Manganese-dependent Superoxide Dismutase Research Articles

Exploratory Studies on Chrysin via Antioxidant, Antimicrobial, ADMET, PASS and Molecular Docking evaluations

IntroductionChrysin, a flavonoid found in Feng Jiao (Chinese name for propolis), exhibits a wide array of pharmacological activities and is recognized for the significant roles it plays in traditional Chinese medicine. The present study aimed to explore the in vitro antioxidant and antimicrobial potentials of chrysin and provide mechanistic insights via in silico experiments. MethodsThe in vitro antioxidant and antimicrobial activities of chrysin, a natural flavone found in propolis was investigated. Molecular docking interactions, the Adsorption, Distribution, Metabolism, Elimination and Toxicity (ADMET) and Prediction of Activity Spectra for Substances (PASS) were used to predict the biological activities of chrysin. ResultsChrysin exhibited a considerable 2,2-diphenyl-1-picrylhydrazyl (DPPH) radicals scavenging activity and ferric reducing antioxidant capacity. Chrysin demonstrated inhibition against S. typhi, and B. subtilis with lowest MIC value of 1.25 mg/mL which was similar to that of chloramphenicol. Drug-likeness and ADMET study of the two compounds showed their physicochemical properties as non-toxic, non-mutagenic, high permeability, good solubility and absorption characteristics. DiscussionThe research below shows that chrysin and silymarin had antioxidant, free radical scavenging, antibacterial, hepatoprotective and anticarcinogenic properties based on PASS prediction of biological activities. Molecular docking of chrysin and silymarin, a standard hepatoprotective drug, with tumour necrosis factor alpha converting enzyme (TACE) and manganese-dependent superoxide dismutase (SOD 2) showed interactions with LEU 401, VAL 402, HIS 405 amino acid residues at the active sites of TACE. Chrysin non-covalently interacted with amino acids of SOD 2 at the active site while silymarin non-covalently interacted at a site different from the active site. Silymarin demonstrated a minimum binding energy of – 9.7 Kcal/mol and – 7.0 Kcal/mol for TACE and SOD 2, respectively while chrysin showed a minimal binding energy of – 9.0 Kcal/mol and – 6.8 Kcal/mol for TACE and SOD2 respectively. ConclusionThe study concluded that chrysin exhibited antioxidant and antimicrobial activity via inhibition and regulation of TACE and SOD2 and possibly offer protection against hepatotoxicity in animal experimental models.

Mitochondrial respiration analysis in permeabilized porcine left ventricular and human right atrial specimens with ischemia-reperfusion.

Mitochondrial function is critical to myocardial ischemia-reperfusion injury and cardioprotection. The measurement of mitochondrial function in isolated mitochondria requires cardiac specimens of about 300 mg and is therefore only possible at the end of an animal experiment or during cardiosurgical interventions in humans. As an alternative, mitochondrial function can be measured in permeabilized myocardial tissue (PMT) specimens of about 2-5 mg, which are retrieved by sequential biopsies in animal experiments and during cardiac catheterization in humans. We attempted to validate measurements of mitochondrial respiration from PMT by comparison with those from isolated mitochondria of left ventricular myocardium from anesthetized pigs undergoing 60 min coronary occlusion and 180 min reperfusion. Mitochondrial respiration was normalized to the content of mitochondrial marker proteins [cytochrome-c oxidase 4 (COX4), citrate synthase, and manganese-dependent superoxide dismutase]. When normalized to COX4, mitochondrial respiration measurements in PMT and isolated mitochondria agreed well in Bland-Altman plots (bias score, -0.03 nmol/min/COX4; 95% confidence interval: 6.31 nmol/min/COX4 and -6.37 nmol/min/COX4) and correlated well (slope of 0.77 and Pearson's R of 0.87). Mitochondrial dysfunction by ischemia-reperfusion was equally reflected in PMT and isolated mitochondria (44 and 48% reduction of ADP-stimulated complex I respiration). Also in isolated human right atrial trabeculae, simulation of ischemia-reperfusion injury by exposure to 60 min hypoxia and 10 min reoxygenation reduced mitochondrial ADP-stimulated complex I respiration by 37% in PMT. In conclusion, mitochondrial function measurements in permeabilized cardiac tissue can substitute for that in isolated mitochondria to reflect mitochondrial dysfunction following ischemia-reperfusion.NEW & NOTEWORTHY Methods to quantify mitochondrial function in translationally relevant models and in human tissue are needed to improve the translation of cardioprotection to patients. Our present approach, using PMT instead of isolated mitochondria for the quantification of mitochondrial ischemia-reperfusion damage, provides a reference for further studies in translationally relevant large animal models and in human tissue, thus possibly improving the translation of cardioprotection to the benefit of patients with acute myocardial infarction.

Inverse Salt Sensitivity of Blood Pressure Is Associated with an Increased Renin-Angiotensin System Activity.

