Reduced Reactive Oxygen Species Generation Research Articles

Improved effect of a mitochondria-targeted antioxidant on hydrogen peroxide-induced oxidative stress in human retinal pigment epithelium cells

BackgroundOxidative damage to retinal pigment epithelial (RPE) cells contributes to the development of age-related macular degeneration, which is among the leading causes of visual loss in elderly people. In the present study, we evaluated the protective role of triphenylphosphonium (TPP)-Niacin against hydrogen peroxide (H2O2)-induced oxidative stress in RPE cells.MethodsThe cellular viability, lactate dehydrogenase release, reactive oxygen species (ROS) generation, and mitochondrial function of retinal ARPE-19 cells were determined under treatment with H2O2 or pre-treatment with TPP-Niacin. The expression level of mitochondrial related genes and some transcription factors were assessed using real-time polymerase chain reaction (RT-qPCR).ResultsTPP-Niacin significantly improved cell viability, reduced ROS generation, and increased the antioxidant enzymes in H2O2-treated ARPE-19 cells. Mitochondrial dysfunction from the H2O2-induced oxidative stress was also considerably diminished by TPP-Niacin treatment, along with reduction of the mitochondrial membrane potential (MMP) and upregulation of the mitochondrial-associated gene. In addition, TPP-Niacin markedly enhanced the expression of transcription factors (PGC-1α and NRF2) and antioxidant-associated genes (especially HO-1 and NQO-1).ConclusionWe verified the protective effect of TPP-Niacin against H2O2-induced oxidative stress in RPE cells. TPP-Niacin is believed to protect against mitochondrial dysfunction by upregulating antioxidant-related genes, such as PGC-1α, NRF2, HO-1, and NQO-1, in RPE cells.

Control of Mesenchymal Stromal Cell Senescence by Tryptophan Metabolites.

Cellular senescence contributes to aging and age-related disorders. High glucose (HG) induces mesenchymal stromal/stem cell (MSC) senescence, which hampers cell expansion and impairs MSC function. Intracellular HG triggers metabolic shift from aerobic glycolysis to oxidative phosphorylation, resulting in reactive oxygen species (ROS) overproduction. It causes mitochondrial dysfunction and morphological changes. Tryptophan metabolites such as 5-methoxytryptophan (5-MTP) and melatonin attenuate HG-induced MSC senescence by protecting mitochondrial integrity and function and reducing ROS generation. They upregulate the expression of antioxidant enzymes. Both metabolites inhibit stress-induced MSC senescence by blocking p38 MAPK signaling pathway, NF-κB, and p300 histone acetyltransferase activity. Furthermore, melatonin upregulates SIRT-1, which reduces NF-κB activity by de-acetylation of NF-κB subunits. Melatonin and 5-MTP are a new class of metabolites protecting MSCs against replicative and stress-induced cellular senescence. They provide new strategies to improve the efficiency of MSC-based therapy for diverse human diseases.

Antioxidative Effects of Chrysoeriol via Activation of the Nrf2 Signaling Pathway and Modulation of Mitochondrial Function.

Retinal pigment epithelium (RPE) cell dysfunction caused by excessive oxidative damage is partly involved in age-related macular degeneration, which is among the leading causes of visual impairment in elderly people. Here, we investigated the protective role of chrysoeriol against hydrogen peroxide (H2O2)-induced oxidative stress in RPE cells. The cellular viability, reactive oxygen species (ROS) generation, and mitochondrial function of retinal ARPE-19 cells were monitored under oxidative stress or pre-treatment with chrysoeriol. The expression levels of mitochondrial-related genes and associated transcription factors were assessed using reverse transcription–quantitative polymerase chain reaction (RT-qPCR). Moreover, the protein expression of antioxidant signal molecules was characterized by Western blot analysis. Chrysoeriol significantly increased cell viability, reduced ROS generation, and increased the occurrence of antioxidant molecules in H2O2-treated ARPE-19 cells. Additionally, mitochondrial dysfunction caused by H2O2-induced oxidative stress was also considerably diminished by chrysoeriol treatment, which reduced the mitochondrial membrane potential (MMP) and upregulated mitochondrial-associated genes and proteins. Chrysoeriol also markedly enhanced key transcription factors (Nrf2) and antioxidant-associated genes (particularly HO-1 and NQO-1). Therefore, our study confirms the protective effect of chrysoeriol against H2O2-induced oxidative stress in RPE cells, thus confirming that it may prevent mitochondrial dysfunction by upregulating antioxidant-related molecules.

