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The mitochondrion, an essential organelle involved in cellular respiration, energy production, and cell death, is the main cellular source of reactive oxygen species (ROS), including superoxide. Mitochondrial diseases resulting from uncontrolled/excess ROS generation are an emerging public health concern and there is current interest in specific mitochondriotropic probes to get information on in-situ ROS production. As such, nitrones vectorized by the triphenylphosphonium (TPP) cation have recently drawn attention despite reported cytotoxicity. Herein, we describe the synthesis of 13 low-toxic derivatives of N-benzylidene-1-diethoxyphosphoryl-1-methylethylamine N-oxide (PPN) alkyl chain-grafted to a pyridinium, triethylammonium or berberinium lipophilic cation. These nitrones showed in-vitro superoxide quenching activity and EPR/spin-trapping efficiency towards biologically relevant free radicals, including superoxide and hydroxyl radicals. Their mitochondrial penetration was confirmed by 31 P NMR spectroscopy, and their anti-apoptotic properties were assessed in Schwann cells treated with hydrogen peroxide. Two pyridinium-substituted PPNs were identified as potentially better alternatives to TPP nitrones conjugates for studying mitochondrial oxidative damage., (© 2023 Wiley-VCH GmbH.)

Oxidative stress is one of the main causes of osteoblast apoptosis induced by post‑menopausal osteoporosis. The authors previously found that metformin can reverse the loss of bone mass in post‑menopausal osteoporosis. The present study aimed to further clarify the effects and mechanisms of action of metformin in post‑menopausal osteoporosis under conditions of oxidative stress. Combined with an in‑depth investigation using the transcriptome database, the association between oxidative stress and mitochondrial dysfunction in post‑menopausal osteoporosis was confirmed. A pre‑osteoblast model of oxidative stress was constructed, and the apoptotic rate following the addition of hydrogen peroxide and metformin was detected using CCK‑8 assay and Annexin V‑FITC/PI staining. Mitochondrial membrane potential was detected using the JC‑1 dye, the intracellular calcium concentration was detected using Fluo‑4 AM, the intracellular reactive oxygen species (ROS) level was observed using DCFH‑DA, and the mitochondrial superoxide level was observed using MitoSOX Red. Bay K8644 was used to increase the level of intracellular calcium. siRNA was used to interfere with the expression of glycogen synthase kinase (GSK)‑3β. Western blot analysis was used to detect the expression of mitochondrial dysfunction‑related proteins. The results revealed that oxidative stress decreased mitochondrial membrane potential and increased intracellular ROS, mitochondrial superoxide and cytoplasmic calcium levels in pre‑osteoblasts; however, metformin improved mitochondrial dysfunction and reversed oxidative stress‑induced injury. Metformin inhibited mitochondrial permeability transition pore opening, suppressed the cytoplasmic calcium influx and reversed pre‑osteoblast apoptosis by promoting GSK‑3β phosphorylation. Moreover, it was found that EGFR was the cell membrane receptor of metformin in pre‑osteoblasts, and the EGFR/GSK‑3β/calcium axis played a key role in metformin reversing the oxidative stress response of pre‑osteoblasts in post‑menopausal osteoporosis. On the whole, these findings provide a pharmacological basis for the use of metformin for the treatment of post‑menopausal osteoporosis.

Background: Recent clinical research suggests that repeated use of opioid pain medications can increase neuropathic pain in people living with human immunodeficiency virus (HIV; PLWH). Therefore, it is significant to elucidate the exact mechanisms of HIV-related chronic pain. HIV infection and chronic morphine induce proinflammatory factors, such as tumor necrosis factor (TNF)α acting through tumor necrosis factor receptor I (TNFRI). HIV coat proteins and/or chronic morphine increase mitochondrial superoxide in the spinal cord dorsal horn (SCDH). Recently, emerging cytoplasmic caspase-11 is defined as a noncanonical inflammasome and can be activated by reactive oxygen species (ROS). Here, we tested our hypothesis that HIV coat glycoprotein gp120 with chronic morphine activates a TNFRI-mtROS-caspase-11 pathway in rats, which increases neuroinflammation and neuropathic pain., Methods: Neuropathic pain was induced by repeated administration of recombinant gp120 with morphine (gp120/M) in rats. Mechanical allodynia was assessed using von Frey filaments, and thermal latency using hotplate test. Protein expression of spinal TNFRI and cleaved caspase-11 was examined using western blots. The image of spinal mitochondrial superoxide was examined using MitoSox Red (mitochondrial superoxide indicator) image assay. Immunohistochemistry was used to examine the location of TNFRI and caspase-11 in the SCDH. Intrathecal administration of antisense oligodeoxynucleotide (AS-ODN) against TNFRI, caspase-11 siRNA, or a scavenger of mitochondrial superoxide was given for antinociceptive effects. Statistical tests were done using analysis of variance (1- or 2-way), or 2-tailed t test., Results: Intrathecal gp120/M induced mechanical allodynia and thermal hyperalgesia lasting for 3 weeks ( P < .001). Gp120/M increased the expression of spinal TNFRI, mitochondrial superoxide, and cleaved caspase-11. Immunohistochemistry showed that TNFRI and caspase-11 were mainly expressed in the neurons of the SCDH. Intrathecal administration of antisense oligonucleotides against TNFRI, Mito-Tempol (a scavenger of mitochondrial superoxide), or caspase-11 siRNA reduced mechanical allodynia and thermal hyperalgesia in the gp120/M neuropathic pain model. Spinal knockdown of TNFRI reduced MitoSox profile cell number in the SCDH; intrathecal Mito-T decreased spinal caspase-11 expression in gp120/M rats. In the cultured B35 neurons treated with TNFα, pretreatment with Mito-Tempol reduced active caspase-11 in the neurons., Conclusions: These results suggest that spinal TNFRI-mtROS-caspase 11 signal pathway plays a critical role in the HIV-associated neuropathic pain state, providing a novel approach to treating chronic pain in PLWH with opioids., Competing Interests: Conflicts of Interest: See Disclosures at the end of the article., (Copyright © 2023 International Anesthesia Research Society.)

The present study aimed to elucidate the effect of sulforaphane (a natural isothiocyanate) on oxidative stress and mitochondrial dysfunction during and at selected periods following status epilepticus (SE) induced in immature 12-day-old rats by Li-pilocarpine. Dihydroethidium was employed for the detection of superoxide anions, immunoblot analyses for 3-nitrotyrosine (3-NT) and 4-hydroxynonenal (4-HNE) levels and respiratory chain complex I activity for evaluation of mitochondrial function. Sulforaphane was given i.p. in two doses (5 mg/kg each), at PD 10 and PD 11, respectively. The findings of the present study indicate that both the acute phase of SE and the early period of epileptogenesis (1 week and 3 weeks following SE induction) are associated with oxidative stress (documented by the enhanced superoxide anion production and the increased levels of 3-NT and 4-HNE) and the persisting deficiency of complex I activity. Pretreatment with sulforaphane either completely prevented or significantly reduced markers of both oxidative stress and mitochondrial dysfunction. Since sulforaphane had no direct anti-seizure effect, the findings suggest that the ability of sulforaphane to activate Nrf2 is most likely responsible for the observed protective effect. Nrf2-ARE signaling pathway can be considered a promising target for novel therapies of epilepsy, particularly when new compounds, possessing inhibitory activity against protein-protein interaction between Nrf2 and its repressor protein Keap1, with less "off-target" effects and, importantly, with an optimal permeability and bioavailability properties, become available commercially., (© 2023. The Author(s).)

Proteinase 3 (PR3) is the main target antigen of antineutrophil cytoplasmic antibodies (ANCAs) in PR3-ANCA-associated vasculitis. A small fraction of PR3 is constitutively exposed on the surface of quiescent blood neutrophils in a proteolytically inactive form. When activated, neutrophils expose an induced form of membrane-bound PR3 (PR3 mb ) on their surface as well, which is enzymatically less active than unbound PR3 in solution due to its altered conformation. In this work, our objective was to understand the respective role of constitutive and induced PR3 mb in the immune activation of neutrophils triggered by murine anti-PR3 mAbs and human PR3-ANCA. We quantified immune activation of neutrophils by the measurement of the production of superoxide anions and secreted protease activity in the cell supernatant before and after treatment of the cells by alpha-1 protease inhibitor that clears induced PR3 mb from the cell surface. Incubation of TNFα-primed neutrophils with anti-PR3 antibodies resulted in a significant increase in superoxide anion production, membrane activation marker exposition, and secreted protease activity. When primed neutrophils were first treated with alpha-1 protease inhibitor, we observed a partial reduction in antibody-induced neutrophil activation, suggesting that constitutive PR3 mb is sufficient to activate neutrophils. The pretreatment of primed neutrophils with purified antigen-binding fragments used as competitor significantly reduced cell activation by whole antibodies. This led us to the conclusion that PR3 mb promoted immune activation of neutrophils. We propose that blocking and/or elimination of PR3 mb offers a new therapeutic strategy to attenuate neutrophil activation in patients with PR3-ANCA-associated vasculitis., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Activation of NADPH oxidase (NOX) enzymes and the generation of reactive oxygen species and oxidative stress regulate vascular and renal function and contribute to the pathogenesis of hypertension. The present study examined the role of NOXA1/NOX1 function in vascular reactivity of renal and mesenteric resistance arteries/arterioles of wild-type and Noxa1 -/- mice. A major finding was that renal blood flow is less sensitive to acute stimulation by angiotensin II (ANG II) in Noxa1 -/- mice compared with wild-type mice, with a direct action on resistance arterioles independent of nitric oxide (NO) bioavailability. These functional results were reinforced by immunofluorescence evidence of NOXA1/NOX1 protein presence in renal arteries, afferent arterioles, and glomeruli as well as their upregulation by ANG II. In contrast, the renal vascular response to the thromboxane mimetic U46619 was effectively blunted by NO and was similar in both mouse genotypes and thus independent of NOXA1/NOX1 signaling. However, phenylephrine- and ANG II-induced contraction of isolated mesenteric arteries was less pronounced and buffering of vasoconstriction after acetylcholine and nitroprusside stimulation was reduced in Noxa1 -/- mice, suggesting endothelial NO-dependent mechanisms. An involvement of NOXA1/NOX1/O 2 •- signaling in response to ANG II was demonstrated with the specific NOXA1/NOX1 assembly inhibitor C25 and the nonspecific NOX inhibitor diphenyleneiodonium chloride in cultured vascular smooth muscle cells and isolated mesenteric resistance arteries. Collectively, our data indicate that the NOX1/NOXA1/O 2 •- pathway contributes to acute vasoconstriction induced by ANG II in renal and mesenteric vascular beds and may contribute to ANG II-induced hypertension. NEW & NOTEWORTHY Renal reactivity to angiotensin II (ANG II) is mediated by superoxide signaling produced by NADPH oxidase (NOX)A1/NOX1. Acute vasoconstriction of renal arteries by ANG was blunted in Noxa1 -/- compared with wild-type mice. NOXA1/NOX1/O 2 •- signaling was also observed in ANG II stimulation of vascular smooth muscle cells and isolated mesenteric resistance arteries, indicating that it contributes to ANG II-induced hypertension. A NOXA1/NOX1 assembly inhibitor (C25) has been characterized that inhibits superoxide production and ameliorates the effects of ANG II.

