Decreased Reactive Oxygen Species Generation Research Articles

Functional Characterization of a Sugar Beet BvbHLH93 Transcription Factor in Salt Stress Tolerance.

The basic/helix–loop–helix (bHLH) transcription factor (TF) plays an important role for plant growth, development, and stress responses. Previously, proteomics of NaCl treated sugar beet leaves revealed that a bHLH TF, BvbHLH93, was significantly increased under salt stress. The BvbHLH93 protein localized in the nucleus and exhibited activation activity. The expression of BvbHLH93 was significantly up-regulated in roots and leaves by salt stress, and the highest expression level in roots and leaves was 24 and 48 h after salt stress, respectively. Furthermore, constitutive expression of BvbHLH93 conferred enhanced salt tolerance in Arabidopsis, as indicated by longer roots and higher content of chlorophyll than wild type. Additionally, the ectopic expression lines accumulated less Na+ and MDA, but more K+ than the WT. Overexpression of the BvBHLH93 enhanced the activities of antioxidant enzymes by positively regulating the expression of antioxidant genes SOD and POD. Compared to WT, the overexpression plants also had low expression levels of RbohD and RbohF, which are involved in reactive oxygen species (ROS) production. These results suggest that BvbHLH93 plays a key role in enhancing salt stress tolerance by enhancing antioxidant enzymes and decreasing ROS generation.

Blockade of mTORC1-NOX signaling pathway inhibits TGF-β1-mediated senescence-like structural alterations of the retinal pigment epithelium.

The retinal pigment epithelium (RPE) undergoes characteristic structural changes and epithelial-mesenchymal transition (EMT) during normal aging, which are exacerbated in age-related macular degeneration (AMD). Although the pathogenic mechanisms of aging and AMD remain unclear, transforming growth factor-β1 (TGF-β1) is known to induce oxidative stress, morphometric changes, and EMT as a senescence-promoting factor. In this study, we examined whether intravitreal injection of TGF-β1 into the mouse eye elicits senescence-like morphological alterations in the RPE and if this can be prevented by suppressing mammalian target of rapamycin complex 1 (mTORC1) or NADPH oxidase (NOX) signaling. We verified that intravitreal TGF-β1-induced stress fiber formation and EMT in RPE cells, along with age-associated morphometric changes, including increased variation in cell size and reduced cell density. In RPE cells, exogenous TGF-β1 increased endogenous expression of TGF-β1 and upregulated Smad3-ERK1/2-mTORC1 signaling, increasing reactive oxygen species (ROS) production and EMT. We demonstrated that inhibition of the mTORC1-NOX4 pathway by pretreatment with 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR), an activator of AMP-dependent protein kinase, or GKT137831, a NOX1/4 inhibitor, decreased ROS generation, prevented stress fiber formation, attenuated EMT, and improved the regularity of the RPE structure in vitro and in vivo. These results suggest that intravitreal TGF-β1 injection could be used as a screening model to investigate the aging-related structural and functional changes to the RPE. Furthermore, the regulation of TGF-β-mTORC1-NOX signaling could be a potential therapeutic target for reducing pathogenic alterations in aged RPE and AMD.

Lack of Autophagy Induction by Lithium Decreases Neuroprotective Effects in the Striatum of Aged Rats

The pharmacological modulation of autophagy is considered a promising neuroprotective strategy. While it has been postulated that lithium regulates this cellular process, the age-related effects have not been fully elucidated. Here, we evaluated lithium-mediated neuroprotective effects in young and aged striatum. After determining the optimal experimental conditions for inducing autophagy in loco with lithium carbonate (Li2CO3), we measured cell viability, reactive oxygen species (ROS) generation and oxygen consumption with rat brain striatal slices from young and aged animals. In the young striatum, Li2CO3 increased tissue viability and decreased ROS generation. These positive effects were accompanied by enhanced levels of LC3-II, LAMP 1, Ambra 1 and Beclin-1 expression. In the aged striatum, Li2CO3 reduced the autophagic flux and increased the basal oxygen consumption rate. Ultrastructural changes in the striatum of aged rats that consumed Li2CO3 for 30 days included electrondense mitochondria with disarranged cristae and reduced normal mitochondria and lysosomes area. Our data show that the striatum from younger animals benefits from lithium-mediated neuroprotection, while the striatum of older rats does not. These findings should be considered when developing neuroprotective strategies involving the induction of autophagy in aging.