High and low sodium diets are associated with increased blood pressure and cardiovascular morbidity and mortality. The paradoxical response of elevated BP in low salt diets, aka inverse salt sensitivity (ISS), is an understudied vulnerable 11% of the adult population with yet undiscovered etiology. A linear relationship between the number of single nucleotide polymorphisms (SNPs) in the dopamine D2 receptor (DRD2, rs6276 and 6277), and the sodium myo-inositol cotransporter 2 (SLC5A11, rs11074656), as well as decreased expression of these two genes in urine-derived renal proximal tubule cells (uRPTCs) isolated from clinical study participants suggest involvement of these cells in ISS. Insight into this newly discovered paradoxical response to sodium is found by incubating cells in low sodium (LS) conditions that unveil cell physiologic differences that are then reversed by mir-485-5p miRNA blocker transfection and bypassing the genetic defect by DRD2 re-expression. The renin-angiotensin system (RAS) is an important counter-regulatory mechanism to prevent hyponatremia under LS conditions. Oversensitive RAS under LS conditions could partially explain the increased mortality in ISS. Angiotensin-II (AngII, 10 nmol/L) increased sodium transport in uRPTCs to a greater extent in individuals with ISS than SR. Downstream signaling of AngII is verified by identifying lowered expression of nuclear factor erythroid 2-related factor 2 (NRF2), CCCTC-binding factor (CTCF), and manganese-dependent mitochondrial superoxide dismutase (SOD2) only in ISS-derived uRPTCs and not SR-derived uRPTCs when incubated in LS conditions. We conclude that DRD2 and SLC5A11 variants in ISS may cause an increased low sodium sensitivity to AngII and renal sodium reabsorption which can contribute to inverse salt-sensitive hypertension.

Evidence for novel cell defense mechanisms sustained by dimethyl fumarate in multiple sclerosis patients: the HuR/SOD2 cascade

BackgroundDimethyl fumarate (DMF) is an effective treatment for relapsing remitting Multiple Sclerosis (MS) and its mechanisms of action encompass immunomodulatory and cytoprotective effects. Despite DMF is known to activate the Nrf2 pathway, Nrf2-independent mechanisms have been also reported and new insights on the underlying molecular mechanisms are still emerging including transcriptional and post-transcriptional events.At this regard, we focused on a small family of RNA-binding proteins, the ELAV-like proteins, that play a pivotal role in post-transcriptional mechanisms and are involved in the pathogenesis of several psychiatric and neurologic disorders. HuR, the ubiquitously expressed member of the family, is implicated in many cellular functions, including survival, inflammation and proper functioning of the immune system. We previously documented the potential entanglement of HuR in MS pathogenesis. In the present work, we explored HuR protein levels in peripheral blood mononuclear cells (PBMCs) from MS patients before and after DMF treatment compared to healthy controls (HC). Considering that HuR may act on various targets, playing a protective role against oxidative stress, our main goals were to evaluate whether manganese-dependent superoxide dismutase transcript (SOD2) could represent a new molecular target of HuR and to study the potential influence of DMF treatment on this interaction. MethodsPBMCs from 20 patients with MS and 20 frequency-matched HC by sex and age were used to evaluate HuR, MnSOD (the protein coded by SOD2) and Nrf2 protein content by Western blot, before and after 12 months of DMF treatment.Immunoprecipitation experiments coupled with RNA extraction in PBMCs were performed to explore whether SOD2 mRNA could be physically bound by HuR and whether the expression of MnSOD protein could be affected by 12 months of DMF treatment. ResultsIn PBMCs, HuR protein binds SOD2 transcript in HC and in MS patients naïve to disease modifying treatment. The expression of MnSOD protein is positively affected by 12 months of DMF treatment. PBMCs from MS patients have a lower HuR and MnSOD protein content compared to matched HC (HuR: p<0.01, MnSOD: p<0.01). Of interest, 12 months of DMF treatment in MS patients restores the amount of both HuR protein and MnSOD enzyme to the levels observed in HC. We also confirmed that Nrf2 is an HuR target, and we report that its levels are significantly increased in MS patients naïve to disease modifying treatment and remain elevated following DMF administration. ConclusionSOD2 transcript is a new target of HuR protein. DMF induces an increased expression of HuR protein, which ultimately interacts more strongly with SOD2 transcript promoting the expression of this antioxidant protein. The activation of this molecular cascade can constitute an additional tool that the cells can exploit to counteract the oxidative stress associated with MS development, and can account for the multifaceted molecular mechanisms underlying DMF efficacy in MS.