Exercise-Linked Irisin Prevents Mortality and Enhances Cognition in a Mice Model of Cerebral Ischemia by Regulating Klotho Expression.

Irisin, which can be released in the hippocampus after physical exercise, is demonstrated to have beneficial effects on neurovascular diseases. This study investigated the impact of exercise linked-irisin on mortality and cognition in a mice model of cerebral ischemia and further explored its underlying mechanism. The cerebrospinal concentrations of irisin and klotho from ischemic stroke patients were measured with an enzyme-linked immunosorbent assay (ELISA). The cognitive function of mice was evaluated by a series of behavioural experiments. The expressions of klotho, MnSOD, and FOXO3a in the hippocampus of mice were detected by Western blot. Superoxide production in the brain tissue of mice was evaluated with the dihydroethidium (DHE) dying. The results demonstrated that stroke patients showed a positive correlation between their CSF irisin concentration and klotho concentration. In addition, when mice subjected to cerebral ischemia, their cognitive function was impaired, the protein expressions of klotho, MnSOD, and FOXO3a downregulated, and the production of reactive oxygen species (ROS) increased compared with the sham group. After pretreatment with exogenous irisin, improved cognitive impairment, upregulated protein expressions of klotho, MnSOD, and FOXO3a, and reduced ROS generation were observed in mice with MCAO. However, the neuroprotective effects of irisin compromised with the evidence of severe cognitive impairment, decreased protein expressions of MnSOD and FOXO3a, and increased ROS production in klotho knockout mice. Thus, our results indicated that exercise-linked irisin could prevent mortality and improve cognitive impairment after cerebral ischemia by regulating klotho expression.

Acetylshikonin Induces Apoptosis in Human Colorectal Cancer HCT-15 and LoVo Cells via Nuclear Translocation of FOXO3 and ROS Level Elevation.

Acetylshikonin, a naphthoquinone, is a pigment compound derived from Arnebia sp., which is known for its anti-inflammatory potential. However, its anticarcinogenic effect has not been well investigated. Thus, in this study, we focused on investigating its apoptotic effects against HCT-15 and LoVo cells, which are human colorectal cancer cells. MTT assay, cell counting assay, and colony formation assay have shown acetylshikonin treatment induced cytotoxic and antiproliferative effects against colorectal cancer cells in a dose- and time-dependent manner. DNA fragmentation was observed via terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay. Also, the increase of subG1 phase in cell cycle arrest assay and early/late apoptotic rates in annexin V/propidium iodide (PI) double staining assay was observed, which indicates an apoptotic potential of acetylshikonin against colorectal cancer cells. 2′,7′-Dichlorofluorescin diacetate (DCF-DA) staining was used to evaluate reactive oxygen species (ROS) generation in acetylshikonin-treated colorectal cancer cells. Fluorescence-activated cell sorting (FACS) analysis showed that acetylshikonin induced an increase in reactive oxygen species (ROS) levels and apoptotic rate in a dose- and time-dependent manner in HCT-15 and LoVo cells. In contrast, cotreatment with N-acetyl cysteine (NAC) has reduced ROS generation and antiproliferative effects in colorectal cancer cells. Western blotting analysis showed that acetylshikonin treatment induced increase of cleaved PARP, γH2AX, FOXO3, Bax, Bim, Bad, p21, p27, and active forms of caspase-3, caspase-7, caspase-9, caspase-6, and caspase-8 protein levels, while those of inactive forms were decreased. Also, the expressions of pAkt, Bcl-2, Bcl-xL, peroxiredoxin, and thioredoxin 1 were decreased. Furthermore, western blotting analysis of cytoplasmic and nuclear fractionated proteins showed that acetylshikonin treatment induced the nuclear translocation of FOXO3, which might result from DNA damage by the increased intracellular ROS level. This study represents apoptotic potential of acetylshikonin against colorectal cancer cells via translocation of FOXO3 to the nucleus and upregulation of ROS generation.