Protective effects of luteolin on the venous endothelium.

Luteolin is a flavonoid with antioxidant properties already demonstrated in studies related to inflammation, tumor, and cardiovascular processes; however, there are no available information regarding its antioxidant effects at the venous endothelial site. We investigated the effects of luteolin (10, 20, and 50μmol/L) in cultures of rat venous endothelial cells. Nitric oxide (NO) and reactive oxygen species (ROS) were analyzed by fluorimetry; 3-nitrotyrosine (3-NT) residues were evaluated by immunofluorescence, and prostacyclin (PGI2) release was investigated by colorimetry. Intracellular NO levels were significantly enhanced after 10min of luteolin incubation, with a parallel decrease in ROS generation. These results were accompanied by a significant reduction in the expression of 3-NT residues and enhanced PGI2 rates. Therefore, luteolin is effective in reducing ROS thereby improving NO availability in venous endothelial cells. Besides, luteolin-induced decrease in 3-NT residues may correlate with the enhancement in endothelial PGI2 bioavailability. These findings suggest the future application of this flavonoid as a protective agent by improving endothelial function in several circulatory disorders related to venous insufficiency.

Role of the Neutral Amino Acid Transporter SLC7A10 in Adipocyte Lipid Storage, Obesity, and Insulin Resistance.

Elucidation of mechanisms that govern lipid storage, oxidative stress, and insulin resistance may lead to improved therapeutic options for type 2 diabetes and other obesity-related diseases. Here, we find that adipose expression of the small neutral amino acid transporter SLC7A10, also known as alanine-serine-cysteine transporter-1 (ASC-1), shows strong inverse correlates with visceral adiposity, insulin resistance, and adipocyte hypertrophy across multiple cohorts. Concordantly, loss of Slc7a10 function in zebrafish in vivo accelerates diet-induced body weight gain and adipocyte enlargement. Mechanistically, SLC7A10 inhibition in human and murine adipocytes decreases adipocyte serine uptake and total glutathione levels and promotes reactive oxygen species (ROS) generation. Conversely, SLC7A10 overexpression decreases ROS generation and increases mitochondrial respiratory capacity. RNA sequencing revealed consistent changes in gene expression between human adipocytes and zebrafish visceral adipose tissue following loss of SLC7A10, e.g., upregulation of SCD (lipid storage) and downregulation of CPT1A (lipid oxidation). Interestingly, ROS scavenger reduced lipid accumulation and attenuated the lipid-storing effect of SLC7A10 inhibition. These data uncover adipocyte SLC7A10 as a novel important regulator of adipocyte resilience to nutrient and oxidative stress, in part by enhancing glutathione levels and mitochondrial respiration, conducive to decreased ROS generation, lipid accumulation, adipocyte hypertrophy, insulin resistance, and type 2 diabetes.

Uric acid released from poly(ε-caprolactone) fibers as a treatment platform for spinal cord injury.