Antioxidant Effects of Pyrazoles in Ischemia/Reperfusion Injury in Cardiomyocytes

Since the turn of the 21st century, cardiovascular diseases (CVDs) have remained the leading cause of death worldwide. In the United States alone, approximately 1.5 million people annually will have a myocardial infarction (MI), or more commonly known as a heart attack. The most common cause of MI is coronary artery disease (CAD), also known as atherosclerosis, where a significant occlusion in the coronary artery leads to an acute mismatch of oxygen supply and demand in the myocardium, leading to myocardial ischemia. Removing the occlusion is critical to salvaging ischemic tissues; however, reperfusion and restoring blood flow to previously anoxic/hypoxic tissues may cause a paradoxical worsening of cardiac cell dysfunction and death. This phenomenon is known as ischemia/reperfusion injury (IRI). Multiple studies have shown that oxidative stress plays a crucial role in the pathophysiology of IRI, and therapeutic studies are now being conducted to find an agent that can counteract the sudden rise in reactive oxygen species (ROS). Pyrazoles are small, heterocyclic molecules found in many biologically active compounds and have been identified as potential therapeutic agents against CVD due to their anti‐inflammatory effects. Most importantly, they are relatively easy to synthesize and can be fine‐tuned to achieve desired electronic and steric effects. Thus, pyrazoles are gaining more attention in the field of drug discovery research. In our present study, the function of pyrazole KT‐01‐14B as an antioxidant was determined by evaluating its effect on the expression of biomarkers indicative of ischemia/reperfusion (I/R)‐induced oxidative stress, which includes manganese‐dependent superoxide dismutase (SOD2), acetylated tubulin, and 4‐hydroxynonenal (4‐HNE) aggresomes. I/R was induced in vivo by using mice models and in vitro by subjecting HL‐1 cardiomyocytes to hypoxia/reoxygenation (H/R). Histologic evaluation of the cardiac tissue in our in vivo models was accomplished using TTC staining, which revealed that pyrazoles significantly reduce infarction size. To measure the mitochondrial superoxide levels in the HL‐1 cardiomyocytes, MitoSOX staining was used, which indicated that superoxide levels were significantly reduced with pyrazole treatment. Further investigation of pyrazole‐treated HL‐1 cardiomyocytes with immunoblotting analysis and immunocytochemistry revealed increased expression of SOD2, increased expression of acetylated tubulin, and decreased 4‐HNE aggresome formation. In addition to studying the effects of pyrazole on the biomarkers, a cell viability assay was performed to determine the effects of pyrazole on cardiomyocyte cell death. These results conclude that the antioxidant function of pyrazoles can prevent cellular damage caused by I/R.

Perillaldehyde Alleviates Spinal Cord Ischemia-Reperfusion Injury Via Activating the Nrf2 Pathway

Spinal Cord ischemia-reperfusion injury (SCII) is one of the most destructive complications in thoracic-abdominal aortic surgery, which can cause physical abnormalities, paralysis and even brain death. Evidence has shown that perillaldehyde (PAH) can ameliorate rat's cerebra ischemia-reperfusion injury. However, the effect of PAH on SCII remains unknown. The current study established SCII rat models and oxygen and glucose deprivation/reoxygenation-induced BV2 microglia models to explore whether PAH could alleviate SCII symptoms and to investigate underlying mechanism. SCII rats underwent severe neurologic motor dysfunction and histopathologic injury compared with the normal rats, which are exhibited by loss of motor neurons and decrease of nissl bodies. Treatment with PAH significantly ameliorated motor dysfunction and neuron damage. PAH downregulated the expression of NLR family pyrin domain containing 3, cleaved/pro caspase-1, interleukin-1β and interleukin-18 in spinal cord tissues of SCII rats. Besides, the contents of oxidative stress-related factors superoxide dismutase, manganese-dependent superoxide dismutase, catalase and glutathione peroxidase were significantly increased and malondialdehyde content was decreased after PAH treatment. PAH treatment upregulated the expression of nuclear factor-E2-related factor 2 and heme oxygenase-1 in spinal cord tissues of SCII rats. Our in vitro study confirmed that PAH inhibited microglial activation by activating the nuclear factor-E2-related factor 2/heme oxygenase-1 pathway, exhibited by alleviated inflammation and oxidative stress. This study elucidates that PAH has the potential value for treating SCII, which provides an experimental basis for clinical trials in the future.

Anakinra Activates Superoxide Dismutase 2 to Mitigate Inflammasome Activity.

Inflammasomes are powerful cytosolic sensors of environmental stressors and are critical for triggering interleukin-1 (IL-1)-mediated inflammatory responses. However, dysregulation of inflammasome activation may lead to pathological conditions, and the identification of negative regulators for therapeutic purposes is increasingly being recognized. Anakinra, the recombinant form of the IL-1 receptor antagonist, proved effective by preventing the binding of IL-1 to its receptor, IL-1R1, thus restoring autophagy and dampening NLR family pyrin domain containing 3 (NLRP3) activity. As the generation of mitochondrial reactive oxidative species (ROS) is a critical upstream event in the activation of NLRP3, we investigated whether anakinra would regulate mitochondrial ROS production. By profiling the activation of transcription factors induced in murine alveolar macrophages, we found a mitochondrial antioxidative pathway induced by anakinra involving the manganese-dependent superoxide dismutase (MnSOD) or SOD2. Molecularly, anakinra promotes the binding of SOD2 with the deubiquitinase Ubiquitin Specific Peptidase 36 (USP36) and Constitutive photomorphogenesis 9 (COP9) signalosome, thus increasing SOD2 protein longevity. Functionally, anakinra and SOD2 protects mice from pulmonary oxidative inflammation and infection. On a preclinical level, anakinra upregulates SOD2 in murine models of chronic granulomatous disease (CGD) and cystic fibrosis (CF). These data suggest that protection from mitochondrial oxidative stress may represent an additional mechanism underlying the clinical benefit of anakinra and identifies SOD2 as a potential therapeutic target.