Objective To study the effect and mechanism of blueberry on regulating the mitochondrial inner membrane protein mitofilin/Mic60 in an in vitro model of metabolic dysfunction-associated liver disease (MAFLD). Methods L02 human hepatocytes were induced by free fatty acids (FFA) to establish MAFLD cell model. A normal group, a model group, an 80 μg/mL blueberry treatment group, a Mic60 short hairpin RNA (Mic60 shRNA) transfection group, and Mic60 knockdown combined with an 80 μg/mL blueberry treatment group were established. The intracellular lipid deposition was observed by oil red O staining, and the effect of different concentrations of blueberry pulp on the survival rate of L02 cells treated with FFA was measured by MTT assay. The levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), triglyceride (TG), total cholesterol (TC), superoxide dismutase (SOD) activity, glutathione (GSH) and malondialdehyde (MDA) contents were measured by visible spectrophotometry. The expression of reactive oxygen species (ROS) in hepatocytes was observed by fluorescence microscopy, and the mRNA and protein expression of Mic60 were detected by real-time quantitative PCR and Western blot analysis, respectively. Results After 24 hours of FFA stimulation, a large number of red lipid droplets in the cytoplasm of L02 cells was observed, and the survival rate of L02 cells treated with 80 μg/mL blueberry was higher. The results of ALT, AST, TG, TC, MDA and the fluorescence intensity of ROS in blueberry treated group were lower than those in model group, while the levels of SOD, GSH, Mic60 mRNA and protein in blueberry treated group were higher than those in model group. Conclusion Blueberry promotes the expression of Mic60, increases the levels of SOD and GSH in hepatocytes, and reduces the production of ROS, thus alleviating the injury of MAFLD hepatocytes and regulating the disorder of lipid metabolism.

Mitochondrial involvement in neurodegenerative diseases is widespread and multifactorial. Targeting mitochondrial pathology is therefore of interest. The recent development of bioactive molecules that modulate mitochondrial dynamics (fusion, fission and motility) offers a new therapeutic approach for neurodegenerative diseases with either indirect or direct mitochondrial involvement. Here, we asked: (1) Can enhanced mitochondrial fusion and motility improve secondary mitochondrial pathology in superoxide dismutase1 (SOD1) mutant amyotrophic lateral sclerosis (ALS)? And: (2) What is the impact of enhancing mitochondria fitness on in vivo manifestations of SOD1 mutant ALS? We observed that small molecule mitofusin activators corrected mitochondrial fragmentation, depolarization and dysmotility in genetically diverse ALS patient reprogrammed motor neurons and fibroblasts, and in motor neurons, sensory neurons and fibroblasts from SOD1 G93A mice. Continuous, but not intermittent, pharmacologic mitofusin activation delayed phenotype progression and lethality in SOD1 G93A mice, reducing neuron loss and improving neuromuscular connectivity. Mechanistically, mitofusin activation increased mitochondrial motility, fitness and residency within neuromuscular synapses; reduced mitochondrial reactive oxygen species production; and diminished apoptosis in SOD1 mutant neurons. These benefits were accompanied by improved mitochondrial respiratory coupling, despite characteristic SOD1 mutant ALS-associated downregulation of mitochondrial respiratory complexes. Targeting mitochondrial dysdynamism is a promising approach to alleviate pathology caused by secondary mitochondrial dysfunction in some neurodegenerative diseases., (© The Author(s) 2022. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Nemaline myopathies (NM) are a group of congenital myopathies that lead to muscle weakness and dysfunction. While 13 genes have been identified to cause NM, over 50% of these genetic defects are due to mutations in nebulin (NEB) and skeletal muscle actin (ACTA1), which are genes required for normal assembly and function of the thin filament. NM can be distinguished on muscle biopsies due to the presence of nemaline rods, which are thought to be aggregates of the dysfunctional protein. Mutations in ACTA1 have been associated with more severe clinical disease and muscle weakness. However, the cellular pathogenesis linking ACTA1 gene mutations to muscle weakness are unclear To evaluate cellular disease phenotypes, iPSC-derived skeletal myocytes (iSkM) harboring an ACTA1 H40Y point mutation were used to model NM in skeletal muscle. These were generated by Crispr-Cas9, and include one non-affected healthy control (C) and 2 NM iPSC clone lines, therefore representing isogenic controls. Fully differentiated iSkM were characterized to confirm myogenic status and subject to assays to evaluate nemaline rod formation, mitochondrial membrane potential, mitochondrial permeability transition pore (mPTP) formation, superoxide production, ATP/ADP/phosphate levels and lactate dehydrogenase release. C- and NM-iSkM demonstrated myogenic commitment as evidenced by mRNA expression of Pax3, Pax7, MyoD, Myf5 and Myogenin; and protein expression of Pax4, Pax7, MyoD and MF20. No nemaline rods were observed with immunofluorescent staining of NM-iSkM for ACTA1 or ACTN2, and these mRNA transcript and protein levels were comparable to C-iSkM. Mitochondrial function was altered in NM, as evidenced by decreased cellular ATP levels and altered mitochondrial membrane potential. Oxidative stress induction revealed the mitochondrial phenotype, as evidenced by collapsed mitochondrial membrane potential, early formation of the mPTP and increased superoxide production. Early mPTP formation was rescued with the addition of ATP to media. Together, these findings suggest that mitochondrial dysfunction and oxidative stress are disease phenotypes in the in vitro model of ACTA1 nemaline myopathy, and that modulation of ATP levels was sufficient to protect NM-iSkM mitochondria from stress-induced injury. Importantly, the nemaline rod phenotype was absent in our in vitro model of NM. We conclude that this in vitro model has the potential to recapitulate human NM disease phenotypes, and warrants further study., Competing Interests: Declaration of competing interest M.W.L. is the founder, CEO, and owner of Diverge Translational Science Laboratory. M.W.L. is or has recently been a member of advisory boards for Solid Biosciences, Taysha Gene Therapies, Astellas Gene Therapies (formerly Audentes Therapeutics), and Ichorion Therapeutics. M.W.L. is also a consultant for Astellas Gene Therapies (formerly Audentes Therapeutics), Encoded Therapeutics, Modis Therapeutics, Lacerta Therapeutics, Dynacure, AGADA Biosciences, Affinia Therapeutics, Biomarin, Locanabio, and Vertex Pharmaceuticals. M.W.L. receives research support from Astellas Gene Therapies, Solid Biosciences, Kate Therapeutics, Prothelia, Cure Rare Disease, and Rocket Pharma, Ultragenyx. S.M.L. is now a Sr. Application Scientist at Curi Bio and D.L.M is on Curi Bio's SAB. All other authors have no disclosures to declare., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Superoxide/hydrogen peroxide production by site IQ in complex I of the electron transport chain is conventionally assayed during reverse electron transport (RET) from ubiquinol to NAD. However, S1QELs (specific suppressors of superoxide/hydrogen peroxide production by site IQ) have potent effects in cells and in vivo during presumed forward electron transport (FET). Therefore, we tested whether site IQ generates S1QEL-sensitive superoxide/hydrogen peroxide during FET (site IQf), or alternatively, whether RET and associated S1QEL-sensitive superoxide/hydrogen peroxide production (site IQr) occurs in cells under normal conditions. We introduce an assay to determine if electron flow through complex I is thermodynamically forward or reverse: on blocking electron flow through complex I, the endogenous matrix NAD pool will become more reduced if flow before the challenge was forward, but more oxidised if flow was reverse. Using this assay we show in the model system of isolated rat skeletal muscle mitochondria that superoxide/hydrogen peroxide production by site IQ can be equally great whether RET or FET is running. We show that sites IQr and IQf are equally sensitive to S1QELs, and to rotenone and piericidin A, inhibitors that block the Q-site of complex I. We exclude the possibility that some sub-fraction of the mitochondrial population running site IQr during FET is responsible for S1QEL-sensitive superoxide/hydrogen peroxide production by site IQ. Finally, we show that superoxide/hydrogen peroxide production by site IQ in cells occurs during FET, and is S1QEL-sensitive., (© 2023 The Author(s).)