Rosuvastatin protects against coronary microembolization-induced cardiac injury via inhibiting NLRP3 inflammasome activation

Coronary microembolization (CME), a common reason for periprocedural myocardial infarction (PMI), bears very important prognostic implications. However, the molecular mechanisms related to CME remain largely elusive. Statins have been shown to prevent PMI, but the underlying mechanism has not been identified. Here, we examine whether the NLRP3 inflammasome contributes to CME-induced cardiac injury and investigate the effects of statin therapy on CME. In vivo study, mice with CME were treated with 40 mg/kg/d rosuvastatin (RVS) orally or a selective NLRP3 inflammasome inhibitor MCC950 intraperitoneally (20 mg/kg/d). Mice treated with MCC950 and RVS showed improved cardiac contractile function and morphological changes, diminished fibrosis and microinfarct size, and reduced serum lactate dehydrogenase (LDH) level. Mechanistically, RVS decreased the expression of NLRP3, caspase-1, interleukin-1β, and Gasdermin D N-terminal domains. Proteomics analysis revealed that RVS restored the energy metabolism and oxidative phosphorylation in CME. Furthermore, reduced reactive oxygen species (ROS) level and alleviated mitochondrial damage were observed in RVS-treated mice. In vitro study, RVS inhibited the activation of NLRP3 inflammasome induced by tumor necrosis factor α plus hypoxia in H9c2 cells. Meanwhile, the pyroptosis was also suppressed by RVS, indicated by the increased cell viability, decreased LDH and propidium iodide uptake in H9c2 cells. RVS also reduced the level of mitochondrial ROS generation in vitro. Our results indicate the NLRP3 inflammasome-dependent cardiac pyroptosis plays an important role in CME-induced cardiac injury and its inhibitor exerts cardioprotective effect following CME. We also uncover the anti-pyroptosis role of RVS in CME, which is associated with regulating mitochondrial ROS.

THAP9-AS1 Promotes Tumorigenesis and Reduces ROS Generation through the JAK2/STAT3 Signaling Pathway by Increasing SOCS3 Promoter Methylation in Osteosarcoma.

Increasing studies have demonstrated that dysfunction of long noncoding RNAs (lncRNAs) plays critical roles in the development of human cancers. THAP9-AS1 has been reported to be dysregulated and associated with tumor progression in some cancers. However, the function and mechanism of THAP9-AS1 in osteosarcoma (OS) remain unclear. In the present study, we found that the expression of THAP9-AS1 was significantly upregulated in OS tissues and associated with the advanced stage of tumors and poor prognosis of patients. Blast comparison results showed that the SOCS3 promoter region and THAP9-AS1 had base complementary pairing binding sites. The interactions between THAP9-AS1, DNA methyltransferases (DNMTs), and SOCS3 were assessed by RIP and ChIP assays. The results of methylation-specific PCR (MSP) and bisulfite sequencing PCR (BSP) validated that THAP9-AS1 enhanced the methylation level of the SOCS3 promoter. The mRNA levels of SOCS3 in OS cells could be reversed by the demethylation agent 5-aza-2′-deoxycytidine. The mRNA expression of SOCS3 was downregulated in OS tissues and negatively correlated with THAP9-AS1 expression in tumors. Moreover, the western blot and immunofluorescence (IF) assay data showed that THAP9-AS1 activated the JAK2/STAT3 signaling pathway by upregulating p-JAK2 and p-STAT3 and the nuclear translocation of p-STAT3. Functionally, ectopic expression of THAP9-AS1 promoted cell proliferation, migration, and invasion and inhibited apoptosis, and this phenomenon could be reversed by SOCS3. Introduction of the JAK/STAT inhibitor AG490 partially abolished the stimulative effect of THAP9-AS1 on cellular processes. In addition, THAP9-AS1 decreased oxidative stress by reducing reactive oxygen species (ROS) and enhancing the mitochondrial membrane potential of OS cells via the SOCS3/JAK2/STAT3 pathway. Stable overexpression of THAP9-AS1 contributed to tumor growth and metastasis in vivo. In total, our findings suggested that upregulation of THAP9-AS1 might recruit DNMTs to epigenetically inhibit SOCS3, thereby activating the JAK2/STAT3 signaling pathway and oncogenesis of OS. These results provide novel insights for the understanding of OS progression.