Salidroside protects cardiac function in mice with diabetic cardiomyopathy via activation of mitochondrial biogenesis and SIRT3.

To investigate the effects and the underlying mechanisms of salidroside on diabetic cardiomyopathy, diabetes was induced in mice by a long-term high-fat diet and a low-dose injection of streptozocin. Measurements of cardiac function, biochemical analysis, and histopathological examinations were conducted to evaluate the therapeutic effects of salidroside. In this study, we found that diabetic mice exhibited decreased cardiac systolic function and impaired mitochondrial ultrastructure. Pre-treatment with salidroside protected mice against myocardial dysfunction, reduced blood glucose, improved insulin resistance, and induced mitochondrial biogenesis. Neonatal rat cardiomyocytes were cultured to explore the mechanisms of salidroside in vitro. Salidroside alleviated decreased expression of peroxisome proliferator-activated receptor-γ coactivator 1-alpha (PGC-1α), mitochondrial transcription factor A (TFAM) via phosphorylation of 5' AMP-activated protein kinase (AMPK), which may be associated with mitochondrial biogenesis. Salidroside also increased sirtuin-3 (SIRT3) expression in cardiomyocytes. Furthermore, salidroside promoted the translocation of SIRT3 from cytoplasm to mitochondria and increased the deacetylation of mitochondrial proteins such as manganese-dependent superoxide dismutase (MnSOD). In Conclusion, salidroside not only improved diabetes, but also ameliorated diabetic cardiomyopathy, which was at least partly associated with the activation of mitochondrial SIRT3, AMPK/Akt, and PGC-1α/TFAM and subsequent improving mitochondrial function.

Association of Val16Ala Polymorphism of Manganese Superoxide Dismutase (MnSOD) with Food Intake and Cardiometabolic Risk Factors in the Elderly in Primary Care in Porto Alegre.

The aging process causes physiological changes on its own. The combination of an unhealthy lifestyle with the presence of genetic polymorphisms, such as the Val16Ala of the antioxidant enzyme manganese-dependent superoxide dismutase (MnSOD) may contribute to a greater occurrence of cardiometabolic risk factors. This study aimed to verify the association of Val16Ala-MnSOD polymorphism with food intake, caloric expenditure, and cardiometabolic risk factors in the elderly. A cross-sectional study with a sample size of 270 elderly individuals assisted in primary health care in the city of Porto Alegre, RS, Brazil. Val16Ala polymorphism, glucose, lipid profile, insulin, HOMA-IR, blood pressure, waist circumference, PCR-us, IL-6, food consumption, and caloric expenditure were evaluated. The average age of the elderly was 68.6 ± 7.6 years. There were statistically significant differences regarding the consumption of two or more servings of fruits and vegetables daily between the elderly VV versus AV (P=0.017). There were also statistically significant differences regarding the consumption of two or more daily servings of legumes and eggs between the elderly AA versus VV (P=0.002). The median of insulin was higher in the elderly AA versus AV (P=0.025) and the median of HOMA-IR was higher in the elderly VV versus AV (P=0.029). AA elderly individuals had higher means of high-density lipoprotein (HDL-c), compared to AV (P=0.029). The results suggest that Val16Ala -MnSOD polymorphism is associated with the consumption of fruits, vegetables, legumes, and eggs, as well as with cardiometabolic risk factors in the elderly.

Placental Mitochondrial Abnormalities in Preeclampsia

Preeclampsia complicates 5–8% of all pregnancies worldwide, and although its pathophysiology remains obscure, placental oxidative stress and mitochondrial abnormalities are considered to play a key role. Mitochondrial abnormalities in preeclamptic placentae have been described, but the extent to which mitochondrial content and the molecular pathways controlling this (mitochondrial biogenesis and mitophagy) are affected in preeclamptic placentae is unknown. Therefore, in preeclamptic (n = 12) and control (n = 11) placentae, we comprehensively assessed multiple indices of placental antioxidant status, mitochondrial content, mitochondrial biogenesis, mitophagy, and mitochondrial fusion and fission. In addition, we also explored gene expression profiles related to inflammation and apoptosis. Preeclamptic placentae were characterized by higher levels of oxidized glutathione, a higher total antioxidant capacity, and higher mRNA levels of the mitochondrial-located antioxidant enzyme manganese-dependent superoxide dismutase 2 compared to controls. Furthermore, mitochondrial content was significantly lower in preeclamptic placentae, which was accompanied by an increased abundance of key constituents of glycolysis. Moreover, mRNA and protein levels of key molecules involved in the regulation of mitochondrial biogenesis were lower in preeclamptic placentae, while the abundance of constituents of the mitophagy, autophagy, and mitochondrial fission machinery was higher compared to controls. In addition, we found evidence for activation of apoptosis and inflammation in preeclamptic placentae. This study is the first to comprehensively demonstrate abnormalities at the level of the mitochondrion and the molecular pathways controlling mitochondrial content/function in preeclamptic placentae. These aberrations may well contribute to the pathophysiology of preeclampsia by upregulating placental inflammation, oxidative stress, and apoptosis.Graphical