Mitochondrial aconitase (ACO2) has been postulated as a redox sensor in the tricarboxylic acid cycle. Its high sensitivity towards reactive oxygen and nitrogen species is due to its particularly labile [4Fe-4S] 2+ prosthetic group which yields an inactive [3Fe-4S] + cluster upon oxidation. Moreover, ACO2 was found as a main oxidant target during aging and in pathologies where mitochondrial dysfunction is implied. Herein, we report the expression and characterization of recombinant human ACO2 and its interaction with frataxin (FXN), a protein that participates in the de novo biosynthesis of Fe-S clusters. A high yield of pure ACO2 (≥99%, 22 ± 2 U/mg) was obtained and kinetic parameters for citrate, isocitrate, and cis-aconitate were determined. Superoxide, carbonate radical, peroxynitrite, and hydrogen peroxide reacted with ACO2 with second-order rate constants of 10 8 , 10 8 , 10 5 , and 10 2  M -1  s -1 , respectively. Temperature-induced unfolding assessed by tryptophan fluorescence of ACO2 resulted in apparent melting temperatures of 51.1 ± 0.5 and 43.6 ± 0.2 °C for [4Fe-4S] 2+ and [3Fe-4S] + states of ACO2, sustaining lower thermal stability upon cluster oxidation. Differences in protein dynamics produced by the Fe-S cluster redox state were addressed by molecular dynamics simulations. Reactivation of [3Fe-4S] + -ACO2 by FXN was verified by activation assays and direct iron-dependent interaction was confirmed by protein-protein interaction ELISA and fluorescence spectroscopic assays. Multimer modeling and protein-protein docking predicted an ACO2-FXN complex where the metal ion binding region of FXN approaches the [3Fe-4S] + cluster, supporting that FXN is a partner for reactivation of ACO2 upon oxidative cluster inactivation., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

The mycotoxin fusaric acid (FA) induces rapid oxidative burst leading to cell death in plants. At the same time, plant defence reactions are mediated by several phytohormones for instance ethylene (ET). However, previously conducted studies leave research gaps on how ET plays a regulatory role under mycotoxin exposure. Therefore, this study aims to the time-dependent effects of two FA concentrations (0.1 mM and 1 mM) were explored on the regulation of reactive oxygen species (ROS) in leaves of wild-type (WT) and ET receptor mutant Never ripe (Nr) tomatoes. FA induced superoxide and H 2 O 2 accumulation in both genotypes in a mycotoxin dose- and exposure time-dependent pattern. 1 mM FA activated NADPH oxidase (+34% compared to the control) and RBOH1 transcript levels in WT leaves. However, superoxide production was significantly higher in Nr with 62% which could contribute to higher lipid peroxidation in this genotype. In parallel, the antioxidative defence mechanisms were also activated. Both peroxidase and superoxide dismutase activities were lower in Nr but ascorbate peroxidase showed one-fold higher activity under 1 mM FA stress than in WT leaves. Interestingly, catalase (CAT) activity decreased upon FA in a time- and concentration-dependent manner and the encoding CAT genes were also downregulated, especially in Nr leaves at 20%. Ascorbate level was decreased and glutathione remained lower in Nr than WT plants under FA exposure. Conclusively, Nr genotype showed more sensitivity to FA-induced ROS suggesting that ET serves defence reactions of plants by activating several enzymatic and non-enzymatic antioxidants to detoxify excess ROS accumulation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Specific targeting of anti-cancer drugs to mitochondria is an emerging strategy to enhance cancer cell killing whilst simultaneously overcoming the problem of drug resistance, low bioavailability and limited clinical success of natural products. We have synthesized a mitochondria targeted derivative of Ethyl Ferulate (EF, a naturally occurring ester of ferulic acid), by conjugating it with triphenylphosphonium ion and compared its cytotoxicity with the parent molecule. Mito-Ethyl Ferulate (M-EF) was found to be more potent than EF (~ 400-fold) in inhibiting the growth of A549 and MCF-7 cells and suppressing the clonogenic potential of A549 cells. Notably, M-EF did not induce any cytotoxicity in normal cells (mouse normal fibroblast cells) up to a concentration of 25 μM. Furthermore, M-EF treatment induced significantly higher cell death in MCF-7 and A549 cells, as compared to EF via induction of apoptosis. M-EF treatment increased mitochondrial superoxide production and induced mitochondrial DNA damage and phosphorylation of JNK and AKT in A549 cells. Furthermore, M-EF induced increase in mitochondrial superoxide production and cytotoxicity was attenuated on pre-treatment with mitochondria-targeted antioxidant (mitoTEMPO) indicating the involvement of mitochondrial ROS in the cytotoxic effects of M-EF. Finally, in silico prediction revealed putative mitochondrial targets of M-EF which are known to regulate mitochondrial ROS and cell viability. In conclusion, the improved cytotoxic efficacy of M-EF exemplifies the use of mitochondria-specific drug delivery in future development of natural product based mitochondrial pharmacology., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Fulvic acid (FA) can significantly alleviate cadmium (Cd) stress, but the specific metabolic response of FA to Cd toxicity is still not clarified. In the present study, we used untargeted metabolomic [gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-mass spectrometry (LC-MS)] analysis to profile cucumber metabolism in response to Cd stress after spray application of FA. Our results showed that 331 differentially enriched metabolites (DEMs) were identified in leaf materials. These DEMs were enriched in 21 shared pathways in comparative groups of "Cd treatment vs. the control treatment" and "FA + Cd treatment vs. the Cd treatment." Specifically, treatment with FA significantly enhanced the organic acid content (citric acid, isocitric acid, 2-oxoglutaric acid, fumaric acid, and malic acid), which would contribute to provide sufficient substrates for the tricarboxylic acid (TCA) cycle and amino acid biosynthesis, thereby ensuring the normal production of energy and amino acid. At the same time, FA significantly increased the amino acid content (aspartate, citrulline, histidine, leucine, and phenylalanine). The accumulation of organic acid and amino acid can act as chelating agents for heavy metal ions and as scavengers of reactive oxygen species (ROS), thereby reducing intracellular oxidative damage. Furthermore, the application of FA improves antioxidant enzymes and accelerates ROS clearance. The improved contents of organic acid and amino acid, and the increased activity of antioxidant enzymes both played a central role in the reduction of malondialdehyde (MDA, 14.08%), hydrogen peroxide (H 2 O 2 , 61.70%) contents, and superoxide anion radical (O 2 - , 30.41%) production rate in plants under Cd stress. Taken together, the present study demonstrates the effects of FA on the antioxidant capacity and carbohydrate and amino acid metabolism of cucumber seedlings exposed to Cd stress, which provides comprehensive insights into the regulation of plants' response to Cd toxicity with FA was applied in cucumber., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Aims/hypothesis: Accumulating evidence suggests that leucocytes play a critical role in diabetes-induced vascular lesions and other abnormalities that characterise the early stages of diabetic retinopathy. However, the role of monocytes has yet to be fully investigated; therefore, we used Ccr2 -/- mice to study the role of CCR2 + inflammatory monocytes in the pathogenesis of diabetes-induced degeneration of retinal capillaries., Methods: Experimental diabetes was induced in wild-type and Ccr2 -/- mice using streptozotocin. After 2 months, superoxide levels, expression of inflammatory genes, leucostasis, leucocyte- and monocyte-mediated cytotoxicity against retinal endothelial cell death, retinal thickness and visual function were evaluated. Retinal capillary degeneration was determined after 8 months of diabetes. Flow cytometry of peripheral blood for differential expression of CCR2 in monocytes was assessed., Results: In nondiabetic mice, CCR2 was highly expressed on monocytes, and Ccr2 -/- mice lack CCR2 + monocytes in the peripheral blood. Diabetes-induced retinal superoxide, expression of proinflammatory genes Inos and Icam1, leucostasis and leucocyte-mediated cytotoxicity against retinal endothelial cells were inhibited in diabetic Ccr2-deficient mice and in chimeric mice lacking Ccr2 only from myeloid cells. In order to focus on monocytes, these cells were immuno-isolated after 2 months of diabetes, and they significantly increased monocyte-mediated endothelial cell cytotoxicity ex vivo. Monocytes from Ccr2-deficient mice caused significantly less endothelial cell death. The diabetes-induced retinal capillary degeneration was inhibited in Ccr2 -/- mice and in chimeric mice lacking Ccr2 only from myeloid cells., Conclusions/interpretation: CCR2 + inflammatory monocytes contribute to the pathogenesis of early lesions of diabetic retinopathy., (© 2022. The Author(s).)

Six gene splice variants of superoxide-generating NADPH oxidase 5 (Nox5) have been identified in humans, and they differ in the sequence of their N-terminal cytoplasmic domains, which comprise four EF-hand motifs. Here, we demonstrated that the Ca 2+ -dependent association and dissociation between the N- and C-terminal cytoplasmic domains of the Nox5β variant are affected by the alanine substitution of the conserved Ile-113 or Leu-115 at the connecting loop between the third and fourth EF-hand motifs. These substitutions impair the cell surface localization of Nox5β. In addition, the Nox5ε/S variant, lacking all EF-hand motifs, does not localize to the plasma membrane. Thus, the Ca 2+ -sensitive intramolecular interaction determines the Nox5 subcellular localization, that is, whether Nox5 variants generate superoxide in the extracellular or intracellular space., (© 2023 Federation of European Biochemical Societies.)

Retinitis pigmentosa (RP) is caused by many different mutations that promote the degeneration of rod photoreceptors and have no direct effect on cones. After the majority of rods have died cone photoreceptors begin to slowly degenerate. Oxidative damage contributes to cone cell death and it has been hypothesized that tissue hyperoxia due to reduced oxygen consumption from the loss of rods is what initiates oxidative stress. Herein, we demonstrate in animal models of RP that reduction of retinal hyperoxia by reducing inspired oxygen to continuous breathing of 11% O 2 reduced the generation of superoxide radicals in the retina and preserved cone structure and function. These data indicate that retinal hyperoxia is the initiating event that promotes oxidative damage, loss of cone function, and cone degeneration in the RP retina., Competing Interests: Declaration of competing interest None., (Copyright © 2023 Elsevier Inc. All rights reserved.)