Ghrelin Ameliorates Diabetic Retinal Injury: Potential Therapeutic Avenues for Diabetic Retinopathy.

Ghrelin has anti-inflammatory, antioxidant, and antiapoptotic effects, and it may be beneficial for the treatment of many ophthalmic diseases, such as cataract, uveitis, and glaucoma. Our previous work proved that ghrelin pretreatment reduced the apoptosis of lens epithelial cells induced by hydrogen peroxide, reduced the accumulation of reactive oxygen species (ROS), and effectively maintained the transparency of lens tissue. However, no study has yet investigated the effect of ghrelin on retina. In this study, we conducted in vitro and in vivo experiments to explore the effect of ghrelin on high-glucose- (HG-) induced ARPE-19 cell damage and diabetic retinopathy in streptozotocin-induced diabetic rats. ARPE-19 cells were incubated in a normal or an HG (30 mM glucose) medium with or without ghrelin. Cell viability was measured by 3-(4, 5-dimethylthiazol-3-yl)-2,5-diphenyl tetrazolium bromide assay, and apoptosis was detected by the Hoechst–PI staining assay. Intracellular reactive oxygen species (ROS) production levels within cells were measured using 2′,7′-dichlorofluorescein diacetate staining, and the contents of superoxide dismutase and malondialdehyde were measured using relevant detection kits. The expression levels of IL-1β and IL-18 were measured using an enzyme-linked immunosorbent assay, and those of NLRP3, IL-1β, and IL-18 were measured using Western blotting. The rat diabetes models were induced using a single intraperitoneal injection of streptozotocin (80 mg/kg). The morphological and histopathological changes in the retinal tissues were examined. The results indicated that ghrelin reduced ROS generation, inhibited cell apoptosis and the activation of NLRP3 inflammasome, inhibited the apoptosis of retinal cells in diabetic rats, and protected the retina against HG-induced dysfunction. In conclusion, ghrelin may play a role in the treatment of ocular diseases involving diabetic retinopathy.

β-Lactams modulate neutrophil extracellular traps formation mediated by mTOR signaling pathway

BackgroundSome biotics, like β-Lactams, have shown immunomodulation effects during sepsis, but the detailed mechanism was still unclear. Here we postulated that neutrophils play an essential role and β-Lactams exert immunomodulation effects through modulating neutrophil extracellular traps (NETs) formation. MethodsNETs formation induced by two β-Lactams, Meropenem (MEM) and ceftazidime/tazobactam (CAZ/TB) in neutrophils from healthy donors and HL-60 cells was performed. Reactive oxygen species (ROS) generation and the activity of nicotinamide adenine dinucleotide phosphate (NAPDH) oxidase were examined. Additionally, the upstream signal pathway of NETs formation, including protein kinase C (PKC), protein kinase B (Akt) and mammalian target of rapamycin (mTOR), were detected. ResultsMEM and CAZ/TB modulate NETs formation in activated PMNs, not resting PMNs. Both reduced ROS generation in resting PMNs and increased in activated PMNs. To test the activity of NADPH oxidase, we detected NADPH in MEM and CAZ/TB pre-cultivated activated PMNs, which showed that MEM and CAZ/TB modulates NETs formation through activation of NADHP oxidase by affecting the subunits of key enzymes. However, MEM reduced levels of phosho-PKC-Akt-mTOR, with no changes in CAZ/TB. ConclusionsWe firstly demonstrate that β-Lactams showed the definitive immunomodulation effects through modulating NETs formation, which is depended on PKC-Akt-mTOR signal pathway.