Mitochondrial Sirtuins and Doxorubicin-induced Cardiotoxicity.

Doxorubicin (DOX) is the most effective and extensively used treatment for many tumors. However, its clinical use is hampered by its cardiotoxicity. DOX-induced mitochondrial dysfunction, which causes reactive oxygen species (ROS) generation, cardiomyocyte death, bioenergetic failure, and decreased cardiac function, is a very important mechanism of cardiotoxicity. These cellular processes are all linked by mitochondrial sirtuins (SIRT3-SIRT4). Mitochondrial sirtuins preserve mitochondrial function by increasing mitochondrial metabolism, inhibiting ROS generation by activating the antioxidant enzyme manganese-dependent superoxide dismutase (MnSOD), decreasing apoptosis by activating the forkhead homeobox type O (FOXO) and P53 pathways, and increasing autophagy through AMP-activated protein kinase (AMPK)/mTOR signaling. Thus, sirtuins function at the control point of many mechanisms involved in DOX-induced cardiotoxicity. In this review, we focus on the role of mitochondrial sirtuins in mitochondrial biology and DOX-induced cardiotoxicity. A further aim is to highlight other mitochondrial processes, such as autophagy (mitophagy) and mitochondrial quality control (MQC), for which the effect of mitochondrial sirtuins on cardiotoxicity is unknown.

MnSOD Val16Ala gene polymorphism is associated with REDOX biomarkers in the elderly of primary health care in the city of Porto Alegre

Studies suggest that redox imbalance may be closely associated with pathological aging, contributing effectively to the genesis of several chronic diseases. One of the major defence enzymes against oxidation is Manganese-dependent superoxide dismutase (MnSOD) that acts within the mitochondria. The gene encoding this enzyme is polymorphic and Val16Ala variant is one of its most investigated polymorphisms regarding aging and oxidative stress. This study aimed to verify the occurrence of the MnSOD Val16Ala gene polymorphism association with markers of REDOX metabolism in the elderly of primary health care. A cross-sectional study was performed. The sample consisted of 270 elderly individuals from Family Health Strategy in the city of Porto Alegre, Rio Grande do Sul, Brazil (EMISUS). The following variables were investigated in all subjects: sociodemographic: gender, age, marital status, schooling and income; Anthropometric: weight, height, body mass index (BMI); REDOX markers: advanced oxidation protein products (AOPP), ischemia-modified albumin (IMA), nitric oxide metabolites (NOx), ferric reducing ability of plasma (FRAP) and malondialdehyde (MDA), MnSOD Val16Ala gene polymorphism. Val16Ala gene polymorphism was evaluated by polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP). Statistically significant associations were observed in the elderly with AA genotype compared to those with VV genotype, concerning AOPP (p = 0.023) and FRAP (p = 0.027) quartile frequencies, respectively. No statistically significant differences were observed between MnSOD genotypes with MDA, NOx and IMA oxidative markers. Val16Ala gene polymorphism is associated with AOPP and FRAP quartiles frequencies in the elderly of primary health care.

Differential Activation of Glioprotective Intracellular Signaling Pathways in Primary Optic Nerve Head Astrocytes after Treatment with Different Classes of Antioxidants.