Spinal cord injury (SCI) is characterized by a primary mechanical phase of injury, resulting in physical tissue damage, and a secondary pathological phase, characterized by biochemical processes contributing to inflammation, neuronal death, and axonal demyelination. Glutamate-induced excitotoxicity (GIE), in which excess glutamate is released into synapses and overstimulates glutamate receptors, is a major event in secondary SCI. GIE leads to mitochondrial damage and dysfunction, release of reactive oxygen species (ROS), DNA damage, and cell death. There is no clinical treatment that targets GIE after SCI, and there is a need for therapeutic targets for secondary damage in patients. Uric acid (UA) acts as an antioxidant and scavenges free radicals, upregulates glutamate transporters on astrocytes, and preserves neuronal viability in in vitro and in vivo SCI models, making it a promising therapeutic candidate. However, development of a drug release platform that delivers UA locally to the injured region in a controlled manner is crucial, as high systemic UA concentrations can be detrimental. Here, we used the electrospinning technique to synthesize UA-containing poly(ɛ-caprolactone) fiber mats that are biodegradable, biocompatible, and have a tunable degradation rate. We optimized delivery of UA as a burst within 20 min from uncoated fibers and sustained release over 2 h with poly(ethylene glycol) diacrylate coating. We found that both of these fibers protected neurons and decreased ROS generation from GIE in organotypic spinal cord slice culture. Thus, fiber mats represent a promising therapeutic for UA release to treat patients who have suffered a SCI.

Exogenous sodium hydrosulfide protects against high glucose‑induced injury and inflammation inhuman umbilical vein endothelial cells by inhibiting necroptosis via the p38 MAPK signaling pathway.

In recent years hydrogen sulfide (H2S) has demonstrated vasculoprotective effects against cell death, which suggests its promising therapeutic potential for numerous types of disease. Additionally, a protective effect of exogenous H2S in HG-induced injuries in HUVECs was demonstrated, suggesting a potential protective effect for diabetic vascular complications. The present study aimed to investigate the mechanism accounting for the cytoprotective role of exogenous H2S against high glucose [HG (40 mM glucose)]-induced injury and inflammation in human umbilical vein endothelial cells (HUVECs). HUVECs were exposed to HG for 24 h to establish an in vitro model of HG-induced cytotoxicity. The cells were pretreated with sodium hydrosulfide (NaHS), a donor of H2S, or inhibitors of necroptosis and p38 MAPK prior to the exposure to HG. Cell viability, intracellular reactive oxygen species (ROS), mitochondrial membrane potential (MMP), IL-1β, IL-6, IL-8, TNF-α, phosphorylated-(p)38 and receptor-interacting protein 3 (RIP3) expression levels were detected using the indicated methods, including Cell Counting Kit 8, fluorescence detection, western blotting, immunofluorescence assay and ELISAs. The results demonstrated that necroptosis and the p38 MAPK signaling pathway mediated HG-induced injury and inflammation. Notably, NaHS was discovered to significantly ameliorate p38 MAPK/necroptosis-mediated injury and inflammation in response to HG, as evidenced by an increase in cell viability, a decrease in ROS generation and loss of MMP, as well as the reduction in the secretion of proinflammatory cytokines. In addition, the upregulated expression of RIP3 induced by HG was repressed by treatment with SB203580, while the HG-induced upregulation of p-p38 expression levels were significantly downregulated following the treatment of Nec-1 and RIP3-siRNA. In conclusion, the findings of the present study indicated that NaHS may protect HUVECs against HG-induced injury and inflammation by inhibiting necroptosis via the p38 MAPK signaling pathway, which may represent a promising drug for the therapy of diabetic vascular complications.

Therapeutic Potential of Polyphenols-Loaded Polymeric Nanoparticles in Cardiovascular System.

Numerous studies document an increased production of reactive oxygen species (ROS) with a subsequent decrease in nitric oxide (NO) bioavailability in different cardiovascular diseases, including hypertension, atherosclerosis, and heart failure. Many natural polyphenols have been demonstrated to decrease ROS generation and/or to induce the endogenous antioxidant enzymatic defense system. Moreover, different polyphenolic compounds have the ability to increase the activity/expression of endothelial nitric oxide synthase (eNOS) with a subsequent enhancement of NO generation. However, as a result of low absorption and bioavailability of natural polyphenols, the beneficial effects of these substances are very limited. Recent progress in delivering polyphenols to the targeted tissues revealed new possibilities for the use of polymeric nanoparticles in increasing the efficiency and reducing the degradability of natural polyphenols. This review focuses on the effects of different natural polyphenolic substances, especially resveratrol, quercetin, curcumin, and cherry extracts, and their ability to bind to polymeric nanoparticles, and summarizes the effects of polyphenol-loaded nanoparticles, mainly in the cardiovascular system.

