Oxidative Cell Injury Research Articles

Resveratrol Attenuates Hydrogen Peroxide-induced Injury of Rat Ovarian Granulosa-lutein Cells by Resisting Oxidative Stress via the SIRT1/Nrf2/ARE Signaling Pathway.

This paper aims to reveal the molecular mechanism of resveratrol against oxidative stress and cell injury. The ovarian granulosa-lutein cell injury and apoptosis induced by oxidative stress may be responsible for female luteal phase deficiency. The antioxidant function of resveratrol has been confirmed; however, its effect on the expression of antioxidant enzymes and regulatory mechanisms in ovarian granulosa-lutein cells remains unclear. This study aimed to investigate the role of the SIRT1/Nrf2/ARE signaling pathway in the effect of resveratrol on the hydrogen peroxide-induced injury of rat ovarian granulosa-lutein cells. In this study, ovarian granulosa-lutein cells extracted from 3-week female SD rats were treated with 200 μM H2O2 in the presence or absence of 20 μM resveratrol. siRNA-SIRT1 and siRNA-Nrf2 were used to inhibit the expression of SIRT1 and Nrf2, respectively. Cell counting kit 8 (CCK-8), cellular morphology, progesterone secretion, and estradiol were used to evaluate cell injury. Hoechst 33258 staining was used to measure cell apoptosis. DHE staining, DCFH-DA staining, malondialdehyde content, protein carbonyl content, total antioxidant capacity and SOD viability were used to estimate the levels of oxidative stress. Western blot analysis was used to detect the levels of apoptosis-related proteins, and SIRT1/Nrf2/ARE signaling pathway-related proteins. The H2O2 treatment-induced rat ovarian granulosa-lutein cells injury was shown as decreased cell viability, impaired cellular morphology, and decreased levels of progesterone and estradiol. The H2O2 treatment also exacerbated cell apoptosis demonstrated as more apoptotic cells stained by Hoechst staining, decreased level of anti-apoptosis protein Bcl-2 and increased level of pro-apoptosis protein Bax. These effects of cell injury and apoptosis induced by H2O2 can be ameliorated by resveratrol. Resveratrol also alleviated oxidative stress induced by H2O2, supported by decreased superoxide anion and cellular total ROS, decreased malondialdehyde and protein carbonyl levels, and increased total antioxidant capacity and SOD viability. Western blot results demonstrated resveratrol reversed the H2O2-induced decrease in levels of antioxidant enzymes containing ARE sequences and activated SIRT1/Nrf2 pathway. Further treatment by siRNA-Nrf2 suggested resveratrol could not activate the expression of antioxidant enzymes under a condition of inhibition of Nrf2. This study demonstrates that resveratrol attenuated oxidative stress to protect H2O2-induced rat ovarian granulosa-lutein cell injury and apoptosis via SIRT1/Nrf2/ARE signaling pathway.

Biological mechanism of cell oxidative stress and death during short-term exposure to nano CuO

It is well known that copper oxide nanoparticles (CuO NPs) are heavily toxic on in vitro systems. In human alveolar epithelial cells, the mechanism of toxicity is mostly related to oxidative insults, coming from intracellularly dissolved copper ions, finally leading to apoptotic or autophagic cell death. Our hypothesis is based on possible early oxidative events coming from specific NP surface reactivity able to undermine the cell integrity and to drive cell to death, independently from Lysosomal-Enhanced Trojan Horse mechanism. Two types of CuO NPs, with different oxidative potential, were selected and tested on A549 cells for 1 h and 3 h at 10, 25, 50 and 100 µg/ml. Cells were then analyzed for viability and oxidative change of the proteome. Oxidative by-products were localized by immunocytochemistry and cell-NP interactions characterized by confocal and electron microscopy techniques. The results show that CuO NPs induced oxidative changes soon after 1 h exposure as revealed by the increase in protein carbonylation and reduced-protein-thiol oxidation. In parallel, cell viability significantly decreased, as shown by MTT assay. Such effects were higher for CuO NPs with more crystalline defects and with higher ROS production than for fully crystalline NPs. At these exposure times, although NPs efficiently interacted with cell surface and were taken up by small endocytic vesicles, no ion dissolution was visible inside the lysosomal compartment and no effects were produced by extracellularly dissolved copper ions. In conclusion, a specific NP surface-dependent oxidative cell injury was demonstrated. More detailed studies are required to understand which targets precociously react with CuO NPs, but these results introduce new paradigms for the toxicity of the metal-based NPs, beyond the Lysosomal-Enhanced Trojan horse-related mechanism, and open-up new opportunities to investigate the interactions and effects at the bio-interface for designing safer as well as more effective CuO-based biocides.

Lipopolysaccharide promotes apoptosis and oxidative injury of porcine small intestinal epithelial cells by down-regulating the expression of glutamine transporter ASCT2.