In this study the antioxidant and neuroprotective activity of an enriched polysaccharide fraction (EPF) obtained from the fruiting body of cultivated P. eryngii was evaluated. Proximate composition (moisture, proteins, fat, carbohydrates and ash) was determined using the AOAC procedures. The EPF was extracted by using, in sequence, hot water and alkaline extractions followed by deproteinization and precipitation with cold ethanol. Total α- and β-glucans were quantified using the Megazyme International Kit. The results showed that this procedure allows a high yield of polysaccharides with a higher content of (1-3; 1-6)-β-D-glucans. The antioxidant activity of EPF was detected from the total reducing power, DPPH, superoxide, hydroxyl and nitric oxide radical scavenging activities. The EPF was found to scavenge DPPH, superoxide, hydroxyl and nitric oxide radicals with a IC50 values of 0.52 ± 0.02, 1.15 ± 0.09, 0.89 ± 0.04 and 2.83 ± 0.16 mg/mL, respectively. As assessed by the MTT assay, the EPF was biocompatible for DI-TNC1 cells in the range of 0.006-1 mg/mL and, at concentrations ranging from 0.05 to 0.2 mg/mL, significantly counteracted H 2 O 2 -induced reactive oxygen species production. This study demonstrated that polysaccharides extracted from P. eryngii might be used as functional food to potentiate the antioxidant defenses and to reduce oxidative stress.

Background and objectives : Taking into consideration the confirmed role of oxidative stress in ischemia/reperfusion injury and the insufficiency in knowledge regarding the phosphodiesterase 5 (PDE5)-mediated effects on the cardiovascular system, the aim of our study was to investigate the influence of two PDE5 inhibitors, tadalafil and vardenafil, with or without the addition of N(G)-nitro-L-arginine methyl ester (L-NAME), on oxidative stress markers, coronary flow and left ventricular function, both ex vivo and in vivo. Methods : This study included 74 male Wistar albino rats divided into two groups. In the first, 24 male Wistar rats were orally treated with tadalafil or vardenafil for four weeks in order to perform in vivo experiments. In the second, the hearts of 50 male Wistar albino were excised and perfused according to the Langendorff technique in order to perform ex vivo experiments. The hearts were perfused with tadalafil (10, 20, 50 and 200 nM), vardenafil (10, 20, 50 and 200 nM) and a combination of tadalafil/vardenafil and L-NAME (30 μM). The CF and oxidative stress markers, including nitrite bioaviability (NO 2 - ), superoxide anion radical (O 2 - ), and the index of lipid peroxidation, were measured in coronary effluent. Results : The L-arginin/NO system acts as the mediator in the tadalafil-induced effects on the cardiovascular system, while it seems that the vardenafil-induced increase in CF was not primarily induced by the NO system. Although tadalafil induced an increase in O 2 - in the two lowest doses, the general effects of both of the applied PDE5 inhibitors on oxidative stress were not significant. The ejection function was above 50% in both groups. Conclusions : Our results showed that both tadalafil and vardenafil improved the coronary perfusion of the myocardium and LV function by increasing the EF.

Hypertension, a multifactorial chronic inflammatory condition, is an important risk factor for neurovascular and neurodegenerative diseases, including stroke and Alzheimer's disease. These diseases have been associated with higher concentrations of circulating interleukin (IL)-17A. However, the possible role that IL-17A plays in linking hypertension with neurodegenerative diseases remains to be established. Cerebral blood flow regulation may be the crossroads of these conditions because regulating mechanisms may be altered in hypertension, including neurovascular coupling (NVC), known to participate in the pathogenesis of stroke and Alzheimer's disease. In the present study, the role of IL-17A on NVC impairment induced by angiotensin (Ang) II in the context of hypertension was examined. Neutralization of IL-17A or specific inhibition of its receptor prevents the NVC impairment ( p < 0.05) and cerebral superoxide anion production ( p < 0.05) induced by Ang II. Chronic administration of IL-17A impairs NVC ( p < 0.05) and increases superoxide anion production. Both effects were prevented with Tempol and NADPH oxidase 2 gene deletion. These findings suggest that IL-17A, through superoxide anion production, is an important mediator of cerebrovascular dysregulation induced by Ang II. This pathway is thus a putative therapeutic target to restore cerebrovascular regulation in hypertension.

Genetic and epidemiologic studies have significantly advanced our understanding of the genetic factors contributing to age-related macular degeneration (AMD). In particular, recent expression quantitative trait loci (eQTL) studies have highlighted POLDIP2 as a significant gene that confers risk of developing AMD. However, the role of POLDIP2 in retinal cells such as retinal pigment epithelium (RPE) and how it contributes to AMD pathology are unknown. Here we report the generation of a stable human RPE cell line ARPE-19 with POLDIP2 knockout using CRISPR/Cas, providing an in vitro model to investigate the functions of POLDIP2 . We conducted functional studies on the POLDIP2 knockout cell line and showed that it retained normal levels of cell proliferation, cell viability, phagocytosis and autophagy. Also, we performed RNA sequencing to profile the transcriptome of POLDIP2 knockout cells. Our results highlighted significant changes in genes involved in immune response, complement activation, oxidative damage and vascular development. We showed that loss of POLDIP2 caused a reduction in mitochondrial superoxide levels, which is consistent with the upregulation of the mitochondrial superoxide dismutase SOD2 . In conclusion, this study demonstrates a novel link between POLDIP2 and SOD2 in ARPE-19, which supports a potential role of POLDIP2 in regulating oxidative stress in AMD pathology.

Aims: Mdivi-1 (Md-1) is a well-known inhibitor of mitochondrial fission and mitophagy. The mitochondrial superoxide scavenger Mito-TEMPO (MT) exerts positive effects on the developmental competence of pig embryos. This study aimed to explore the adverse effects of Md-1 on developmental capacity in porcine embryos and the protective effects of MT against Md-1-induced injury., Main Methods: We exposed porcine embryos to Md-1 (10 and 50 μM) for 2 days after in vitro fertilization (IVF). MT (0.1 μM) treatment was applied for 4 days after exposing embryos to Md-1. We assessed blastocyst development, DNA damage, mitochondrial superoxide production, and mitochondrial distribution using TUNEL assay, Mito-SOX, and Mito-tracker, respectively. Subsequently, the expression of PINK1, DRP1, and p-DRP1Ser616 was evaluated via immunofluorescence staining and Western blot analysis., Key Findings: Md-1 compromised the developmental competence of blastocysts. Apoptosis and mitochondrial superoxide production were significantly upregulated in 50 μM Md-1-treated embryos, accompanied by a downregulation of p-DRP1Ser616, PINK1, and LC3B levels and lower mitophagy activity at the blastocyst stage. We confirmed the protective effects of MT against the detrimental effect of Md-1 on blastocyst developmental competence, mitochondrial fission, and DRP1/PINK1-mediated mitophagy activation. Eventually, MT recovered DRP1/PINK1-mediated mitophagy and mitochondrial fission by inhibiting superoxide production in Md-1-treated embryos., Significance: MT protects against detrimental effects of Md-1 on porcine embryos by suppressing superoxide production. These findings expand available scientific knowledge on improving outcomes of IVF., Competing Interests: Declaration of competing interest The authors declare that there are no conflicts of interest., (Copyright © 2022. Published by Elsevier Inc.)

This study aimed to investigate the phytotoxic effect of copper (Cu) and copper nanoparticles (CuONPs) and ameliorative potential of nitric oxide (NO) against these toxic materials in Sorghum vulgare Pers. seedlings. Data suggested that exposure of Cu and CuONPs significantly reduced growth, chlorophyll, carotenoids and protein in root and shoot, which coincided with increased Cu accumulation. However, addition of sodium nitroprusside (SNP, a donor of NO) lowered Cu and CuONPs mediated toxicity through restricting Cu accumulation and improving photosynthetic pigments and total soluble protein contents. Data further suggested that exposure of Cu and CuONPs significantly increased hydrogen peroxide (H2 O2 ), superoxide radicals (O2 •- ), and malondialdehyde (MDA) contents. Enhanced level of oxidative stress severely inhibited the enzymatic activities of glutathione reductase (GR), ascorbate peroxidase (APX), dehydroascorbate reductase (DHAR) and monodehydroascorbate reductase (MDHAR) but enhanced superoxide dismutase (SOD) and catalase (CAT) activity. However, addition of SNP positively regulated antioxidants enzymes activity, particularly the enzymes involved in the ascorbate-glutathione cycle to overcome Cu- and CuONPs-induced stress in Sorghum seedlings. Further, Cu and CuONPs enhanced accumulation of free proline through inducing Δ1 -pyrroline-5-carboxylate synthetase (P5CS) activity while lowering the proline dehydrogenase (PDH) activity. However, addition of SNP reversed these responses. Therefore, overall results revealed that SNP has enough potential of reducing the toxicity of Cu and CuONPs in Sorghum seedlings through regulation of proline metabolism and activity of enzymes of the ascorbate-glutathione cycle. These findings can be employed in developing new resistant varieties of Sorghum having enhanced tolerance against Cu or CuONP stress and improved productivity.