Cell proliferation of rat bladder urothelium induced by nicotine is suppressed by the NADPH oxidase inhibitor, apocynin

Tobacco smoking is a major risk factor for human cancers including urinary bladder carcinoma. In a previous study, nicotine enhanced rat urinary bladder carcinogenesis in a two-stage carcinogenesis model. Nicotine also induced cytotoxicity in the bladder urothelium in a short-term study. In the present study, male rats were treated with nicotine (40 ppm) in drinking water co-administered with the NADPH oxidase inhibitor, apocynin (0, 250 or 750 mg/kg) in diet for 4 weeks. The apocynin treatment induced no clinical toxic effects. Reduction of reactive oxygen species (ROS) by apocynin was confirmed by immunohistochemistry of 8-OHdG in the bladder urothelium. Incidences of simple hyperplasia, cell proliferation and apoptosis were reduced by apocynin treatment in the bladder urothelium. However, despite reduction of cell proliferation (labeling index), apocynin did not affect the incidence of simple hyperplasia, apoptosis, or ROS generation in the kidney pelvis urothelium, in addition to 8-OHdG positivity induced by nicotine being lower. In vitro, apocynin (500 μM) reduced ROS generation, but induced cell proliferation in bladder cancer cell lines (T24 and UMUC3 cells). These data suggest that oxidative stress may play a role in the cell proliferation of the bladder urothelium induced by nicotine.

ROS-Mediated Necroptosis Is Involved in Iron Overload-Induced Osteoblastic Cell Death.

Excess iron has been reported to lead to osteoblastic cell damage, which is a crucial pathogenesis of iron overload-related osteoporosis. However, the cytotoxic mechanisms have not been fully documented. In the present study, we focused on whether necroptosis contributes to iron overload-induced osteoblastic cell death and related underlying mechanisms. Here, we showed that the cytotoxicity of iron overload in osteoblastic cells was mainly due to necrosis, as evidenced by the Hoechst 33258/PI staining, Annexin-V/PI staining, and transmission electronic microscopy. Furthermore, we revealed that iron overload-induced osteoblastic necrosis might be mediated via the RIPK1/RIPK3/MLKL necroptotic pathway. In addition, we also found that iron overload was able to trigger mitochondrial permeability transition pore (mPTP) opening, which is a critical downstream event in the execution of necroptosis. The key finding of our experiment was that iron overload-induced necroptotic cell death might depend on reactive oxygen species (ROS) generation, as N-acetylcysteine effectively rescued mPTP opening and necroptotic cell death. ROS induced by iron overload promote necroptosis via a positive feedback mechanism, as on the one hand N-acetylcysteine attenuates the upregulation of RIPK1 and RIPK3 and phosphorylation of RIPK1, RIPK3, and MLKL and on the other hand Nec-1, siRIPK1, or siRIPK3 reduced ROS generation. In summary, iron overload induced necroptosis of osteoblastic cells in vitro, which is mediated, at least in part, through the RIPK1/RIPK3/MLKL pathway. We also highlight the critical role of ROS in the regulation of iron overload-induced necroptosis in osteoblastic cells.

Role of binding immunoglobulin protein (BiP) in Graves' orbitopathy pathogenesis.