The present study aimed to investigate the effects of lipopolysaccharide (LPS) stimulation on oxidative damage, apoptosis, and glutamine (Gln) transporter Alanine-Serine-Cysteine transporter 2 (ASCT2) expression in porcine small intestinal epithelial cells (IPEC-J2), and preliminarily elucidated the relationship between ASCT2 expression level and oxidative damage and apoptosis of IPEC-J2 cells. IPEC-J2 cells were treated without (control group, CON, N = 6) or with 1 μg/mL LPS (LPS group, LPS, N = 6). Cell viability, lactate dehydrogenase (LDH) content, malonaldehyde (MDA), anti-oxidant enzymes (superoxide dismutase [SOD], catalase [CAT], glutathione peroxidase [GSH-Px], and total anti-oxidant capacity [T-AOC]), apoptosis of IPEC-J2 cells, the expression of Caspase3, the expression of ASCT2 mRNA and ASCT2 protein was detected. The results showed that LPS stimulation of IPEC-J2 cells significantly reduced the cell viability, and anti-oxidant enzymes activity (SOD, CAT, and GSH-Px), and significantly increased LDH and MDA release. Flow cytometry results showed that LPS stimulation significantly increased the late apoptosis rate and the total apoptosis rate of IPEC-J2 cells. The immunofluorescence results showed that the fluorescence intensity of LPS stimulated IPEC-J2 cells was significantly enhanced. LPS stimulation significantly decreased the mRNA and protein expression of ASCT2 in IPEC-J2 cells. The correlation analysis showed that ASCT2 expression was negatively correlated with apoptosis, and positively correlated with the anti-oxidant capacity of IPEC-J2 cells. According to the results of this study, it can be preliminarily concluded that LPS promotes the apoptosis and oxidative injury of IPEC-J2 cells by down-regulating the expression of ASCT2.

The effect of Myrtus communis L. extract on nephrolithiasis model in rats.

Nephrolithiasis is a common urological disease that can lead to renal failure. Oxidative stress has been shown to be a contributing factor for nephrolithiasis and many agents have been studied to prevent and treat oxidative stress-related nephrolithiasis and renal damage. Myrtus communis (MC) extract has been shown to be an important antioxidant in different animal models. In this study, MC extract was administered preventively or therapeutically to rats with kidney stones, and its effectiveness was investigated. Wistar albino rats were divided into four groups (n=8); control (C), ethylene glycol (EG), EG+preventive MC, and EG+curative MC groups. The nephrolithiasis model was created by adding 0.75% EG to drinking water for 8 weeks. Ultimately, 24-hour urine was collected to measure calcium, citrate, and creatinine levels. After decapitation, kidney tissues were harvested for histological analyses, measurement of osteopontin and 8-hydroxydeoxyguanosine (8-OHdG) levels, and N-acetyl-β-glucosaminidase (NAG), myeloperoxidase (MPO) and caspase-3 activities. In 24-hour urine samples, calcium, citrate and creatinine levels were decreased in the EG group, while oxalate levels were increased and in treatment groups these parameters returned to control levels. MPO, 8-OHdG, caspase-3 and NAG activity were significantly increased in tissue and these changes were reversed in both MC groups. Histological findings also supported the biochemical parameters. MC can reduce oxidative stress and histopathological changes in kidney tissues in rat nephrolithiasis model when used as either a preventive or therapeutic agent. If supported with further clinical trials, MC might have clinical implications in preventing oxidative renal cell injury and ultimately kidney stone formation.

Comparative Biochemical Effects of Natural and Synthetic Pesticides on Preserved Vigna Subterranea in Male Albino Rats

Preservatives both natural and synthetic have been widely used to store food in order to ensure all season availability and safety for human consumption. This study was carried out to evaluate the biochemical effects of natural and synthetic pesticides preserved Vigna subterranea in male albino rats. Thirty six adult male albino weighing between 150 – 200 g were randomly allocated into 6 groups A, B, C, D, E and F (n=6 each). Vigna subterranea were preserved for six months in 5 air tight containers: preserved with neem ash, Pepper, Dichlovors, Aluminium Phosphide and without preservative. The preserved beans were milled into powder and used as dietary supplement for the rats. All the animals freely received animal feed and water. Group A, B, C, D, E and F were fed with animal feed and water only, Vigna subterranea without preservative, neem Ash preserved Vigna subterranea, Pepper preserved Vigna subterranea, Dichlovors preserved Vigna subterranea and Aluminium Phosphide preserved Vigna subterranea respectively. The dietary intervention lasted for 2 months after which the animals were sacrificed. The blood sample of each rat was collected and analyzed using standard methods. The results from the analysis of the liver function across the groups with preservatives showed increase in liver enzymes. The serum AST shows significant increase (p < 0.05) in Group D compared to Group A. ALT significantly (p < 0.05) increased in all the test groups except the group fed with pepper compared to the control which indicates there may be possible injury to the liver. The result showed elevation (p < 0.05) of C-reactive protein in all the test group compared to Group A and significantly (p < 0.05) reduced in the group fed with neem Ash, which may indicate injury and inflammation in the test animals of the other preservatives. In the kidney, the serum urea findings showed significant increase (p<0.05) in all the groups fed with preservatives compared to the Group A and the concentrations of Creatinine increased significantly (p<0.05) in all the groups fed with the preserved V. subterranea compared to the control indicating there may be possible damage to the kidney. A significant decrease (p<0.05) in the antioxidants-CAT, SOD and GPx and increase (p<0.05) in MDA in the group fed with preservatives indicates there may be increase in lipid peroxidation, leading to oxidative stress and cell injury in the test animals. The results of this study suggest that the use of some of these preservatives which could be natural and synthetic may be toxic and could lead to cellular and tissue damage.