Assessment of the photoprotective potential and toxicity of Antarctic red macroalgae extracts from Curdiea racovitzae and Iridaea cordata for cosmetic use

Botanical and marine extracts have been used in cosmetic products as natural antioxidants or UV-absorbers to reduce the use of organic UV filters. However, despite the high UV exposure of Antarctic macroalgae that induces the production of photoprotective compounds, the use of marine macroalgae extracts for photoprotection has been little explored. Thus our objective was to assess the photoprotective potential and the toxicity of extracts from two red macroalgae Curdiea racovitzae and Iridaea cordata. Chemical characterization was based on the qualitative and quantitative analyses of mycosporine-like amino acids (MAAs) by high performance liquid chromatography coupled to mass spectrometry. Photoprotective properties were assessed by UV spectrophotometry and photostability assay. Antioxidant capacity was studied by DPPH and superoxide radical scavenging assays. UVB and UVA photoprotection was assessed in keratinocytes by cell viability analysis after UVB irradiation and quantification of intracellular reactive oxygen species (ROS) induced by UVA, respectively. Toxicity was estimated by cytotoxicity, phototoxicity and HET-CAM (Hen's egg test-chorioallantoic membrane) assays. Red algae extracts had high concentrations of MAAs, mainly consisted of palythine. The extracts absorbed in the UVB and UVA-II and were photostable. The C. racovitzae extract showed antioxidant activity against superoxide radicals. In addition, the C. racovitzae (1.0 and 0.5 mg/mL) and I. cordata (1.0 mg/mL) extracts had an UVB photoprotection similar to benzophenone-3, which was dependent on the total MAAs concentration. Besides, the C. racovitzae extract decreased ROS generation by 31 and 24% at 100 and 50 μg/mL, respectively. Furthermore, it was more effective in reducing ROS generation than quercetin and Helioguard 365®. Finally, both extracts were considered to be non-cytotoxic, non-phototoxic and non-irritant. Therefore, the C. racovitzae extract is a good candidate to be added to anti-aging and sunscreen formulations in order to protect against oxidative stress and to improve the absorption spectra of UV filters.

Effect of Glucose Metabolism-Associated Protein-1 on Exendin-4-Treated Pancreatic Islet Cell Function

Exendin-4 regulates blood sugar. Glucose metabolism-associated protein-1 (GMRP-1) regulates islet cell function. Therefore, this paper intends to analyze the effect of GMRP-1 on Exendin-4treated islet cells. The islet cell MIN6 cells were separated into control group, Exendin-4 group, GMRP-1 overexpressing+ Exendin-4 group, and GMRP-1 siRNA+ Exendin-4 group followed by analysis of the expression of GMRP-1 by quantitative real-time PCR and Western blot, cell proliferation by MTT, cell apoptosis by flow cytometry, Bcl-2, Bax, Fas and Fas-L expression by real-time PCR, as well as caspase-3 activity, reactive oxygen species (ROS) and superoxide dismutase (SOD) activity. Exendin-4 and GMRP-1 overexpression+ Exendin-4 significantly increased GMRP-1 expression and promoted cell proliferation, decreased apoptotic rate and Caspase-3 activity, increased Bcl-2 and Fas-L expression, reduced Fas and Bax expression, as well as decreased ROS generation and increased SOD activity (P < 0 05), and the changes in GMRP-1 overexpression+ Exendin-4 group was more significant (P < 0 01). However, GMRP-1 siRNA+ Exendin-4 group significantly down-regulated GMRP-1 expression, decreased MIN6 cell prolifer- ation, increased cell apoptosis and Caspase-3 activity, decreased Bcl-2 and Fas-L expression, elevated Fas and Bax expression, increased ROS content and decreased SOD activity compared to control group and Exendin-4 group (P < 0 05). Up-regulation of GMRP-1 expression promoted Exendin-4-induced proliferation and inhibited apoptosis of islet cells. Down-regulation of GMRP-1 expression reversed Exendin-4's effect on islet cells.