The mitochondrial bc1 complex is a major source of mitochondrial superoxide. While bc1-generated superoxide plays a beneficial signaling role, excess production of superoxide lead to aging and degenerative diseases. The catalytic core of bc1 comprises three peptides -cytochrome b, Fe-S protein, and cytochrome c1. All three core peptides exhibit accelerated evolution in anthropoid primates. It has been suggested that the evolution of cytochrome b in anthropoids was driven by a pressure to reduce the production of superoxide. In humans, the bc1 core peptides exhibit anthropoid-specific substitutions that are clustered near functionally critical sites that may affect the production of superoxide. Here we compare the high-resolution structures of bovine, mouse, sheep and human bc1 to identify structural changes that are associated with human-specific substitutions. Several cytochrome b substitutions in humans alter its interactions with other subunits. Most significantly, there is a cluster of seven substitutions, in cytochrome b, the Fe-S protein, and cytochrome c1 that affect the interactions between these proteins at the tether arm of the Fe-S protein and may alter the rate of ubiquinone oxidation and the rate of superoxide production. Another cluster of substitutions near heme bH and the ubiquinone reduction site, Qi, may affect the rate of ubiquinone reduction and thus alter the rate of superoxide production. These results are compatible with the hypothesis that cytochrome b in humans (and other anthropoid primates) evolve to reduce the rate of production of superoxide thus enabling the exceptional longevity and exceptional cognitive ability of humans., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Objectives: Erythrocytes are exposed to oxidative stress during storage and can be stored for up to 42 days (in AS-7) under blood bank conditions for transfusion. Vitamin-C and Vitamin-E have proved beneficial in diminishing oxidative stress. Therefore, this study aims to investigate the combined effects of Vitamin-C and Vitamin-E on erythrocytes during storage., Materials and Methods: Blood was collected from male Wistar rats and erythrocytes were isolated and stored in AS-7 (Additive Solution) at 4 °C for 35 days. Erythrocytes were grouped into i) Controls and ii) Experimentals [Vitamin-C (10 mM) and Vitamin-E (2 mM)]. Antioxidant and oxidative stress markers were assessed at weekly intervals. Statistical analyses were performed by using GraphPad Prism software., Results: Hemoglobin increased on days 7 and 14 in the Experimentals. Superoxide dismutase activity elevated on days 7 & 14 in Controls and on day 7 in Experimentals. Catalase activity increased on day 21 in both groups. Protein carbonyls decreased on days 21 and 28 in Experimentals. Thiobarbituric acid reactive substances decreased from day 14 in both groups. Conjugate dienes decreased on days 21 & 35 in the Experimentals. Glutathione increased from day 14 in both groups. Superoxides decreased on days 14, 28 & 35 in Controls and from day 14 in Experimentals., Conclusion: Vitamin-C and Vitamin-E have been beneficial in terms of hemoglobin, antioxidants, protein & lipid oxidations and superoxides in stored erythrocytes. Therefore, this study provides new avenues for the development of effective storage solutions which will have a clinical impact in erythrocyte transfusions., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Société française de transfusion sanguine (SFTS). Published by Elsevier Masson SAS. All rights reserved.)

Reactive oxygen species (ROS) represent a group of high reactive molecules with dualistic natures since they can induce cytotoxicity or regulate cellular physiology. Among the ROS, the superoxide anion radical (O2·-) is a key redox signaling molecule prominently generated by the NADPH oxidase (NOX) enzyme family and by the mitochondrial electron transport chain. Notably, altered redox balance and deregulated redox signaling are recognized hallmarks of cancer and are involved in malignant progression and resistance to drugs treatment. Since oxidative stress and metabolism of cancer cells are strictly intertwined, in this review, we focus on the emerging roles of NOX enzymes as important modulators of metabolic reprogramming in cancer. The NOX family includes seven isoforms with different activation mechanisms, widely expressed in several tissues. In particular, we dissect the contribute of NOX1, NOX2, and NOX4 enzymes in the modulation of cellular metabolism and highlight their potential role as a new therapeutic target for tumor metabolism rewiring.

Classically, superoxide anion O 2 •- and reactive oxygen species ROS play a dual role. At the physiological balance level, they are a by-product of O 2 reduction, necessary for cell signalling, and at the pathological level they are considered harmful, as they can induce disease and apoptosis, necrosis, ferroptosis, pyroptosis and autophagic cell death. This revision focuses on understanding the main characteristics of the superoxide O 2 •- , its generation pathways, the biomolecules it oxidizes and how it may contribute to their modification and toxicity. The role of superoxide dismutase, the enzyme responsible for the removal of most of the superoxide produced in living organisms, is studied. At the same time, the toxicity induced by superoxide and derived radicals is beneficial in the oxidative death of microbial pathogens, which are subsequently engulfed by specialized immune cells, such as neutrophils or macrophages, during the activation of innate immunity. Ultimately, this review describes in some depth the chemistry related to O 2 •- and how it is harnessed by the innate immune system to produce lysis of microbial agents.

A novel electrochemical biosensor was developed to monitor fibroblast cells stress levels for the first time in situ under external stimuli based on the recognition of superoxide anion released upon cell damage. The biosensor comprised metallized polycaprolactone electrospun fibers covered with zinc oxide for improved cell adhesion and signal transduction, whilst stable bioconjugates of mercaptobenzoic acid-functionalized gold nanoparticles/superoxide dismutase were employed as recognition bioelements. Biosensors were first tested and optimized for in situ generated superoxide detection by fixed potential amperometry at +0.3 V, with minimal interferences from electroactive species in cell culture media. L929 fibroblast cells were then implanted on the optimized biosensor surface and the biosensor morphologically characterized by scanning electron microscopy (SEM) and fluorescence microscopy, which illustrated the network-type pattern of fibroblasts adjacent to the fiber scaffold. Fibroblast stress was induced by zymosan and monitored at the cells integrated biosensor using fixed potential amperometry (CA) with a sensitivity of 26 nA cm -2 μg mL -1 zymosan and electrochemical impedance spectroscopy (EIS), with similar sensitivity of the biosensor considering the Rs and Z' parameters of around 0.13 Ω cm 2 μg -1 mL and high correlation factors R 2 of 0.9994. The obtained results underline the applicability of the here developed biosensor for the electrochemical screening of the fibroblast cells stress. The concept in using low-cost biocompatible polymeric fibers as versatile scaffolds for both enzyme immobilization and cell adhesion, opens a new path in developing biosensors for the in-situ investigation of a variety of cellular events., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Nanozymes with superoxide dismutase (SOD)-like activity have attracted increasing interest due to their ability to scavenge superoxide anion, the origin of most reactive oxygen species in vivo. However, SOD nanozymes reported thus far have yet to approach the activity of natural enzymes. Here, we report a carbon dot (C-dot) SOD nanozyme with a catalytic activity of over 10,000 U/mg, comparable to that of natural enzymes. Through selected chemical modifications and theoretical calculations, we show that the SOD-like activity of C-dots relies on the hydroxyl and carboxyl groups for binding superoxide anions and the carbonyl groups conjugated with the π-system for electron transfer. Moreover, C-dot SOD nanozymes exhibit intrinsic targeting ability to oxidation-damaged cells and effectively protect neuron cells in the ischemic stroke male mice model. Together, our study sheds light on the structure-activity relationship of C-dot SOD nanozymes, and demonstrates their potential for treating of oxidation stress related diseases., (© 2023. The Author(s).)

Parkinson's disease (PD) is characterized by dopaminergic cell loss in the substantia nigra, and PD brains show neuroinflammation, oxidative stress, and mitochondrial dysfunction. The study evaluated the neuroprotective activity of 1α,25-dihydroxy vitamin D3 (VD3), on the rotenone (ROT)-induced cytotoxicity in PC12 cells. The viability parameters were assessed by the MTT and flow cytometry, on cells treated or not with VD3 and/or ROT. Besides, ROS production, cell death, mitochondrial transmembrane potential, reduced GSH, superoxide accumulation, molecular docking (TH and Keap1-Nrf2), and TH, Nrf2, NF-kB, and VD3 receptor protein contents by western blot were evaluated. VD3 was shown to improve the viability of ROT-exposed cells. Cells exposed to ROT showed increased production of ROS and superoxide, which decreased after VD3. ROT decrease in the mitochondrial transmembrane potential was prevented, after VD3 treatment and, VD3 was shown to interact with tyrosine hydroxylase (TH) and Nrf2. While ROT decreased TH, Nrf2, and NF-kB expressions, these effects were reversed by VD3. In addition, VD3 also increased VD3 receptor protein contents and values went back to those of controls after ROT exposure. VD3 protects PC12 cells against ROT damage, by decreasing oxidative stress and improving mitochondrial function. One target seems to be the TH molecule and possibly an indirect Nrf2 activation could also justify its neuroprotective actions on this PC12 cell model of PD., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

To understand the mitigation effects of melatonin on the chilling-induced photoinhibition in tomato, four groups of seedlings were labelled: NW (normal temperature + water), NM (normal temperature + melatonin), CW (chilling + water) and CM (chilling+ melatonin). We measured chlorophyll fluorescence, key photosynthetic parameters and the cycle efficiency for chloroplast ascorbic acid-glutathione (AsA-GSH). The results showed that, compared with the NW control, photosynthesis rate in CW was decreased by 50.3%-72.6%, chloroplast malondialdehyde content was decreased by 17.5%-132.7%, superoxide anion production was increased by 86.5%-235.9%, and H 2 O 2 was increased by 96.6%-208.4%. These trends were significantly alleviated by exogenous melatonin, with photosynthetic rates in CM being increased by 22.7%-24.7% compared with in CW, malondialdehyde content being decreased by 16.6%-29.0%, the rate of superoxide anion production being decreased by 14.9%-22.7%, H 2 O 2 content being decreased by 10.7%-27.1%. Compared with CW, the quantum yield of photochemical energy in PS Ⅱ was increased by 15.8% in CM, the quantum yield of regulated non-photochemical energy loss was increased by 7.2%, the quantum yield of non-regulated non-photochemical energy loss was decreased by 24.7%, and the activities of key metabolic enzymes in the AsA-GSH cycle were increased to different degrees. We concluded that exogenous melatonin application could alleviate photoinhibition in tomato seedlings under chilling by balancing the partitioning of absorption energy in PS Ⅱ and by enhancing the ROS scavenging efficiency of the AsA-GSH cycle in the chloroplast.