Raman Spectroscopy and Imaging Studies of Human Digestive Tract Cells and Tissues—Impact of Vitamin C and E Supplementation

Cancers of digestive tract such as colorectal cancer (CRC) and gastric cancer (GC) are the most commonly detected types of cancer worldwide and their origin can be associated with oxidative stress conditions. Commonly known and followed antioxidants, such as vitamin C and E, are widely considered as potential anti-cancer agents. Raman spectra have great potential in the biochemical characterization of matter based on the fact that each molecule has its own unique vibrational properties. Raman spectroscopy allows to precisely characterize components (proteins, lipids, nucleic acids). The paper presents the application of the Raman spectroscopy technique for the analysis of tissue samples and cells of the human colon and stomach. The main goal of this study is to show the differences between healthy and cancerous tissues from the human digestive tract and human normal and cancer colon and gastric cell lines. The paper presents the spectroscopic characterization of normal colon cells, CCD-18 Co, in physiological and oxidative conditions and effect of oxidative injury of normal colon cells upon supplementation with vitamin C at various concentrations based on Raman spectra. The obtained results were related to the Raman spectra recorded for human colon cancer cells-CaCo-2. In addition, the effect of the antioxidant in the form of vitamin E on gastric cancer cells, HTB-135, is presented and compared with normal gastric cells-CRL-7869. All measured gastric samples were biochemically and structurally characterized by means of Raman spectroscopy and imaging. Statistically assisted analysis has shown that normal, ROS injured and cancerous human gastrointestinal cells can be distinguished based on their unique vibrational properties. ANOVA tests, PCA (Principal Component Analysis) and PLSDA (Partial Least Squares Discriminant Analysis) have confirmed the main role of nucleic acids, proteins and lipids in differentiation of human colon and stomach normal and cancer tissues and cells. The conducted research based on Raman spectra proved that antioxidants in the form of vitamin C and E exhibit anti-cancer properties. In consequence, conducted studies proved that label-free Raman spectroscopy may play an important role in clinical diagnostic differentiation of human normal and cancerous gastrointestinal tissues and may be a source of intraoperative information supporting histopathological analysis.

Effect of Esculetin on Tert-Butyl Hydroperoxide-Induced Oxidative Injury in Retinal Pigment Epithelial Cells In Vitro.

Esculetin is a coumarin-derived compound with antioxidant and anti-inflammatory properties. The current study aims to evaluate the therapeutic implications of esculetin on retinal dysfunction and uncover the underlying mechanisms. Tert-butyl hydroperoxide (t-BHP) at a concentration of 300 μM was used to induce oxidative stress in human retinal pigment epithelial cell line (ARPE-19) cells. Esculetin at concentrations below 250 μM did not cause cytotoxicity to ARPE-19 cells. Cell viability analysis confirmed that t-BHP induced oxidative injury of ARPE-19 cells. However, ARPE-19 cells were protected from t-BHP-induced oxidative injury by esculetin in a concentration-dependent manner. As a result of the TUNEL assay to confirm apoptosis, esculetin treatment reduced the number of TUNEL-positive cells. Esculetin down-regulated the expression levels of Bax, Caspase-3, and PARP and up-regulated the expression level of Bcl2. Collectively, this study demonstrates that esculetin exerts potent antioxidant properties in ARPE-19 cells, inhibiting t-BHP-induced apoptosis under the regulation of apoptotic factors.

Fucoxanthin ameliorates oxidative injury and inflammation of human bronchial epithelial cells induced by cigarette smoke extract via the PPARγ/NF‑κB signaling pathway.