Inflammation and adipogenesis represent the main pathogenic mechanisms of Graves' orbitopathy (GO), and oxidative stress is a well-known inducer of GO pathology. Endoplasmic reticulum (ER) stress has been suggested as a major contributor to inflammation and reactive oxygen species (ROS) generation. In this study, we investigated the role of the ER-stress chaperone protein, binding immunoglobulin protein (BiP), in GO pathogenesis. Using primary cultures of orbital fibroblasts from patients with GO, we examined the role of BiP in GO pathogenesis by silencing its expression with small-interfering RNA (siRNA). Inflammatory cytokine expression was analysed by Western blotting and ELISA. Intracellular ROS levels induced by hydrogen peroxide or cigarette smoke extract were measured by 5-(and 6)-carboxy-20,70-dichlorodihydrofluorescein diacetate staining and flow cytometry. After adipogenic differentiation in BiP siRNA-transfected cells, the cells were stained with Oil Red O, and the levels of adipogenic transcription factors were determined by Western blot analysis. BiP mRNA expression levels were significantly higher in GO orbital tissues than in non-GO orbital tissues. Silencing BiP attenuated the expression of pro-inflammatory cytokines (interleukin-6, intercellular adhesion molecule-1, and monocyte chemotactic protein-1) in primary cultured GO orbital fibroblasts. Silencing BiP also reduced ROS generation, hyaluronan production, and adipocyte differentiation. These findings suggest that ER stress is involved in the aetiology of GO and that modulation of ER stress has therapeutic potential for GO.

Melatonin restores neutrophil functions and prevents apoptosis amid dysfunctional glutathione redox system.

Melatonin is a chronobiotic hormone, which can regulate human diseases like cancer, atherosclerosis, respiratory disorders, and microbial infections by regulating redox system. Melatonin exhibits innate immunomodulation by communicating with immune system and influencing neutrophils to fight infections and inflammation. However, sustaining redox homeostasis and reactive oxygen species (ROS) generation in neutrophils are critical during chemotaxis, oxidative burst, phagocytosis, and neutrophil extracellular trap (NET) formation. Therefore, endogenous antioxidant glutathione (GSH) redox cycle is highly vital in regulating neutrophil functions. Reduced intracellular GSH levels and glutathione reductase (GR) activity in the neutrophils during clinical conditions like autoimmune disorders, neurological disorders, diabetes, and microbial infections lead to dysfunctional neutrophils. Therefore, we hypothesized that redox modulators like melatonin can protect neutrophil health and functions under GSH and GR activity-deficient conditions. We demonstrate the dual role of melatonin, wherein it protects neutrophils from oxidative stress-induced apoptosis by reducing ROS generation; in contrast, it restores neutrophil functions like phagocytosis, degranulation, and NETosis in GSH and GR activity-deficient neutrophils by regulating ROS levels both in vitro and in vivo. Melatonin mitigates LPS-induced neutrophil dysfunctions by rejuvenating GSH redox system, specifically GR activity by acting as a parallel redox system. Our results indicate that melatonin could be a potential auxiliary therapy to treat immune dysfunction and microbial infections, including virus, under chronic disease conditions by restoring neutrophil functions. Further, melatonin could be a promising immune system booster to fight unprecedented pandemics like the current COVID-19. However, further studies are indispensable to address the clinical usage of melatonin.

Ceria Nanoparticles Mitigate the Iron Oxidative Toxicity of Human Retinal Pigment Epithelium.

Oxidative injury is implicated in retinal damage observed in age-related macular degeneration (AMD), as well as other degenerative conditions. Abnormally elevated levels of iron accumulation within the retinal pigment epithelium have been detected in eyes with AMD, and it is suspected to play a role in the pathogenesis through the production of reactive oxygen species (ROS). Ceria nanoparticles (CNP) have the ability to scavenge ROS. This study sought to evaluate the ability of CNP to mitigate iron-induced oxidative stress and assess cell viability in the human ARPE-19 cell line in vitro. Cell viability was measured by an MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay and compared between experimental groups undergoing 48-hr exposure to a ferrous iron solution with and without 24-hr CNP pre-treatment. The CNP effect on ROS formation was evaluated additionally by H2DCFDA (2,7-dichlorodihydrofluorescein diacetate) fluorescent probe assay and superoxide dismutase assay. CNP demonstrated a three-fold increase in cell viability and a reduction in ROS generation. The results show a promising treatment modality for diseases causing oxidative damage in the eye.