Optic nerve head astrocytes are the specialized glia cells that provide structural and trophic support to the optic nerve head. In response to cellular injury, optic nerve head astrocytes undergo reactive astrocytosis, the process of cellular activation associated with cytoskeletal remodeling, increases in the rate of proliferation and motility, and the generation of Reactive Oxygen Species. Antioxidant intervention has previously been proposed as a therapeutic approach for glaucomatous optic neuropathy, however, little is known regarding the response of optic nerve head astrocytes to antioxidants under physiological versus pathological conditions. The goal of this study was to determine the effects of three different antioxidants, manganese (III) tetrakis (1-methyl-4-pyridyl) porphyrin (Mn-TM-2-PyP), resveratrol and xanthohumol in primary optic nerve head astrocytes. Effects on the expression of the master regulator nuclear factor erythroid 2-related factor 2 (Nrf2), the antioxidant enzyme, manganese-dependent superoxide dismutase 2 (SOD2), and the pro-oxidant enzyme, nicotinamide adenine dinucleotide phosphate oxidase 4 (NOX4), were determined by quantitative immunoblotting. Furthermore, efficacy in preventing chemically and reactive astrocytosis-induced increases in cellular oxidative stress was quantified using cell viability assays. The results were compared to the effects of the prototypic antioxidant, Trolox. Antioxidants elicited highly differential changes in the expression levels of Nrf2, SOD2, and NOX4. Notably, Mn-TM-2-PyP increased SOD2 expression eight-fold, while resveratrol increased Nrf2 expression three-fold. In contrast, xanthohumol exerted no statistically significant changes in expression levels. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) uptake and lactate dehydrogenase (LDH) release assays were performed to assess cell viability after chemically and reactive astrocytosis-induced oxidative stress. Mn-TM-2-PyP exerted the most potent glioprotection by fully preventing the loss of cell viability, whereas resveratrol and xanthohumol partially restored cell viability. Our data provide the first evidence for a well-developed antioxidant defense system in optic nerve head astrocytes, which can be pharmacologically targeted by different classes of antioxidants.

Multiple Bactericidal Mechanisms of the Zinc Ionophore PBT2.

Globally, more antimicrobials are used in food-producing animals than in humans, and the extensive use of medically important human antimicrobials poses a significant public health threat in the face of rising antimicrobial resistance (AMR). The development of novel ionophores, a class of antimicrobials used exclusively in animals, holds promise as a strategy to replace or reduce essential human antimicrobials in veterinary practice. PBT2 is a zinc ionophore with recently demonstrated antibacterial activity against several Gram-positive pathogens, although the underlying mechanism of action is unknown. Here, we investigated the bactericidal mechanism of PBT2 in the bovine mastitis-causing pathogen, Streptococcus uberis In this work, we show that PBT2 functions as a Zn2+/H+ ionophore, exchanging extracellular zinc for intracellular protons in an electroneutral process that leads to cellular zinc accumulation. Zinc accumulation occurs concomitantly with manganese depletion and the production of reactive oxygen species (ROS). PBT2 inhibits the activity of the manganese-dependent superoxide dismutase, SodA, thereby impairing oxidative stress protection. We propose that PBT2-mediated intracellular zinc toxicity in S. uberis leads to lethality through multiple bactericidal mechanisms: the production of toxic ROS and the impairment of manganese-dependent antioxidant functions. Collectively, these data show that PBT2 represents a new class of antibacterial ionophores capable of targeting bacterial metal ion homeostasis and cellular redox balance. We propose that this novel and multitarget mechanism of PBT2 makes the development of cross-resistance to medically important antimicrobials unlikely.IMPORTANCE More antimicrobials are used in food-producing animals than in humans, and the extensive use of medically important human antimicrobials poses a significant public health threat in the face of rising antimicrobial resistance. Therefore, the elimination of antimicrobial crossover between human and veterinary medicine is of great interest. Unfortunately, the development of new antimicrobials is an expensive high-risk process fraught with difficulties. The repurposing of chemical agents provides a solution to this problem, and while many have not been originally developed as antimicrobials, they have been proven safe in clinical trials. PBT2, a zinc ionophore, is an experimental therapeutic that met safety criteria but failed efficacy checkpoints against both Alzheimer's and Huntington's diseases. It was recently found that PBT2 possessed potent antimicrobial activity, although the mechanism of bacterial cell death is unresolved. In this body of work, we show that PBT2 has multiple mechanisms of antimicrobial action, making the development of PBT2 resistance unlikely.

SOD2 genetic polymorphism (rs4880) has no impact on 6‐month response to antidepressant treatment and inflammatory biomarkers in depressed patients

Two thirds of patients suffering from a major depressive episode (MDE) do not reach a complete response with antidepressant drugs. This lack of response is due to several factors, including genetic determinants. Since major depressive disorder is associated with inflammatory and oxidative stress abnormalities, the metabolism of superoxide anions might be involved in non-response to antidepressant drugs. Superoxide anions are metabolized by manganese-dependent superoxide dismutase (SOD2) in the mitochondria. A functional genetic polymorphism (SOD2, rs4880), responsible of a 40% reduction in enzyme activity, is associated with anti-inflammatory response of rosuvastatin. We investigated the association of ala-allele of SOD2 rs4880 and both antidepressant efficacy and inflammatory parameters in patients treated for a MDE with antidepressant drugs. The Hamilton Depression Rating Scale (HDRS) score and levels of plasma CRP and inflammatory cytokines were assessed at baseline, one month (M1), 3months (M3) and 6months (M6) after antidepressant treatment. They were compared according to SOD2 genetic polymorphism. Of the 484 patients studied, 361 (74.6%) carried the ala-allele (Ala group), 123 (25.4%) of them had Val/Val genotype (Val/Val group). No significant difference was observed between the Ala and Val/Val groups neither for baseline clinical characteristics, nor for HDRS scores, response/remission rates, plasma CRP and cytokine levels throughout the study. The rs4880 SOD2 genetic polymorphism was not associated with the clinical response and cytokines levels after antidepressant treatment. These data suggest that SOD2 is not a major genetic determinant of antidepressant response. Other genes of the oxidative stress pathways should be explored in further studies.