Chronic obstructive pulmonary disease (COPD) is a prevalent and long-term airway disease. It has been reported that fucoxanthin (FX) exhibits anti-inflammatory and antioxidant effects. However, the underlying mechanism of FX in COPD remains unknown. Therefore, to investigate the effect of FX on COPD, BEAS-2B cells were treated with cigarette smoke extract (CSE). The viability of BEAS-2B cells treated with increasing doses of FX was assessed by Cell Counting Kit-8. Lactate dehydrogenase (LDH) levels were measured using a corresponding kit. In addition, ELISA was carried out to detect the content of TNF-α, IL-1β and IL-6. Additionally, a TUNEL assay and western blot analysis were performed to assess the cell apoptosis rate. Furthermore, 2',7'-dichlorodihydrofluorescein diacetate was used to measure reactive oxygen species levels, while the contents of oxidative stress-associated indexes were determined using the corresponding kits. Bioinformatics analysis using the search tool for interactions of chemicals database predicted that peroxisome proliferator-activated receptor γ (PPARγ) may be a target of FX. The binding capacity of FTX with PPARγ was confirmed by molecular docking. The protein expression levels of the PPARγ/NF-κB signaling-associated factors were detected by western blot analysis. Finally, the regulatory mechanism of FX in COPD was revealed following cell treatment with the PPARγ inhibitor, T0070907. The results demonstrated that FX enhanced CSE-induced BEAS-2B cell viability and attenuated CSE-induced BEAS-2B cell inflammation and oxidative damage, possibly via triggering PPARγ/NF-κB signaling. Pre-treatment of BEAS-2B cells with the PPARγ inhibitor, T0070907, could reverse the protective effects of FX on CSE-induced BEAS-2B cells. Overall, the present study suggested that FX could ameliorate oxidative damage as well as inflammation in CSE-treated human bronchial epithelial in patients with COPD via modulating the PPARγ/NF-κB signaling pathway.

AP2a-Mediated Upregulation of miR-125a-5p Ameliorates Radiation-Induced Oxidative Stress Injury via BRD4/Nrf2/HO-1 Signaling.

Radiation therapy is widely used to restrain tumor progression, but it is always accompanied by damage to healthy tissues. We aimed to probe the impact and mechanism of activator protein 2a (AP2a) and miR-125a-5p in radiation-induced oxidative stress injury. Human umbilical vein endothelial cells (HUVECs) were treated with X rays to induce radiation injury in vitro. Cell viability was measured using MTT assays. Flow cytometry assay was employed to detect the apoptosis rate. Oxidative stress markers were evaluated by detection kits. Gene or protein levels were determined by RT-qPCR or Western blotting. Validation of the interaction of miR-125a-5p with BRD4 and AP2a was conducted by dual luciferase assay or ChIP. MiR-125a-5p and AP2a were decreased in irradiated HUVECs, whereas BRD4 was increased. MiR-125a-5p overexpression or BRD4 silencing alleviated the cell viability decline, apoptosis, and oxidative stress injury caused by radiation treatment. MiR-125a-5p repressed the BRD4 level. The protective effects of miR-125a-5p overexpression in the radiation-induced oxidative injury were impeded by BRD4 overexpression. Moreover, AP2a bound to the promoter of miR-125a-5p. MiR-125a-5p inhibition reversed the effects of AP2a overexpression on radiational oxidative injury by modulating Nrf2/HO-1 signaling. AP2a transcriptionally activated miR-125a-5p ameliorated oxidative stress injury of HUVECs caused by radiation through Nrf2/HO-1 signaling.

Role of microRNAs in the regulation of blood-brain barrier function in ischemic stroke and under hypoxic conditions in vitro

The blood-brain barrier (BBB) is a highly specialized structure that separates the brain from the blood and allows the exchange of molecules between these two compartments through selective channels. The breakdown of the BBB is implicated in the development of severe neurological diseases, especially stroke and traumatic brain injury. Oxygen-glucose deprivation is used to mimic stroke and traumatic brain injury in vitro. Pathways that trigger BBB dysfunction include an imbalance of oxidative stress, excitotoxicity, iron metabolism, cytokine release, cell injury, and cell death. MicroRNAs are small non-coding RNA molecules that regulate gene expression and are emerging as biomarkers for the diagnosis of central nervous system (CNS) injuries. In this review, the regulatory role of potential microRNA biomarkers and related therapeutic targets on the BBB is discussed. A thorough understanding of the potential role of various cellular and linker proteins, among others, in the BBB will open further therapeutic options for the treatment of neurological diseases.

Protective effect of fluoxetine against oxidative stress induced by renal ischemia-reperfusion injury via the regulation of miR-450b-5p/Nrf2 axis.