Brain-Specific Serine-47 Modification of Cytochrome c Regulates Cytochrome c Oxidase Activity Attenuating ROS Production and Cell Death: Implications for Ischemia/Reperfusion Injury and Akt Signaling.

We previously reported that serine-47 (S47) phosphorylation of cytochrome c (Cytc) in the brain results in lower cytochrome c oxidase (COX) activity and caspase-3 activity in vitro. We here analyze the effect of S47 modification in fibroblast cell lines stably expressing S47E phosphomimetic Cytc, unphosphorylated WT, or S47A Cytc. Our results show that S47E Cytc results in partial inhibition of mitochondrial respiration corresponding with lower mitochondrial membrane potentials (ΔΨm) and reduced reactive oxygen species (ROS) production. When exposed to an oxygen-glucose deprivation/reoxygenation (OGD/R) model simulating ischemia/reperfusion injury, the Cytc S47E phosphomimetic cell line showed minimal ROS generation compared to the unphosphorylated WT Cytc cell line that generated high levels of ROS upon reoxygenation. Consequently, the S47E Cytc cell line also resulted in significantly lower cell death upon exposure to OGD/R, confirming the cytoprotective role of S47 phosphorylation of Cytc. S47E Cytc also resulted in lower cell death upon H2O2 treatment. Finally, we propose that pro-survival kinase Akt (protein kinase B) is a likely mediator of the S47 phosphorylation of Cytc in the brain. Akt inhibitor wortmannin abolished S47 phosphorylation of Cytc, while the Akt activator SC79 maintained S47 phosphorylation of Cytc. Overall, our results suggest that loss of S47 phosphorylation of Cytc during brain ischemia drives reperfusion injury through maximal electron transport chain flux, ΔΨm hyperpolarization, and ROS-triggered cell death.

Brain Factor-7\xae improves learning and memory deficits and attenuates ischemic brain damage by reduction of ROS generation in stroke in vivo and in vitro

Brain Factor-7® (BF-7), silk fibroin peptide, is known to be effective in improvement of memory and learning ability. In this study, the effects of BF-7 (10 mg/kg, p.o., pre-treatment for 7 days and post-treatment for 7 days) on neuroprotection and memory and learning functions were investigated in a rat model of transient focal cerebral ischemia and a gerbil model of transient global forebrain ischemia. Furthermore, to find the mechanism of BF-7, we examined the neuroprotective and antioxidative effects of BF-7 in vitro using neuroblastoma (SH-SY5Y) cells. In vivo model, treatment with BF-7 significantly reduced the number of errors in 8-arm maze test and significantly increased latency time in passive avoidance test at 7 days after focal ischemia compared to those in the vehicle-treated group. In addition, treatment with BF-7 significantly decreased the infarct size or neuronal death at 7 day following transient ischemia compared to that in the vehicle-treated group. In vitro model, 10 or 20 μg/ml of BF-7 treatment significantly increased cell viability in dose-dependent manner. In addition, oxidative stress was significantly attenuated in the ischemic cells, showing that 10 or 20 μg/ml of BF-7 treatment significantly reduced the generation of reactive oxygen species (ROS) compared to that in the ischemic cells. These results indicate that BF-7 treatment can attenuate ischemic damages and improve memory deficits via reduction of ROS generation.

Fumonisin B1 triggers the formation of bovine neutrophil extracellular traps

Fumonisin B1 (FB1) is a congener of fumonisins produced by Fusarium species that may be found as corn contaminants threatening health of humans and animals. FB1 causes a variety of toxicity effects, including hepatotoxic, nephrotoxic and cytotoxic effects. However, detailed mechanisms associated with FB1 immunotoxicity in neutrophils are still unclear. To accomplish this, we utilized neutrophils to study the mechanisms of FB1 immunotoxicity. In the current study, we found that FB1 induced the formation of neutrophil extracellular traps (NETs), increased reactive oxygen species (ROS) levels, and decreased SOD and CAT activities. Concurrently, FB1 treatment led to the concentration-dependent phosphorylation of ERK-1/2 and p38 in neutrophils. Moreover, we demonstrated that FB1-induced NET formation was dependent of NADPH oxidase activity. Pretreatment of neutrophils with DPI, U0126 and SB202190 significantly reduced ROS generation, and prevented NET formation, further suggesting that ROS dependent activation of ERK 1/2 and p38 pathways, which possibly mediate FB1-induced NET release in neutrophils. Thus, NET formation and ROS production could be attributed to FB1 immunotoxicity, which might enrich the toxicological mechanisms of FB1.