Upregulation Sestrin2 protects against hydrogen peroxide-induced oxidative damage bovine mammary epithelial cells via a Keap1-Nrf2/ARE pathway.

Sestrin2 (SESN2) is a highly conservative oxidative stress protein that can regulate energy metabolism, cell proliferation, apoptosis, and mitochondria autophagy processes. It plays a role as an antioxidant in various diseases. The aims of the present study were to explore the underlying role of SESN2 after hydrogen peroxide (H2 O2 ) treatment in bovine mammary epithelial cells (MAC-T cells) by the methods of knockout or overexpression of SESN2. The results show that knockout of Sestrin2 exacerbate apoptosis, upregulatethe expressions of Bax/Bcl2 in H2 O2 -treated MAC-T cells. Moreover, knockout of SESN2 also promoted reactive oxygen species (ROS) generation and exacerbated oxidative damage in H2 O2 -treated MAC-T cells. On the contrary, overexpression of SESN2 decreased apoptosis by downregulation of Bax/Bcl2 level decreased ROS generation and blocked oxidative damage in H2 O2 -treated MAC-T cells. In addition, results indicate that the Kelch-like ECH-associated protein-1 (Keap1)-nuclear factor (erythroid-derived 2) like2 (Nrf2)/antioxidant response element (ARE) signaling pathway was activated by H2 O2 ; upregulation of SESN2 could relieve oxidative stress by inducing the expression of Keap1, Nrf2, HO-1, and NDPH: quinone oxidoreductase-1 protein. In conclusion, this study demonstrates that expression of SESN2 was significantly increased after H2 O2 treatment and that SESN2 can alleviate oxidative stress and cell apoptosis in H2 O2 -treated MAC-T cells through activation of the Keap1-Nrf2/ARE pathway.

Mitochondrial APE1 promotes cisplatin resistance by downregulating ROS in osteosarcoma.

Apurinic/apyrimidinic endonuclease 1 (APE1) is a primary nuclear-localized multifunctional protein in osteosarcoma. However, the cytoplasmic localization of APE1 was found to be functional and to increase with cisplatin resistance, yet the molecular mechanism is unknown. In the present study, we explored the cisplatin resistance mechanism in osteosarcoma from the new perspective of APE1 extranuclear biological activity. Using cisplatin-resistant and cisplatin-sensitive osteosarcoma cell lines, we found that mitochondrial APE1 (mtAPE1) was overexpressed in cisplatin-resistant cells but not in sensitive cells. Overexpression of mtAPE1 reduced cisplatin-induced apoptosis, while knockdown of APE1 reversed this phenomenon and caused oxidative DNA damage via overproduction of reactive oxygen species (ROS). We further demonstrated that high mtAPE1 expression could downregulate ROS production by decreasing the phosphorylation of Rac1 (p-Rac1), further promoting cisplatin resistance in osteosarcoma. Our findings suggest that mitochondrial APE1 promotes cisplatin resistance by decreasing ROS generation, which may provide new ideas for researching the molecular mechanism of osteosarcoma chemoresistance and strategies to overcome cisplatin resistance in osteosarcoma.