Nitric Oxide (NO) is a highly diffusible, ubiquitous signaling molecule and a free radical that is naturally synthesized by our body. The pleiotropic effects of NO in biological systems are due to its reactivity with different molecules, such as molecular oxygen (O 2 ), superoxide anion, DNA, lipids, and proteins. There are several contradictory findings in the literature pertaining to its role in oncology. NO is a Janus-faced molecule shown to have both tumor promoting and tumoricidal effects, which depend on its concentration, duration of exposure, and location. A high concentration is shown to have cytotoxic effects by triggering apoptosis, and at a low concentration, NO promotes angiogenesis, metastasis, and tumor progression. Upregulated NO synthesis has been implicated as a causal factor in several pathophysiological conditions including cancer. This dichotomous effect makes it highly challenging to discover its true potential in cancer biology. Understanding the mechanisms by which NO acts in different cancers helps to develop NO based therapeutic strategies for cancer treatment. This review addresses the physiological role of this molecule, with a focus on its bimodal action in various types of cancers., Competing Interests: Declaration of Competing Interest The authors declare that there are no conflicts of interest., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Free radicals, such as hydroxyl radical, superoxide, and lipid-derived radical, have unpaired electrons, making them a highly reactive chemical species. They play important physiological roles, for example, in the removal of xenobiotic substances, such as bacteria and viruses, and in the production of chemical mediators, like prostaglandins and leukotrienes. However, excessive production of free radicals can cause structural defects in biomolecules like DNA and proteins, resulting in a loss of their normal functions. Hence, free radicals have been implicated in the onset and progression of various diseases such as cancer, atherosclerosis, and neurodegenerative diseases. However, there is very little clarity on the substantial amount, type, and location of free radicals in vivo, under pathological conditions. An investigation on the actual state of free radicals in vivo could lead to the diagnosis of pathological conditions and the elucidation of the mechanisms of their onset and progression; therefore, the development of in vivo radical detection methods is being widely pursued. Toward this end, nuclear medical imaging methods have recently attracted attention. In this study, we discuss the development of a nuclear medical imaging probe for the specific targeting of lipid radicals.

Irisin is a hormone able to reproduce some of the positive effects of physical activity and diet. Recently, we demonstrated the presence of Irisin at the ovarian level as a potential physiological regulator of follicular function. Adipose tissue is crucial for reproductive function through its metabolic activity and the production of adipokines. At present, the exact nature of adipocyte precursors is still under debate, but an important role has been assigned to the population of adipose tissue mesenchymal stromal cells (ASCs) of perivascular origin. It should be noted that, when appropriately stimulated, ASCs can differentiate into preadipocytes and, subsequently, adipocytes. Therefore, this present study was undertaken to explore the potential effect of Irisin on ASCs, known for their high differentiative potential. Since Irisin expression in ASCs was confirmed by PCR, we tested its potential effects on the main functional activities of these cells, including proliferation (BrdU uptake); metabolic activity (ATP production); redox status, evaluated as the generation of free molecules such as superoxide anion and nitric oxide; and scavenger activities, assessed as both enzymatic (superoxide dismutase) and non-enzymatic antioxidant power. Moreover, we tested the effect of Irisin on ASCs adipogenic differentiation. BrdU uptake was significantly ( p < 0.001) inhibited by Irisin, while ATP production was significantly ( p < 0.05) increased. Both superoxide anion and nitric oxide generation were significantly increased ( p < 0.001) by Irisin, while scavenger activity was significantly reduced ( p < 0.05). Irisin was found to significantly ( p < 0.05) inhibit ASCs adipogenic differentiation. Taken together, the present results suggest a potential local role of Irisin in the regulation of adipose tissue function.

Brevilin A (BA), a sesquiterpene lactone isolated from Centipeda minima herb, has been identified to exhibit potent anticancer activity. However, the potential pharmacological effect and mechanism of BA in regulating endothelial cell (EC) angiogenesis, a key event in tumor growth, is poorly understood. In this study, BA was shown to significantly prevent vascular endothelial growth factor (VEGF) induced EC angiogenic capacities in vitro , ex vivo, and in vivo . Subsequent functional assays revealed that BA dose dependently inhibited VEGF-stimulated survival, proliferation, migration, and triggered apoptosis activity in human umbilical vein endothelial cells (HUVECs), as well as suppressed the expression of antiapoptotic protein Bcl-2, increased the expression of proapoptotic protein caspase-3 and Bax, and suppressed PI3K/AKT pathway. Meanwhile, BA was also able to depolarize mitochondrial membranal permeability (MMP), accelerate mitochondrial superoxide accumulation, induce intracellular reactive oxygen species (ROS) production, and decreased intracellular glutathione (GSH) in HUVECs. Furthermore, both mitochondria-specific superoxide scavenger Mito-TEMPOL and broad-spectrum antioxidant N-acetyl-cysteine (NAC) dramatically abolished BA-induced mitochondrial dysfunction and mitochondrial ROS production, causing the reversion of PI3K/AKT pathway and repression of apoptosis, eventually correcting the impaired endothelial behavior in survival, growth, migration, and angiogenesis. Collectively, our data for the first time identified a new mechanism for antiangiogenic effect of BA in vascular EC, one that is based on the regulation of mitochondrial-dependent ROS overproduction., Competing Interests: The authors declare that they have no competing interests., (Copyright © 2022 Bailing Wei et al.)

Biscembranoids are the distinctive tetraterpenoids owing a 14/6/14 membered tricyclic scaffold that have been mainly discovered in the soft corals, especially the genera Sarcophyton , Lobophytum and Sinularia . Recent findings have demonstrated the great anti-inflammatory potential of biscembranoid analogues in human neutrophils, motivating more chemical and biological explorations targeting these marine-derived natural products. In the current study, the chemical diversity of biscembranoids derived from the cultured-type Sarcophyton trocheliophorum von Marenzeller was illustrated through MS/MS molecular networking (MN) profiling approach. Based on the MN patterns, the prioritization of unknown biscembranoid derivatives was putatively analyzed. As a result, the biscembrane targeting isolation afforded two new metabolites, sarcotrochelides A ( 1 ) and B ( 2 ), along with six known analogues ( 3 - 8 ). Their structures and relative configurations were determined by spectroscopic methods. In vitro neutrophil inflammatory inhibition was further investigated for all isolates based on reduced superoxide anion (O 2 •- ) generation detections. Compounds 5 - 8 showed significant dose-dependently inhibitory effects, suggesting the cruciality of 6,7-dihydrooxepin-2(5H)-one moiety and saturated γ-lactone ring in their reactive oxygen species (ROS)-dependent anti-inflammatory properties.

3,4-Methylenedioximethamphetamine (MDMA; "ecstasy") is a psychotropic drug with well-known neurotoxic effects mediated by hitherto not fully understood mechanisms. The Na + - and K + -activated adenosine 5'-triphosphatase (Na + /K + ATPase), by maintaining the ion gradient across the cell membrane, regulates neuronal excitability. Thus, a perturbation of its function strongly impacts cell homeostasis, ultimately leading to neuronal dysfunction and death. Nevertheless, whether MDMA affects the Na + /K + ATPase remains unknown. In this study, we used synaptosomes obtained from whole mouse brain to test the effects of MDMA, three of its major metabolites [α-methyldopamine, N-methyl-α-methyldopamine and 5-(glutathion-S-yl)-α-methyldopamine], serotonin (5-HT), dopamine, 3,4-dihydroxy-L-phenylalanine (L-Dopa) and 3,4-dihydroxyphenylacetic acid (DOPAC) on the Na + /K + ATPase function. A concentration-dependent increase of Na + /K + ATPase activity was observed in synaptosomes exposed to the tested compounds (concentrations ranging from 0.0625 to 200 µM). These effects were independent of protein kinases A and C activities. Nevertheless, a rescue of the compounds' effects was observed in synaptosomes pre-incubated with the antioxidant N-acetylcysteine (1 mM), suggesting a role for reactive species-regulated pathways on the Na + /K + ATPase effects. In agreement with this hypothesis, a similar increase in the pump activity was found in synaptosomes exposed to the chemical generator of superoxide radicals, phenazine methosulfate (1-250 µM). This study demonstrates the ability of MDMA metabolites, monoamine neurotransmitters, L-Dopa and DOPAC to alter the Na + /K + ATPase function. This could represent a yet unknown mechanism of action of MDMA and its metabolites in the brain., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Anti- insecticidal potential of daidzein was studied by feeding second instar larvae of Spodoptera litura (Fabricius) on artificial diet incorporated with different concentrations (5 ppm, 25 ppm, 125 ppm, 625 ppm) of diadzein. Results revealed high larval mortality, prolongation of pupal and total developmental period of the larvae treated with diadzein. Anti-nutritional/post ingestive toxicity of diadzein was also revealed by the decrease in the nutritional indices such as relative growth rate (RGR), relative consumption rate (RCR), efficiency of conversion of digested food (ECD), efficiency of conversion of ingested food (ECI) and approximate digestibility (AD). The suppression of immune function due to decline in the total hemocytes count was also observed in treated S. litura larvae. Profiles of detoxifying enzymes viz. superoxide dismutases (SOD), catalase (CAT), ascorbate peroxidases (APOX) and glutathione S-transferase (GST) were also significantly increased with diadzein treatment. The hydrogen peroxide content (H 2 O 2 ), lipid peroxide content (LP) and protein carbonyl content were also significantly enhanced in the treated larvae thus, indicating oxidative stress in the insect. Our findings suggest that daidzein can be used as the alternative to conventional pesticides for controlling S. litura population., Competing Interests: Declaration of competing interest This manuscript has not been submitted or accepted for publication elsewhere. All authors have approved the content and carry no conflict of interests., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Background: Age-associated comorbidities are higher in people with HIV (PWH) than HIV-negative individuals. This is partially attributed to immune activation and CD38 expression on T cells driving chronic inflammation. However, the exact contribution of CD38-expressing T cells on the proinflammatory response is not completely understood., Methods: CD38-expressing CD8 + T lymphocytes were measured from PWH and HIV-negative individuals. Mitochondrial mass, superoxide content, membrane depolarization of CD4 + and CD8 + T lymphocytes, and cytokine production after HIV(Gag)-specific peptide stimulation from CD38 + CD8 + T lymphocytes of PWH were measured to link biological effects of CD38 expression on cellular metabolism., Results: The frequency of activated CD8 + CD38 + T cells persists in PWH on ART compared with HIV-negative individuals. Higher CD38 expression is associated with mitochondrial biogenesis and HIV(Gag)-specific proinflammatory cytokine production in PWH. Blockade of CD38 results in lower Gag-specific cytokine production., Conclusions: ART only partially reduced HIV-induced CD38 expression on CD8 + T cells. CD8 + CD38 + T cells are highly activated in vivo, and HIV-specific stimulation in vitro augments CD38 expression, contributing to a proinflammatory response despite virologic control with ART. Therefore, CD38 is a potential therapeutic target for mitigating chronic inflammation that likely drives cellular aging, comorbidities, and end-organ disease in PWH., Competing Interests: P.M. is supported by NIH grant 5K12HL143886-02 and reports personal fees from Med Learning Group and NAACME, outside the submitted work. S.K. reports grants from Arbutus Pharmaceuticals, Merck Inc, Silverback Therapeutics, Zhuhai Yufan Biotechnologies, The Liver Company, and Regeneron Pharmaceuticals, outside the submitted work. The remaining authors have no funding or conflicts of interest to disclose., (Copyright © 2022 Wolters Kluwer Health, Inc. All rights reserved.)