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.

Theaflavin-3, 3'-Digallate Suppresses RANKL-Induced Osteoclastogenesis and Attenuates Ovariectomy-Induced Bone Loss in Mice.

Theaflavin-3, 3′-digallate (TF3) is extracted from black tea and has strong antioxidant capabilities. The aim of this study was to assess the influences of TF3 on osteoclastogenesis and explore the underlying mechanisms. TF3 efficiently decreased receptor activator of nuclear factor-kappa B ligand (RANKL)-induced osteoclast formation and reactive oxygen species (ROS) generation in a dose-dependent manner. Mechanistically, TF3 reduced ROS generation by activating nuclear factor erythroid 2-related factor 2 (Nrf2) and its downstream heme oxygenase-1 (HO-1) and also inhibited the mitogen-activated protein kinases (MAPK) pathway. Moreover, micro-computed tomography (CT) analysis, hematoxylin and eosin (H&E) staining, and TRAP staining of the femurs of C57BL/6J female mice showed that TF3 markedly attenuated bone loss and osteoclastogenesis in mice. Immunofluorescence staining, 2′,7′-dichlorofluorescein diacetate (DCFH-DA) staining, and measurement of the levels of malonaldehyde (MDA) and superoxide dismutase (SOD) revealed that TF3 increased the expression of Nrf2 and decreased the intracellular ROS level in vivo. These findings indicated that TF3 may have the potential to treat osteoporosis and bone diseases related to excessive osteoclastogenesis via inhibiting the intracellular ROS level.

Foliar application of gibberellic acid endorsed phytoextraction of copper and alleviates oxidative stress in jute (Corchorus capsularis L.) plant grown in highly copper-contaminated soil of China.

Copper (Cu) is an abundant essential micronutrient element in various rocks and minerals and is required for a variety of metabolic processes in both prokaryotes and eukaryotes. However, excess Cu can disturb normal development by adversely affecting biochemical reactions and physiological processes in plants. The present study was conducted to explore the potential of gibberellic acid (GA3) on fibrous jute (Corchorus capsularis L.) seedlings grown on Cu mining soil obtained from Hubei Province China. Exogenous application of GA3 (10, 50, and 100 mg/L) on 60-day-old seedlings of C. capsularis which was able to grow in highly Cu-contaminated soil (2221 mg/kg) to study different morphological, physiological, and Cu uptake and accumulation in different parts of C. capsularis seedlings. According to the results, increasing concentration of GA3 (more likely 100 mg/L) alleviates Cu toxicity in C. capsularis seedlings by increasing plant growth, biomass, photosynthetic pigments, and gaseous exchange attributes. The results also showed that exogenous application of GA3 reduced oxidative stress in C. capsularis seedlings by the generation of extra reactive oxygen species (ROS). The reduction in oxidative stress in C. capsularis seedlings is because that plant has strong enzymatic antioxidants [superoxidase dismutase (SOD), peroxidase (POD), ascorbate peroxidase (APX), and catalase (CAT)], which ultimately increased their activities to overcome oxidative damage in the cells/tissues. In addition to the plant growth, biomass, and photosynthesis, foliar application of GA3 also helps to increase metal (Cu) concentration in different parts of the plants when compared to 0 mg/L of application of GA3. From these findings, we can conclude that foliar application of GA3 plays a promising role in reducing ROS generation in the plant cells/tissues and increased phytoextraction of Cu in different plant parts. However, more investigation is needed on field experiments to find a combination of GA3 with a very higher concentration of Cu using fibrous C. capsularis.