The present study was performed to assess the protective effect of fluoxetine (FLX) on renal ischemia-reperfusion injury (IRI) via the regulation of miR-450b-5p/Nrf2 axis in male rats. In vivo, these male rats were randomly divided into different treatment groups. The rats were administered with FLX (20 mg/kg, intraperitoneally) once daily for 3 days before operation. The pathomorphological changes of renal tissues were assessed by histological examination and Masson staining. In vitro, HK-2 cells were used to detect the activity by CCK-8 assay in Hypoxia/Reoxygenation (H/R) group and Hypoxia/Reoxygenation+Fluoxetine (H/R+FLX) group. In addition, the oxidative stress biomarkers were evaluated. Subsequently, Nrf2, NF-κB, and Nrf2-dependent antioxidant enzymes, were detected by Western blot assay. In vivo, the pathological changes and serological renal function were significantly relieved in the rats with the pre-treatment of FLX, compared to IRI group. After FLX stimulation, the expression levels of oxidative stress indices significantly decreased, while tissue antioxidant indices significantly increased, compared to IRI group. The differently expressed miRNAs on renal IRI in male rats were screened out by miRNA microarray, especially showing that miR-450b-5p was selected as the target miRNA. Following miR-450b-5p agomir injection, the pathological changes and oxidative stress biomarkers significantly aggravated, whether in IRI group or IRI+FLX group. Bioinformatics analysis and double-luciferase reporter assay demonstrated that miR-450b-5p directly targeted Nrf2. The expression level of NF-κB significantly increased, while the expression levels of Nrf2 and Nrf2-dependent antioxidant enzymes significantly decreased after miR-450b-5p agomir injection. Furthermore, the expression levels of Nrf2 and it-dependent antioxidant enzymes were apparently increased in ischemic kidney after the transfection of miR-450b-5p mimic+recombination protein Nrf2, as well as the decreased expression levels of intracellular ROS and iNOS. In vitro, FLX significantly increased HK-2 cell viability, and relieved H/R HK-2 cell oxidative injury via down-regulating ROS and iNOS. In addition, H/R-induced oxidative damage was recovered with miR-450b-5p mimic and recombination protein Nrf2. Consequently, FLX played an important protective role in renal IRI-induced oxidative damage by promoting antioxidation via targeting miR-450b-5p/Nrf2 axis.

Protective activity of Malus doumeri leaf extract on H2O2-induced oxidative injury in H9C2 rat cardiomyocytes.

In this study, Malus doumeri leaf extract (MDLE) was used to test its anti-oxidation capacity in vitro, it has been preliminarily analyzed for H2O2-induced oxidative damage in H9C2 cells and its main active components. The antioxidant capacity through DPPH (1, 1-Diphenyl-2-Picrylhydrazyl), ABTS+• [2,2,2'-azino-BIS-(3-ethylbenzo-thiazoline-6-sulfonic acid)] radical ion, •OH (hydroxyl radical), and (superoxide anion) were determined in vitro. The proliferation of H9C2 cells was examined by MTT [3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-Tetrazolium bromide]. MDA (malondialdehyde), SOD (superoxide dismutase), CAT (catalase), GSH (glutathione), and GSH-Px (glutathione peroxidase) were determined by colorimetry. Apoptosis induced by oxidative damage was detected by flow cytometry. The mRNA expression of antioxidant related genes of SOD, CAT, GSH, and GSH-Px were checked by qRT-PCR (quantitative real-time polymerase chain reaction). The MDLE main active components were analyzed by HPLC (high-performance liquid chromatography). MDLE had significant scavenging effects on DPPH, ABTS+•, •OH, and superoxide anion radicals in a concentration-dependent manner. H2O2 treatment could significantly lead to oxidative stress injury of H9C2 cells, and MDLE treatment significantly improved the degree of H9C2 cell damage, and showed a positive correlation with concentration. MDLE can also reduce apoptosis caused by oxidative damage. MDLE treatment could significantly reduce MDA content and increase CAT, SOD, GSH, and GSH-Px contents and expression. In addition, by HPLC analysis, the following six bioactive components were detected from MDLE: chlorogenic acid, isoquercitrin, quercetin, baicalin, and phloretin. Therefore, MDLE has a good protective effect on myocardial cells.

Ginseng polysaccharide attenuates red blood cells oxidative stress injury by regulating red blood cells glycolysis and liver gluconeogenesis

Ethnopharmacological relevancePanax ginseng C.A. Mey (PG) is famous for “Qi-tonifying” effect, which has a medicinal history of more than 2 millennia. Modern pharmacology has confirmed that the “Qi-tonifying” effect of PG may be closely related to its pharmacological properties such as anti-oxidation, antineoplastic and treatment of cardiovascular disease. As one of the earliest cells affected by oxidative stress, RBCs are widely used in the diagnosis of diseases. Ginseng polysaccharide (GPS), is one of the major active components of PG, which plays an important role in resisting oxidative stress, affecting energy metabolism and other effects. However, the molecular mechanism explaining the “Qi-tonifying” effect of GPS from the perspective of RBCs oxidative damage has not been reported. Aim of the studyThis study aimed to investigate the protective effect of GPS on oxidatively damaged RBCs using in vitro and in vivo models and explore the molecular mechanisms from the perspective of glycolysis and gluconeogenesis pathways. To provides a theoretical basis for the future research of antioxidant drugs. Materials and methodsEstablished three different in vitro and in vivo research models: an in vitro model of RBCs exposed to hydrogen peroxide (H2O2) (40 mM), an in vivo model of RBCs from rats subjected to exhaustive swimming, and an in vitro model of BRL-3A cells exposed to H2O2 (25 μM). All three models were also tested in the presence of different concentrations of GPS. ResultsThe findings showed that GPS was the most potent antagonist of H2O2-induced hemolysis and redox inbalance in RBCs. In exhaustive exercise rats, GPS ameliorated RBVs hemolysis, including reducing whole-blood viscosity (WBV), improving deformability, oxygen-carrying and -releasing capacities, which was related to the enhancing of antioxidant capacity. Moreover, GPS promoted RBCs glycolysis in rats with exhaustive exercise by recovering the activities of glycolysis-related enzymes and increasing band 3 protein expression, thereby regulating the imbalance of energy metabolism caused by oxidative stress. Furthermore, we demonstrated that GPS improved antioxidant defense system, enhanced energy metabolism, and regulated gluconeogenesis via activating PPAR gamma co-activator 1 alpha (PGC-1α) pathway in H2O2-exposed BRL-3A cells. Mechanistically, GPS promoted glycolysis and protected RBCs from oxidative injury was partly dependent on the regulation of gluconeogenesis, as inhibition of gluconeogenesis by metformin (Met) attenuates the regulation of antioxidant enzymes and key enzymes of glycolytic by GPS in exhaustive exercise rats. ConclusionThis study demonstrates that GPS protects RBCs from oxidative stress damage by promoting RBCs glycolysis and liver gluconeogenesis pathways. These results may contribute to the study of new RBCs treatments to boost antioxidant capacity and protect RBCs against oxidative stress.

