of climate change, particularly in the Alps, which are warming faster than the global average. Previously, we have shown that alpine plants, including Primula minima, can be gradually heat hardened under field conditions in situ to achieve maximum tolerance within a week. Here, we investigated the antioxidant mechanisms of P. minima leaves that had been heat hardened (H) without or with (H+D) additional drought stress. Lower free-radical scavenging and ascorbate concentrations were found in H and H+D leaves, while concentrations of glutathione disulphide (GSSG) were higher under both treatments without any change in glutathione (GSH) and little change in glutathione reductase activity. In contrast, ascorbate peroxidase activity in H leaves was increased, and H+D leaves had >two-fold higher catalase, ascorbate peroxidase and glucose-6-phosphate dehydrogenase activities compared with the control. In addition, the glutathione reductase activity was higher in H+D compared with H leaves. Our results highlight that the stress load from heat acclimation to maximum tolerance is associated with a weakened low-molecular-weight antioxidant defence, which may be compensated for by an increased activity of antioxidant enzymes, particularly under drought conditions.

Effects of Deacetylase Inhibition on the Activation of the Antioxidant Response and Aerobic Metabolism in Cellular Models of Fanconi Anemia.

Fanconi anemia (FA) is a rare genetic disease characterized by a dysfunctional DNA repair and an oxidative stress accumulation due to defective mitochondrial energy metabolism, not counteracted by endogenous antioxidant defenses, which appear down-expressed compared to the control. Since the antioxidant response lack could depend on the hypoacetylation of genes coding for detoxifying enzymes, we treated lymphoblasts and fibroblasts mutated for the FANC-A gene with some histone deacetylase inhibitors (HDACi), namely, valproic acid (VPA), beta-hydroxybutyrate (OHB), and EX527 (a Sirt1 inhibitor), under basal conditions and after hydrogen peroxide addition. The results show that VPA increased catalase and glutathione reductase expression and activity, corrected the metabolic defect, lowered lipid peroxidation, restored the mitochondrial fusion and fission balance, and improved mitomycin survival. In contrast, OHB, despite a slight increase in antioxidant enzyme expressions, exacerbated the metabolic defect, increasing oxidative stress production, probably because it also acts as an oxidative phosphorylation metabolite, while EX527 showed no effect. In conclusion, the data suggest that VPA could be a promising drug to modulate the gene expression in FA cells, confirming that the antioxidant response modulation plays a pivotal in FA pathogenesis as it acts on both oxidative stress levels and the mitochondrial metabolism and dynamics quality.

Overexpression of Mitochondrial Catalase within Adipose Tissue Does Not Confer Systemic Metabolic Protection against Diet-Induced Obesity.

Obesity is associated with significant metabolic co-morbidities, such as diabetes, hypertension, and dyslipidaemia, as well as a range of cardiovascular diseases, all of which lead to increased hospitalisations, morbidity, and mortality. Adipose tissue dysfunction caused by chronic nutrient stress can result in oxidative stress, mitochondrial dysfunction, inflammation, hypoxia, and insulin resistance. Thus, we hypothesised that reducing adipose tissue oxidative stress via adipose tissue-targeted overexpression of the antioxidant mitochondrial catalase (mCAT) may improve systemic metabolic function. We crossed mCAT (floxed) and Adipoq-Cre mice to generate mice overexpressing catalase with a mitochondrial targeting sequence predominantly in adipose tissue, designated AdipoQ-mCAT. Under normal diet conditions, the AdipoQ-mCAT transgenic mice demonstrated increased weight gain, adipocyte remodelling, and metabolic dysfunction compared to the wild-type mice. Under obesogenic dietary conditions (16 weeks of high fat/high sucrose feeding), the AdipoQ-mCAT mice did not result in incremental impairment of adipose structure and function but in fact, were protected from further metabolic impairment compared to the obese wild-type mice. While AdipoQ-mCAT overexpression was unable to improve systemic metabolic function per se, our results highlight the critical role of physiological H2O2 signalling in metabolism and adipose tissue function.