Metabolism of Reactive Oxygen Species in Osteosarcoma and Potential Treatment Applications.

Background: The present study was designed to explore the underlying role of hypoxia-inducible factor 1α (HIF-1α) in reactive oxygen species (ROS) formation and apoptosis in osteosarcoma (OS) cells induced by hypoxia. Methods: In OS cells, ROS accumulated and apoptosis increased within 24 h after exposure to low HIF-1α expression levels. A co-expression analysis showed that HIF was positively correlated with Forkhead box class O1 (FoxO1) expression and negatively correlated with CYP-related genes from the National Center for Biotechnology Information’s Gene Expression Omnibus (NCBI GEO) datasets. Hypoxia also considerably increased HIF-1α and FoxO1 expression. Moreover, the promoter region of FoxO1 was directly regulated by HIF-1α. We inhibited HIF-1α via siRNA and found that the ROS accumulation and apoptosis induced by hypoxia in OS cells decreased. In this study, a murine xenograft model of BALB-c nude mice was adopted to test tumour growth and measure the efficacy of 2-ME + As2O3 treatment. Results: Ad interim knockdown of HIF-1α also inhibited manganese-dependent superoxide dismutase (MnSOD), catalase and sestrin 3 (Sesn3) expression in OS cells. Furthermore, hypoxia-induced ROS formation and apoptosis in OS cells were associated with CYP450 protein interference and were ablated by HIF-1α silencing via siRNA. Conclusions: Our data reveal that HIF-1α inhibits ROS accumulation by directly regulating FoxO1 in OS cells, which induces MnSOD, catalase and Sesn3 interference, thus resulting in anti-oxidation effects. The combination of an HIF-1α inhibitor (2-mercaptoethanol,2-ME) and ROS inducer (arsenous oxide, As2O3) can prohibit proliferation and migration and promote apoptosis in MG63 cells in vitro while inhibiting tumour growth in vivo.

Dexmedetomidine alleviates doxorubicin cardiotoxicity by inhibiting mitochondrial reactive oxygen species generation.

Cardiotoxicity largely limits the application of doxorubicin (Dox) for cancer treatment. Dexmedetomidine (Dex), a selective agonist of α2-adrenergic receptor, has been suggested to exert cardioprotection against myocardial injury. However, the effect and underlying mechanisms of Dex on Dox cardiotoxicity remain unknown. In this study, C57BL/6 mice were treated with Dox followed by Dex administration. Cardiomyocytes were co-incubated with Dox and Dex in vitro. The results showed that Dex markedly attenuated cardiac dysfunction induced by Dox. TUNEL staining exhibited that Dex inhibited Dox-induced cardiomyocyte apoptosis in myocardium. Moreover, the expression of anti-apoptotic protein Bcl-2 was increased, whereas the expression of pro-apoptotic protein Bax was decreased by Dex. Dox-induced the increase of reactive oxygen species (ROS), superoxide anion, and mitochondrial ROS (mROS) generation in myocardial tissues were significantly inhibited after Dex administration. In in vitro study, it was further confirmed that Dex prevented Dox-induced cardiomyocyte apoptosis and injury. However, the stimulation of mROS generation reversed the effect of Dex in cardiomyocytes. Mechanically, Dex blocked Dox-induced the ubiquitination of peroxisome proliferator-activated receptor gamma coactivator 1-α (PGC-1α), leading to the restoration of PGC-1α and downstream oxidative stress-protective molecules uncoupling protein 2 and manganese-dependent superoxide dismutase expression. Taken together, this study demonstrates that Dex exerts cardioprotection against Dox cardiotoxicity by attenuating mitochondrial dysfunction, oxidative stress, and cardiomyocyte apoptosis via inhibiting PGC-1α-signaling pathway inactivation. This suggests that Dex may be a potential therapeutic strategy for Dox cardiotoxicity treatment.

Precompetitional Weight Reduction Modifies Prooxidative-Antioxidative Status in Judokas.