The purpose of this research was to determine expression profiles of metacaspase (MC) genes during ripening and senescence of two apple cultivars with distinct ripening behavior. "Golden Delicious" and "Fuji" harvested at commercial maturity were used as materials. Our data revealed that flesh firmness, respiration rate, ethylene production, metacaspase (MC) activity, superoxide anion (O 2 •- ) production rate, relative electrical conductivity (REC), hydrogen peroxide (H 2 O 2 ), and malondialdehyde (MDA) contents in "Golden Delicious" were higher than in "Fuji" during ripening. At 35 days, no DNA ladder was observed in both cultivars, and tonoplast disintegration was only observed in "Golden Delicious" by transmission electron microscope (TEM), indicating that programmed cell death (PCD) was initiated earlier in "Golden Delicious" than in "Fuji." A total of 18 MC genes were detected to be expressed in both cultivars. For those genes expressed only in "Golden Delicious," MdMC06, MdMC10, MdMC12, and MdMC21 might play a role in the early stage of ripening, whereas MdMC19 might be associated with the late stage of ripening. MdMC15 was expressed only in "Fuji." The remaining MC genes were differentially expressed in both cultivars during ripening. These results would provide useful information to further underlie the relationships among MC genes, PCD and storability of different apple cultivars. PRACTICAL APPLICATIONS: Apple is one of the most popular fruits in the world. Different apple cultivars vary in their ripening behavior and storability, but the molecular mechanism has not yet been fully elucidated. "Golden Delicious" and "Fuji" are two important apple cultivars worldwide. Our data indicated that PCD was initiated earlier in "Golden delicious" than in "Fuji" during postharvest ripening and senescence. MdMCs showed variable expression patterns in both cultivars during ripening. MdMC15 and MdMC19 might be closely associated with the early stage of PCD. These results would provide useful information to further decode the molecular mechanisms responsible for different storage storability of apple cultivars., (© 2022 Wiley Periodicals LLC.)

The NOD-like receptor pyrin domain 3 (NLRP3) inflammasome is activated during atherogenesis, but how this occurs is unclear. Here, we explored the mechanisms activating and regulating NLRP3 inflammasomes via the acid sphingomyelinase (ASM)-ceramide signaling pathway. As a neointima formation model, partial left carotid ligations were performed on endothelial cell (EC)-specific ASM transgene mice (Smpd1 trg /EC cre ) and their control littermates (Smpd1 trg /WT and WT/WT) fed on the Western diet (WD). We found neointima formation remarkably increased in Smpd1 trg /EC cre mice over their control littermates. Next, we observed enhanced colocalization of NLRP3 versus adaptor protein ASC (the adaptor molecule apoptosis-associated speck-like protein containing a CARD) or caspase-1 in the carotid ECs of WD-treated Smpd1 trg /EC cre mice but not in their control littermates. In addition, we used membrane raft (MR) marker flotillin-1 and found more aggregation of ASM and ceramide in the intima of Smpd1 trg /EC cre mice than their control littermates. Moreover, we demonstrated by in situ dihydroethidium staining, carotid intimal superoxide levels were much higher in WD-treated Smpd1 trg /EC cre mice than in their control littermates. Using ECs from Smpd1 trg /EC cre and WT/WT mice, we showed ASM overexpression markedly enhanced 7-ketocholesterol (7-Ket)-induced increases in NLRP3 inflammasome formation, accompanied by enhanced caspase-1 activity and elevated interleukin-1β levels. These 7-Ket-induced increases were significantly attenuated by ASM inhibitor amitriptyline. Furthermore, we determined that increased MR clustering with NADPH oxidase subunits to produce superoxide contributes to 7-Ket-induced NLRP3 inflammasome activation via a thioredoxin-interacting protein-mediated controlling mechanism. We conclude that ceramide from ASM plays a critical role in NLRP3 inflammasome activation during hypercholesterolemia via MR redox signaling platforms to produce superoxide, which leads to TXNIP dissociation., Competing Interests: Conflict of Interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Context: Endoplasmic reticulum (ER) stress contributes to endothelium pathological conditions. Chitooligosaccharides (COS) have health benefits, but their effect on endothelial cells is unknown. We demonstrate for the first time a protective effect of COS against ER-induced endothelial cell damage., Objective: To evaluate the protective effect of COS on ER stress-induced apoptosis in endothelial cells., Material and Methods: Endothelial (EA.hy926) cells were pre-treated with COS (250 or 500 μg/mL) for 24 h, and then treated with 0.16 μg/mL of Tg for 24 h and compared to the untreated control. Apoptosis and necrosis were detected by Annexin V-FITC/propidium iodide co-staining. Reactive oxygen species (ROS) were measured with the DCFH 2 -DA and DHE probes. The protective pathway and ER stress markers were evaluated by reverse transcription-polymerase chain reaction, western blot, and immunofluorescence analyses., Results: COS attenuated ER stress-induced cell death. The viability of EA.hy926 cells treated with Tg alone was 44.97 ± 1% but the COS pre-treatment increased cells viability to 74.74 ± 3.95% in the 250 μg/mL COS and 75.34 ± 2.4% in the 500 μg/mL COS treatments. Tg induced ER stress and ROS, which were associated with ER stress-mediated death. Interestingly, COS reduced ROS by upregulating nuclear factor-E2-related factor 2 (Nrf2), and the oxidative enzymes, superoxide dismutase1 (SOD1) and catalase. COS also suppressed up-regulation of the ER-related apoptosis protein, CHOP induced by Tg., Conclusions: COS protected against ER stress-induced apoptosis in endothelial cells by suppressing ROS and up-regulation Nrf2 and SOD1. These findings support the use of COS to protect endothelial cells.

Pigeon pea protein was sequentially digested with pepsin followed by pancreatin and the hydrolysate separated into 18 fractions using reversed-phase high-performance liquid chromatography. Fractions were analyzed for in vitro antioxidant properties (radical scavenging, metal chelation, and ferric iron reducing ability) in addition to inhibition of renin and angiotensin-converting enzyme (ACE). The most active fractions were analyzed by mass spectroscopy followed by identification of 10 peptide sequences (7 pentapeptides and 3 hexapeptides). All the peptides showed a wide range of multifunctional activity by scavenging hydroxyl (31.9-66.8%) and superoxide (25.6-100.0%) radicals in addition to ACE inhibition (7.4-100%) with significant (p < .05) differences between the peptides. AGVTVS, TKDIG, TSRLG, GRIST, and SGEKI were the most active; however, AGVTVS had the highest hydrophobic residue and exhibited the strongest activity against ACE, renin as well as superoxide and hydroxyl radicals. PRACTICAL APPLICATIONS: There is an increasing attraction of researchers to food peptides especially from legume proteins. Enzymatic digestion as well as high performance liquid chromatography (HPLC) purification has become an important process used to separate peptides with significant biological activities and health-promoting effects. There is useful information regarding the bioactive and functional (in vitro antioxidant, antidiabetic, in vitro/in vivo antihypertensive) properties of hydrolyzed and ultra-filtered pigeon pea fractions but scant research output still exists for purified peptides from pigeon pea establishing their therapeutic potential. The present study aimed to separate peptide fractions from pigeon pea hydrolysate and identify available amino acid sequences from the parent protein. Therefore, peptide sequences generated from the most bioactive fractions showed prospects for the expanded industrial utilization of pigeon pea. Further promoting its application as functional ingredient or additive for alleviating angiotensin-converting enzyme-related diseases., (© 2022 Wiley Periodicals LLC.)