Synthetic Secoisolariciresinol Diglucoside (LGM2605) Prevents Asbestos-Induced Inflammation and Genotoxic Cell Damage in Human Mesothelial Cells.

Although alveolar macrophages play a critical role in malignant transformation of mesothelial cells following asbestos exposure, inflammatory and oxidative processes continue to occur in the mesothelial cells lining the pleura that may contribute to the carcinogenic process. Malignant transformation of mesothelial cells following asbestos exposure occurs over several decades; however, amelioration of DNA damage, inflammation, and cell injury may impede the carcinogenic process. We have shown in an in vitro model of asbestos-induced macrophage activation that synthetic secoisolariciresinol diglucoside (LGM2605), given preventively, reduced inflammatory cascades and oxidative/nitrosative cell damage. Therefore, it was hypothesized that LGM2605 could also be effective in reducing asbestos-induced activation and the damage of pleural mesothelial cells. LGM2605 treatment (50 µM) of huma n pleural mesothelial cells was initiated 4 h prior to exposure to asbestos (crocidolite, 20 µg/cm2). Supernatant and cells were evaluated at 0, 2, 4, and 8 h post asbestos exposure for reactive oxygen species (ROS) generation, DNA damage (oxidized guanine), inflammasome activation (caspase-1 activity) and associated pro-inflammatory cytokine release (IL-1β, IL-18, IL-6, TNFα, and HMGB1), and markers of oxidative stress (malondialdehyde (MDA) and 8-iso-prostaglandin F2a (8-iso-PGF2α). Asbestos induced a time-dependent ROS increase that was significantly (p < 0.0001) reduced (29.4%) by LGM2605 treatment. LGM2605 pretreatment also reduced levels of asbestos-induced DNA damage by 73.6% ± 1.0%. Although levels of inflammasome-activated cytokines, IL-1β and IL-18, reached 29.2 pg/mL ± 0.7 pg/mL and 43.9 pg/mL ± 0.8 pg/mL, respectively, LGM2605 treatment significantly (p < 0.0001) reduced cytokine levels comparable to baseline (non-asbestos exposed) values (3.8 pg/mL ± 0.2 pg/mL and 5.4 pg/mL ± 0.2 pg/mL, respectively). Furthermore, levels of IL-6 and TNFα in asbestos-exposed mesothelial cells were high (289.1 pg/mL ± 2.9 pg/mL and 511.3 pg/mL ± 10.2 pg/mL, respectively), while remaining undetectable with LGM2605 pretreatment. HMGB1 (a key inflammatory mediator and initiator of malignant transformation) release was reduced 75.3% ± 0.4% by LGM2605. Levels of MDA and 8-iso-PGF2α, markers of oxidative cell injury, were significantly (p < 0.001) reduced by 80.5% ± 0.1% and 76.6% ± 0.3%, respectively. LGM2605, given preventively, reduced ROS generation, DNA damage, and inflammasome-activated cytokine release and key inflammatory mediators implicated in asbestos-induced malignant transformation of normal mesothelial cells.

Effects of blocking lactate production by 2-DG on hypoxic injury of HT22 neurons and its mechanisms