Objective The main aim of the study was an assessment of the influence of rapid weight loss on oxidative stress parameters in judokas differing in weight reduction value. Materials and Methods The study included 30 judokas with an age range of 18-30 years (mean age: 22.4 ± 3.40 years). Enzymatic and nonenzymatic antioxidative markers, lipid peroxidation markers, and total oxidative stress were assessed three times: one week before a competition (the first stage), after gaining the desired weight (the second stage), and one week after the competition (the third stage). Results Between the first and the second stage, the concentration of lipid hydroperoxides (LPH) decreased significantly. The superoxide dismutase (SOD), copper- and zinc-containing superoxide dismutase (Cu,Zn-SOD), ceruloplasmin (CER), malondialdehyde (MDA), LPH, and total oxidative stress (TOS) concentrations were the lowest one week after the competition. Linear regression indicated that the emphases on increased weight reduction increased the activity of glutathione peroxidase (GPx), glutathione reductase (GR), glutathione S-transferase (GST), and protein sulfhydryl (PSH) between the first and the second stage of the study. Moderate weight reduction (2-5%) resulted in elevated levels of SOD, Mn-SOD, LPH, MDA, and TOS in comparison to low and high reductions. An opposite relation was observed in PSH. In judokas, the precompetitional weight reduction range was 0.44-6.10% (mean: 2.93% ± 1.76%) of the initial body weight. Concentrations of superoxide dismutase (SOD; p < .01), manganese-dependent superoxide dismutase (Mn-SOD; p < .001), and ceruloplasmin (CER; p < .05) decreased between the first and the third stage of the study as well between the second and third one. Before competitions, a decrease in lipid hydroperoxide (LPH; p < .01) concentration was observed. A reduction of malondialdehyde (MDA; p < .05), LPH (p < .01), and total oxidative stress (TOS; p < .05) levels between the first and the final stage occurred. The increase in weight reduction was linearly correlated with the rise of glutathione peroxidase (GPx; p < .05), glutathione reductase (GR; p < .05), glutathione S-transferase (GST; p < .05), and protein sulfhydryl (PSH; p < .05) concentrations between the first and the second stage of the study. Moderate weight reduction (2-5%) resulted in elevated levels of SOD (p < .05), Mn-SOD (p < .05), LPH (p < .05), MDA (p < .05), and TOS (p < .05) in comparison to low and high reductions. An opposite relation was observed in PSH (p < .005). Conclusions The effect of weight reduction in judo athletes on prooxidative-antioxidative system diversity depends on the weight reduction value.

Protocatechuic Acid-Ameliorated Endothelial Oxidative Stress through Regulating Acetylation Level via CD36/AMPK Pathway.

As one of the main metabolites of anthocyanin, protocatechuic acid (PCA) possesses strong antioxidant activity. In the present study, we explored the capacity of PCA on the alleviation of endothelial oxidative stress and investigated the underlying mechanisms using RNA sequencing (RNA-Seq). In comparison with palmitic acid (PA)-treated cells, PCA (100 μM) significantly decreased the generations of 3-nitrotyrosine (3-NT) and 8-hydroxydeoxyguanosine (8-OHdG) (0.82 ± 0.01 vs 1.16 ± 0.05 and 0.80 ± 0.01 vs 1.48 ± 0.15, respectively, p < 0.01), two biomarkers of oxidative damage, and restored the levels of nitric oxide (NO) (0.97 ± 0.04 vs 0.54 ± 0.02, p < 0.01) and mitochondrial membrane potential (MMP) (0.96 ± 0.03 vs 0.86 ± 0.02, p < 0.01) in human umbilical vein endothelial cells (HUVECs). PCA also obviously reduced the level of reactive oxygen species (ROS) (0.86 ± 0.15 vs 2.67 ± 0.09, p < 0.01) in aorta from high-fat diet (HFD)-fed mice. RNA-Seq and Western blot analysis indicated that PCA markedly reduced the expression of cluster of differentiation 36 (CD36), a membrane fatty acid transporter, and reduced the generations of adenosine triphosphate (ATP) and acetyl coenzyme A (Ac-CoA). These effects of PCA were associated with decreased level of acetylated-lysine and restored the activity of manganese-dependent superoxide dismutase (MnSOD) through reducing the generation of Ac-CoA or activating Sirt1 and Sirt3 via a CD36/AMP-kinase (AMPK) dependent pathway.

Quercetin Enhances the Function and Reduces Apoptosis of Mouse Islets

BackgroundQuercetin (QE) is an antioxidant, anti-inflammatory, flavonoid compound. It was shown that islets are susceptible to oxidative stress due to their inherent low antioxidant capacity. In the present study, we investigated whether treatment of mouse islets with QE could enhance their function before transplantation. MethodsBalb/c mouse islets were treated with various concentrations of QE and their viability, function, and nitric oxide (NO) and the expression of inducible NO synthase (iNOS) were determined before and after cytokine treatment. The expression of antioxidant genes was determined. Apoptosis and apoptosis-associated gene expression was measured using INS-1 cells with or without QE treatment before and after cytokine treatment. ResultsThe QE-treated islets and INS-1 cells showed higher cell function compared to untreated control. The expression of heme oxygenase-1, manganese-dependent superoxide dismutase, and B-cell lymphoma-2 (Bcl-2) were enhanced, and the expression of NO, iNOS, and Bcl-2-associated X protein were reduced before and after cytokine treatment. ConclusionsOur results show that QE could enhance the viability and reduce apoptosis of mouse islets and improve their function before transplantation.