Fucoidan from acid-processed Hizikia fusiforme attenuates oxidative damage and regulate apoptosis

We had observed in our previous study that the active fucoidan (JHCF4), isolated from the crude fucoidan in acid-processed Hizikia fusiforme, possessed an anticancer effect. In this study, the antioxidant effect of JHCF4 was evaluated. Among the fractions, JHCF4 showed the highest scavenging activity against 2, 2-diphenyl-1-picrylhydrazyl (DPPH), alkyl, and hydroxyl radicals, as well as protective effect against reactive oxygen species (ROS) in 2, 2′-azobis (2-amidinopropane) dihydrochloride (AAPH)-treated Vero cells. Furthermore, JHCF4 showed a protective activity against AAPH-induced apoptosis, as observed by nuclear staining with Hoechst 33342. Our results showed that JHCF4 can up-regulate Bcl-xL, down-regulate Bax and cleave caspase-3 with increased concentrations in AAPH-induced Vero cells. JHCF4 induced anti-apoptosis via a mitochondria-mediated pathway. Additionally, JHCF4 was selected for further in vivo screening in a zebrafish model, which markedly decreased ROS generation and lipid peroxidation. Thus, JHCF4 showed a potential protective activity against AAPH-induced ROS both in vitro and in the zebrafish model.

The simultaneous removal of the combined pollutants of hexavalent chromium and o-nitrophenol by Chlamydomonas reinhardtii.

Microalgae have been used for the removal of heavy metals or synthetic organics; however, the simultaneous removal of both types of compounds is always technically difficult. In this study, a green algae, Chlamydomonas reinhardtii, was first used to simultaneously remove hexavalent chromium [Cr(VI)] and o-nitrophenol (ONP), and the balance among biomass, oxidative damage and removal rate was also investigated. The results showed that treatment with Cr(VI) or ONP decreased the photosynthetic and superoxide dismutase activities and increased the production of reactive oxygen species (ROS) and malondialdehyde content. However, combined treatment with Cr(VI) (≤4mg/L) and ONP (≤15mg/L) significantly decreased ROS generation and alleviated cell damage in C. reinhardtii. In addition, the removal rates of Cr(VI) and ONP by C. reinhardtii cells significantly increased from 37.4% to 54.9% and from 35.8% to 45.9%, respectively, and the cells could be reused at least four times. Moreover, the increased acidity in the medium and Cr(VI) reductase content in C. reinhardtii caused Cr(VI) to be reduced to Cr(III). The addition of an exogenous antioxidant decreased the removal rates of Cr(VI) and ONP. These results indicated that the presence of Cr(VI) could induce ROS generation in C. reinhardtii and enhance ONP degradation, which consumed ROS, alleviated cell damage, and thus benefited Cr(VI) reduction. As a result, C. reinhardtii could be used as a theoretical candidate for the simultaneous removal of combined Cr(VI) and ONP contamination.

Antioxidant potential of non-oil seed legumes of Indonesian’s ethnobotanical extracts

This study investigated the in vitro antioxidant properties (DPPH, ABTS, CUPRAC and FRAP), total phenolic content and flavonoid content of extracts from three non-oil seed legumes (Phaseolus lunatus red and white, and Canavalia ensiformis), local edible seeds from Indonesia, obtained using different solvent system (distilled water, 70% ethanol, and 100% ethanol). The variety of legume was a major source of variation in the phenolic contents, flavonoid content and antioxidant activity. HPLC analysis of the non-oil seed legume extracts identified gallic acid, epicatechin and coumaric acid. Among the varieties of non-oil seed legume extracts, the phenolic content varied from 15.21–38.60 mg gallic acid equivalents/g dry weight and the flavonoid content was 11.73–24.61 mg catechin equivalents/g dry weight. The antioxidant activity of the extracts suppressed the reactive oxygen species (ROS) generation and cellular damage induced by UV-B in HaCaT cells. These results showed that antioxidant activity (1.83–19.42% of inhibition DPPH; 2.99–37.29% of inhibition ABTS; 0.20–2.47 µM CUPRAC value; and 0.96–1.10 µM of FRAP value) of extracts possessed strong radical scavenging activity as well as inhibited ROS generation in a dose-dependent manner without showing any cytotoxicity. Collectively, the data presented that antioxidant of the extracts have potent antioxidant activity and decreasing ROS generation in HaCaT cells. It can be intimately used as alternative criterion for antioxidant and antiradical activities that can be utilized as a functional food and nutraceutical ingredients.