To investigate the effects of blocking lactate synthesis on the HT22 cell injuries caused by hypoxia. 2-deoxy-D-glucose (2-DG) is a non-metabolized glucose analogue that can inhibit lactate synthesis by blocking glycolysis. HT22 cells were divided into 4 groups: Control group, 2-DG group, Hypoxia group and 2-DG+Hypoxia group. The cells in control group and 2-DG treatment group were cultured in a 37℃, 5% CO2 incubator, and thecells in hypoxia group and 2-DG + Hypoxia group were cultured in a hypoxia incubator. The concentrations of 2-DG were 2.5 and 5 mmol/L, the concentration of oxygen was 0.3%, and the treatment time was 24 h. Cell activity was detected by CCK-8 assay, the levels of lactate in cell culture medium were detected by spectrophotometry, cell morphology was observed by fluorescence staining, the level of reactive oxygen species (ROS) was detected by flow cytometry, and the activities of superoxide dismutase (SOD) and catalase (CAT) were determined by enzyme activity kits. The protein expression levels of p-p38, t-p38 and β-actin were detected by Western blot. Compared with that in control group, the lactate level in culture medium and cell activity were decreased significantly (P＜0.01), the number of adherent cells was decreased, the level of ROS was increased (P＜0.01), and the enzyme activity of CAT was decreased (P＜0.05) in the 2-DG group. In the hypoxia group, the level of lactate in the culture medium was increased significantly (P＜0.01), the cell activity was decreased (P＜0.01), the number of adherent cells was decreased, the ROS levels were increased (P＜0.01), and the enzyme activities of CAT and SOD were decreased (P＜0.01 or P＜0.05). In 2-DG+Hypoxia group, the level of lactate was decreased significantly (P＜0.05), the cell viability was decreased significantly (P＜0.01), the number of cells was decreased significantly, and the ability of adhere to the wall was weakened significantly. The level of ROS was increased significantly (P＜0.01), the enzyme activities of CAT and SOD were decreased significantly (P＜0.01), the protein expression level of p-p38 was increased significantly (P＜0.05), and there was no change in t-p38. Compared with hypoxia groups, in 2-DG combined with hypoxia group, the level of lactate induced by hypoxia, the cell activity, and the enzyme activity level of CAT were decreased significantly (all P＜0.01), while the level of ROS was increased significantly (P＜ 0.01). Blocking lactate can reduce the cell activity level under hypoxia and aggravate the oxidative stress injury of HT22 cells. The mechanisms may be related to increasing ROS level and activating p38 signal pathway.

Wheat Straw Burial Improves Physiological Traits, Yield and Grain Quality of Rice by Regulating Antioxidant System and Nitrogen Assimilation Enzymes under Alternate Wetting and Drying Irrigation

Wheat straw burial has great potential to sustain rice production under alternate wetting and drying (AWD) irrigation. A field experiment was conducted with three wheat straw burial treatments, including without straw burial (NSB), with light straw burial of 300 kg/hm2 (LSB) and dense straw burial of 800 kg/hm2 (DSB), as well as three AWD regimes: alternate wetting/moderate drying (AWMD), alternate wetting/severe drying (AWSD) and alternate wetting/critical drying (AWCD). The rice growth and grain quality were higher in LSB and NSB than those in NSB under the same AWD regime. The AWMD × DSB treatment resulted in the highest yield, brown rice rate, milled rice rate, amylose content and protein content. Conversely, the AWCD × NSB treatment led to the lowest yield, brown rice rate, milled rice rate, amylose content and protein content. The active absorption area and nitrate reductase activity of roots were higher in the AWMD × DSB treatment than those in the AWCD × NSB treatment, as the former increased organic carbon and nitrogen contents in the rhizosphere, whereas the latter reduced their availability. Total soluble protein content and glutamine synthetase activity were greater in the AWMD × DSB treatment than those in the AWCD × NSB treatment. The activities of superoxide dismutase and catalase were higher in the AWMD × DSB treatment compared with the AWCD × NSB treatment, leading to the amelioration of oxidative cell injury, as shown by a lower malonaldehyde level. This study suggested that farmers should implement AWMD irrigation after leaving the straw residues in the field, followed by deep tillage to improve soil quality and mitigate the drought stress cycles of AWD. This approach can improve rice growth and grain quality and alleviate the problems of disposal of straw residues and water scarcity for sustainable rice production.

The structural characteristic of bamboo shoot shell polysaccharides extracted using ultrasound‐assisted phosphotungstic acid hydrolysis and its protection against cell oxidative injury

SummaryIn this study, crude polysaccharide (BSSP) was extracted from bamboo (Phyllostachys edulis) shoot shell using the ultrasonic‐assisted phosphotungstic acid hydrolysis method. Then, BSSP was further fractionated by DEAE Sepharose fast flow column and serial NaCl solution elutions to obtain two fractions, BSSP‐A and BSSP‐B. The results showed that the maximum yield of BSSP was 195.2 mg g−1 (19.52%) under optimised conditions. Structural analysis indicated that both BSSP‐A and BSSP‐B were heteropolysaccharides with α‐pyranoside bond structure and mainly consisted of xylose by scanning electronic microscopy, Fourier transform infrared spectroscopy, ultraviolet–visible spectroscopy, thermal characterisation and nuclear magnetic resonance spectroscopy. BSSP‐B showed a better protective effect than BSSP‐A against H2O2‐induced oxidative injury in WRL‐68 cells by protecting the cell membrane structure, maintaining mitochondrial membrane potential and increasing intracellular antioxidant enzymes activities. Immunoblotting indicated that polysaccharides might up‐regulate the expression of antioxidant enzymes through the Keap1/Nrf2 signalling pathway and thus alleviate H2O2‐induced cell injury. The findings of this study suggested that polysaccharides from bamboo shoot shell may act as potent antioxidants in functional food industries.