Circadian rhythm plays a significant role in maintaining the function of the cardiovascular system. Emerging studies have demonstrated that circadian disruption enhances the risk of cardiovascular diseases by activating the sympathetic nervous system; however, the underlying mechanisms remain unknown. Therefore, this study aimed to clarify the role of oxidative stress in the rostral ventrolateral medulla (RVLM) in sympathetic hyperactivity induced by circadian disruption. Rats were randomly divided into two groups: the normal light and dark (LD) group and the circadian disruption (CD) group. Sympathetic nerve activity of rats was assessed by recording renal sympathetic nerve activity (RSNA) and indirect methods such as plasma level of norepinephrine (NE). The level of oxidative stress in the RVLM was detected by dihydroethidium probes. Moreover, the expression levels of the oxidative stress-related proteins in the RVLM were detected by Western blotting. Circadian disruption significantly increased blood pressure (BP), RSNA, and plasma levels of NE. Compared to the LD group, the CD group exhibited a more significant depressor response to i.v. hexamethonium bromide, a ganglionic blocker. Furthermore, the reactive oxygen species (ROS) production in the RVLM of rats with circadian disruption was significantly increased. In addition, BP and RSNA of rats with circadian disruption exhibited a greater decrease in the effects of microinjection of tempol, a superoxide scavenger, into the RVLM, compared to artificial cerebrospinal fluid (aCSF). Further investigation of the molecular mechanism by Western blotting showed that nuclear factor-erythroid-2-related factor 2 (Nrf2)/heme oxygenase 1 (HO1)/NAD(P)H: quinone oxidoreductase 1 (NQO1) signaling was down-regulated in the RVLM of circadian disruption rats. These data suggest that oxidative stress in the RVLM mediates sympathetic hyperactivity induced by circadian disruption and possibly by down-regulating Nrf2/HO1/NQO1 signaling., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier B.V. All rights reserved.)

A characteristic of cytochrome P450 (CYP) enzymes is their ability to generate H 2 O 2 , either directly or indirectly via superoxide anion, a reaction referred to as "NADPH oxidase" activity. H 2 O 2 production by CYPs can lead to the accumulation of cytotoxic reactive oxygen species which can compromise cellular functioning and contribute to tissue injury. Herein we determined if form selective CYP inhibitors could distinguish between the activities of the monooxygenase and NADPH oxidase activities of rat recombinant CYP1A2, CYP2E1, CYP3A1 and CYP3A2 and CYP1A1/2-enriched β-naphthoflavone-induced rat liver microsomes, CYP2E1-enriched isoniazide-induced rat liver microsomes and CYP3A subfamily-enriched dexamethasone-induced rat liver microsomes. In the presence of 7,8-benzoflavone (2.0 μM) for CYP1A2 and 4-methylpyrazole (32 μM) or DMSO (16 mM) for CYP2E1, monooxygenase activity was blocked without affecting NADPH oxidase activity for both the recombinant enzymes and microsomal preparations. Ketoconazole (1.0 μM), a form selective inhibitor for CYP3A subfamily enzymes, completely inhibited monooxygenase activity of rat recombinant CYP3A1/3A2 and CYP3A subfamily in rat liver microsomes; it also partially inhibited NADPH oxidase activity. 7,8-benzoflavone is a type I ligand, which competes with substrate binding, while 4-methylpyrazole and DMSO are type II heme binding ligands. Interactions of heme with these type II ligands was not sufficient to interfere with oxygen activation, which is required for NADPH oxidase activity. Ketoconazole, a type II ligand known to bind multiple sites on CYP3A subfamily enzymes in close proximity to heme, also interfered, at least in part, with oxygen activation. These data indicate that form specific inhibitors can be used to distinguish between monooxygenase reactions and H 2 O 2 generating NADPH oxidase of CYP1A2 and CYP2E1. Mechanisms by which ketoconazole inhibits CYP3A NADPH oxidase remain to be determined., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Chitooligosaccharides (COS) with two different molecular weights are acylated with four nonpolar amino acids: glycine (Gly), alanine (Ala), valine (Val) and leucine (Leu) to obtain 2-aminoacetyl-chitooligosaccharide (2-GlyCOS), 2-aminopropionyl-chitooligosaccharide (2-AlaCOS), 2-amino-3-methylbutyryl-chitooligosaccharide (2-ValCOS), and 2-amino-4-methylpentanoyl-chitooligosaccharide (2-LeuCOS). The structure of the derivatives was characterized by FT-IR spectroscopy, 13 C NMR spectroscopy, and elemental analysis. The antioxidant activities of the derivatives, such as hydroxyl radical (·OH) scavenging ability, superoxide anion (O 2 · - ) scavenging ability, reducing ability, and DPPH radical scavenging ability, were investigated using various established systems. Compared with chitooligosaccharide and nonpolar amino acids, all derivatives have strong scavenging ability toward hydroxyl radicals and superoxide anions, and the clearance rate was 19.05% and 67.70% separately. The reducing ability and DPPH free radical scavenging ability of the derivatives are only 0.021Abs and 32.97%. Among them, only 2-AlaLCOS has significant reducing ability, and the value can reach 0.143Abs. The above results showed that the antioxidant activity of some derivatives was higher than that of chitooligosaccharide. The water solubility of the new derivatives was also greatly improved compared to that of nonpolar amino acids. Therefore, the application of 2-aminoacyl-chitooligosaccharides (2-AACOS) in antioxidants has laid a foundation and has certain potential application value in the fields of medicine, agriculture, and animal husbandry., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

This study's objective was to investigate the effects of dietary Se (in the form of selenomethionine) on the antioxidant activity and selenoprotein gene expressions in layer breeder roosters. One hundred and eighty, 36-wk-old Jingfen layer breeder roosters were randomly allocated to one of 5 dietary treatments (0, 0.25, 0.5, 1, or 2 mg/kg Se) for 6 wk on a corn-soybean meal-based diet. Antioxidant parameters and selenoprotein gene expressions were assessed at the end of the experiment. The results showed that Se supplementation significantly increased the activity of T-SOD, CAT, GSH-Px, and superoxide anion scavenging ability in plasma (P ≤ 0.05), and activities of T-SOD, CAT, GSH-Px, superoxide anion scavenging ability, and hydroxyl radical scavenging ability in the liver, kidney, and testis (P < 0.05). Moreover, MDA levels were significantly reduced in plasma, liver, kidney, and testis (P < 0.01), compared to the control group. Furthermore, the dietary administration of Se significantly increased TrxR2 and GPx4 mRNA levels in kidney and testis, and ID1 mRNA levels in liver and kidney. Most of the antioxidant parameters and selenoprotein-related gene expressions significantly increased, and MDA significantly decreased at dietary supplementation with 0.5 mg/kg Se. Whereas a higher dose of Se level (1 or 2 mg/kg) inhibited the activities of some of the antioxidant enzymes and selenoprotein-related gene expressions in selected tissues. In conclusion, dietary Se supplementation with 0.5 mg/kg significantly improved roosters' antioxidant status and selenoprotein-related gene expression in liver, kidney, and testis, while higher doses led to inhibit these; dietary Se might increase reproductive performance by enhancing their antioxidant status in roosters., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

The present study evaluated the priming efficacy of chitosan and chitosan-derived nanoparticles (CNPs) against bacterial wilt of tomato. In the current study, seed-treated CNPs plus pathogen-inoculated tomato seedlings recorded significant protection of 62 % against pathogen-induced wilt disease and subsequently better growth. The induced resistance was witnessed by a prominent increase in lignin, callose and H 2 O 2 deposition, followed by superoxide radical accumulation in leaves. Additionally, chitosan and CNPs-treated tomato plants recorded a remarkable increase in the upregulation of phenylalanine ammonia-lyase (PAL), peroxidase (POX), polyphenol oxidase (PPO), catalase (CAT) and β-1, 3 glucanase (GLU) in comparison with untreated plants. The chitosan and CNPs-induced antioxidant enzymes were positively correlated with the stimulation of corresponding gene expression in CNPs treated plants related to pathogen-inoculated ones. The results of this study describe that how the application of chitosan and CNPs elicit defense responses at the cellular, biochemical and gene expression in tomato plants against bacterial wilt disease, thereby improve growth and yield., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Sugars are essential metabolites for energy and anabolism that can also act as signals to regulate plant physiology and development. Experimental tools to disrupt major sugar signalling pathways are limited. We performed a chemical screen for modifiers of activation of circadian gene expression by sugars to discover pharmacological tools to investigate and manipulate plant sugar signalling. Using a library of commercially available bioactive compounds, we identified 75 confident hits that modified the response of a circadian luciferase reporter to sucrose in dark-adapted Arabidopsis thaliana seedlings. We validated the transcriptional effect on a subset of the hits and measured their effects on a range of sugar-dependent phenotypes for 13 of these chemicals. Chemicals were identified that appear to influence known and unknown sugar signalling pathways. Pentamidine isethionate was identified as a modifier of a sugar-activated Ca 2+ signal that acts as a calmodulin inhibitor downstream of superoxide in a metabolic signalling pathway affecting circadian rhythms, primary metabolism and plant growth. Our data provide a resource of new experimental tools to manipulate plant sugar signalling and identify novel components of these pathways., (© 2022 The Authors. New Phytologist © 2022 New Phytologist Foundation.)

Abstract: The transient receptor potential ion channel TRPM3 is highly prevalent on nociceptive dorsal root ganglion (DRG) neurons, but its functions in neuronal plasticity of chronic pain remain obscure. In an animal model of nonspecific low back pain (LBP), latent spinal sensitization known as nociceptive priming is induced by nerve growth factor (NGF) injection. Here, we address the TRPM3-associated molecular basis of NGF-induced latent spinal sensitization at presynaptic level by studying TRPM3-mediated calcium transients in DRG neurons. By investigating TRPM3-expressing HEK cells, we further show the dynamic mitochondrial activity downstream of TRPM3 activation. NGF enhances TRPM3 function, attenuates TRPM3 tachyphylaxis, and slows intracellular calcium clearance; TRPM3 activation triggers more mitochondrial calcium loading than depolarization does, causing a steady-state mitochondrial calcium elevation and a delayed recovery of cytosolic calcium; mitochondrial calcium buffering accounts for approximately 40% of calcium influx subsequent to TRPM3 activation. TRPM3 activation provokes an outbreak of pulsatile superoxide production (mitoflash) that comes in the form of a surge in frequency being tunable. We suggest that mitoflash pulsations downstream of TRPM3 activation might be an early signaling event initiating pain sensitization. Tuning of mitoflash activity would be a novel bottom-up therapeutic strategy for chronic pain conditions such as LBP and beyond., (Copyright © 2022 International Association for the Study of Pain.)