Anilinopyrazines as potential mitochondrial uncouplers

Mitochondrial protonophores transport protons through the mitochondrial inner membrane into the matrix to uncouple nutrient oxidation from ATP production thereby decreasing the proton motive force. Mitochondrial uncouplers have beneficial effects of decrease reactive oxygen species generation and have the potential for treating diseases such as obesity, neurodegenerative diseases, non-alcoholic fatty liver disease (NAFLD), diabetes, and many others. In this study, we report the structure-activity relationship profile of the pyrazine scaffold bearing substituted aniline rings. Our work indicates that a trifluoromethyl group is best at the para position while the trifluoromethoxy group is preferred in the meta position of the aniline rings of 2,3-substituted pyrazines. As proton transport and cycling requires the formation of a negative charge that has to traverse the mitochondrial membrane, a stabilizing internal hydrogen bond is a key feature for efficient mitochondrial uncoupling activity.

Fisetin protects against cardiac cell death through reduction of ROS production and caspases activity

Myocardial infarction (MI) is a leading cause of death worldwide. Reperfusion is considered as an optimal therapy following cardiac ischemia. However, the promotion of a rapid elevation of O2 levels in ischemic cells produces high amounts of reactive oxygen species (ROS) leading to myocardial tissue injury. This phenomenon is called ischemia reperfusion injury (IRI). We aimed at identifying new and effective compounds to treat MI and minimize IRI. We previously studied heart regeneration following myocardial injury in zebrafish and described each step of the regeneration process, from the day of injury until complete recovery, in terms of transcriptional responses. Here, we mined the data and performed a deep in silico analysis to identify drugs highly likely to induce cardiac regeneration. Fisetin was identified as the top candidate. We validated its effects in an in vitro model of MI/IRI in mammalian cardiac cells. Fisetin enhances viability of rat cardiomyocytes following hypoxia/starvation – reoxygenation. It inhibits apoptosis, decreases ROS generation and caspase activation and protects from DNA damage. Interestingly, fisetin also activates genes involved in cell proliferation. Fisetin is thus a highly promising candidate drug with clinical potential to protect from ischemic damage following MI and to overcome IRI.

SFPQ is involved in regulating arsenic-induced oxidative stress by interacting with the miRNA-induced silencing complexes

Arsenic exposure contributed to the development of human diseases. Arsenic exerted multiple organ toxicities mainly by triggering oxidative stress. However, the signaling pathway underlying oxidative stress is unclear. We previously found that the expression of SFPQ, a splicing factor, was positively associated with urinary arsenic concentration in an arsenic-exposed population, suggesting an oxidative stress regulatory role for SFPQ. To test this hypothesis, we established cell models of oxidative stress in human hepatocyte cells (L02) treated with NaAsO2. Reactive oxygen species (ROS) synthesis displayed a time- and dose-dependent increase with NaAsO2 treatment. SFPQ suppression resulted in a 36%–53% decrease in ROS generation, leading to enhanced cellular damage determined by 8-OHdG, comet tail moment, and micronucleus analysis. Particularly, SFPQ deficiency attenuated expression of the oxidase genes DUOX1, DUOX2, NCF2, and NOX2. A fluorescent-based RNA electrophoretic mobility shift assay (FREMSA) and dual-luciferase reporter system revealed that miR-92b-5p targeted DUOX2 mRNA degradation. An RNA immunoprecipitation assay showed an interaction between SFPQ and miR-92b-5p of the miRNA-induced silencing complex (miRISC). Notably, NaAsO2 treatment diminished the interaction between SFPQ and miR92b-5p, accompanied by decreased binding between miR-92b-5p and 3′-UTR of DUOX2. However, SFPQ deficiency suppressed the dissociation of miR-92b-5p from 3′-UTR of DUOX2, indicating that miR-92b-5p regulated the SFPQ-dependent DUOX2 expression. Taken together, we reveal that SFPQ responds to arsenic-induced oxidative stress by interacting with the miRISC. These findings offer new insight into the potential role of SFPQ in regulating cellular stress response.

Nobiletin Regulates ROS/ADMA/DDAHII/eNOS/NO Pathway and Alleviates Vascular Endothelium Injury by Iron Overload.

Iron overload is harmful to health and associates with intracellular excessive reactive oxygen species (ROS) generation. Nobiletin (Nob) is known to be antioxidant and anti-inflammatory. However, whether Nob can protect endothelial cells against iron overload has not been studied, and the specific mechanism has not yet been elucidated. In this study, we have identified the protective effects of Nob, and its underlying molecular mechanism in human umbilical vein endothelial cells (HUVECs) suffered from iron overload via ROS/ADMA/DDAHII/eNOS/NO pathway. We found that compared with 50μM iron dextran treatment, co-treatment with 20μM Nob increased cell viability and decreased lactate dehydrogenase activity. Besides, Nob could upregulate DDAHII expression and activity, promote eNOS phosphorylation to produce more NO, reduce ADMA content, and therefore increase superoxide dismutase, catalase, and glutathione peroxidase activities, and decrease malondialdehyde level and ROS generation. Nob also inhibited mitochondrial permeability transition pore (mPTP) openness and cleaved caspase-3 expression, and decreased apoptosis induced by iron overload. These results were consistent when Nob was replaced by the positive control reagents L-arginine (a competitive substrate of ADMA), cyclosporin A (an mPTP closing agent), or edaravone (a free radical scavenger). The addition of pAD/DDAHII-shRNA adenovirus reversed the above effects of Nob. These data suggested that the protective mechanism of Nob was to inhibit ROS burst, upregulate DDAHII expression and activity, promote eNOS phosphorylation, produce NO, reduce ADMA content, and ultimately alleviate iron overload damage in vascular endothelium.

Cordycepin attenuates Salivary Hypofunction through the Prevention of Oxidative Stress in Human Submandibular Gland Cells.

Xerostomia (dry mouth) is a significant age-related condition. Meanwhile, cordycepin, the natural therapeutic agent, has demonstrated an anti-aging effect. Therefore, the present study aimed to investigate the preventive effects of cordycepin on secretory function in an in vitro model of hydrogen peroxide (H2O2)-induced salivary hypofunction. After being exposed to H2O2, human submandibular gland (HSG) cells were treated with various concentrations of cordycepin (6.25-50 µM) for 24, 48, and 72h. To evaluate cell proliferation and reactive oxygen species (ROS) generation, 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide and 2, 7'-dichlorodihydrofluorescein diacetate assays were performed. The amylase activity was kinetically measured by 2-chloro-p-nitrophenol linked with maltotrioside. The expression of salivary, antioxidant and apoptotic markers at mRNA and protein levels were performed by reverse transcriptase polymerase chain reaction (RT-PCR) and immunofluorescence analysis, respectively. We demonstrated that cordycepin (6.25-25 µM) contributed to significant increases in expression of the salivary marker genes, alpha-amylase 1 (AMY1A) and aquaporin-5 (AQP5), and in amylase secretion without changes in cell viability. Under oxidative stress, HSG cells showed remarkable dysfunction. Cordycepin rescued the protective effects partially by decreasing ROS generation and restoring the expression of the salivary proteins, AMY and AQP5 via anti-oxidant and anti-apoptotic activity. In addition, the amount of amylase that was secreted from HSG cells cultured in cordycepin was increased. In conclusion, cordycepin demonstrated a protective effect on H2O2-induced HSG cells by decreasing ROS generation and upregulating the salivary function markers, AMY1A and AQP5, at both the transcriptional and translational levels.