Downregulation of miR-192 Alleviates Oxidative Stress-Induced Porcine Granulosa Cell Injury by Directly Targeting Acvr2a.

Follicular atresia is primarily caused by breakdown to granulosa cells (GCs) due to oxidative stress (OS). MicroRNAs (miRNAs) elicit a defense response against environmental stresses, such as OS, by acting as gene-expression regulators. However, the association between miRNA expression and OS in porcine GCs (PGCs) is unclear. Here, we examined the impact of H2O2-mediated OS in PGCs through miRNA-Seq. We identified 22 (14 upregulated and 8 downregulated) and 33 (19 upregulated and 14 downregulated) differentially expressed miRNAs (DEmiRNAs) at 100 μM and 300 μM H2O2, respectively, compared with the control group. Among the DEmiRNAs, mi-192 was most induced by H2O2-mediated OS, and the downregulation of miR-192 alleviated PGC oxidative injury. The dual-luciferase reporter assay results revealed that miR-192 directly targeted Acvr2a. The Acvr2a level was found to be remarkably decreased after OS. Furthermore, grape seed procyanidin B2 (GSPB2) treatment significantly reduced the H2O2-induced upregulation of miR-192, and decreased PGC apoptosis and oxidative damage. Meanwhile, GSPB2 prevented an H2O2-induced increase in caspase-3 activity, which was enhanced by the application of the miR-192 inhibitor. These results indicate that GSPB2 protects against PGC oxidative injury via the downregulation of miR-192, the upregulation of Acvr2a expression, and the suppression of the caspase-3 apoptotic signaling pathway.

Network pharmacology, molecular docking and in vivo and in vitro experiments to explore the molecular mechanism of licorice green tea beverage to scavenge oxygen free radicals.

Excessive oxygen free radicals can lead to aging, cancer, and other diseases. Therefore, searching for effective antioxidants to scavenge oxygen free radicals has become the focus of modern medicine. In this study, the molecular mechanism of Licorice Green Tea Beverage (LGTB) in scavenging oxygen free radicals was investigated by means of network pharmacology, molecular docking and experimental verification. Network pharmacology studies have shown that paeonol, eugenol, cinnamaldehyde, swertisin, rutin, glycyrrhetinic acid, oleic, pelargonidin-3-O-glucoside and quercetin, kaferempol were the main active components of LGTB, and SOD and CAT are important targets for LGTB in scavenging oxygen free radicals. The results of molecular docking showed that these representative compounds had good affinity to SOD and CAT target proteins. In vitro free radical scavenging experiments showed that LTGB had significant scavenging effects on both DPPH and ABTS radicals, and had strong total reducing power. In vitro cell experiments showed that LGTB could protect HaCaT cells from oxidative stress induced by H2 O2 . The mechanism of LGTB was related to the increase of SOD and CAT activity. Western blotting showed that LGTB could inhibit PI3K/AKT/HIF-1 signaling pathway and improve the antioxidant capacity of HaCaT cells. In vivo experiments showed that LGTB could significantly increase mouse visceral index, increase serum SOD and GSH-Px activity, decrease the content of MDA, and improve liver and kidney pathological state. This study reported the molecular mechanism of LTGB scavenging oxygen free radicals, which provided scientific basis for the treatment and clinical research of aging and other diseases caused by excessive free radicals. PRACTICAL APPLICATIONS: Free radicals are produced by the normal response of cells during aerobic respiration and perform various functions, such as signaling and providing protection against infection. However, excessive free radicals can lead to aging, cancer, and other diseases. The antioxidant can overcome the harm caused by excessive free radicals. In this study, we investigated the molecular mechanism of scavenging oxygen free radicals of Licorice Green Tea Beverage (LGTB) through network pharmacology and molecular docking, and its efficacy was verified by free radical scavenging experiment in vitro, HaCaT cell oxidative stress injury induced by H2 O2 , D-galactose to establish an aging model in mice and Western blotting experiment. It not only elucidates its mechanism at the system level, but also proves its validity at the biological level. It provides the theoretical basis and experimental evidence for the follow-up research and promotion of the product.

TPPU Downregulates Oxidative Stress Damage and Induces BDNF Expression in PC-12 Cells.

Objective Ischemia-reperfusion is an ongoing clinical challenge that can lead to a series of pathological changes including oxidative stress. The inhibition of soluble epoxide hydrolase inhibitor (sEH) by 1-(1-propanoylpiperidin-4-yl)-3-[4-(trifluoromethoxy)phenyl]urea (TPPU) results in an anti-inflammatory, cardioprotective, and blood vessel growth-promoting effects. Therefore, this study focused on the protective effect of TPPU on a rat pheochromocytoma (PC-12) cell oxidative stress model induced by H2O2. Methods CCK-8 and Hoechst 33342 were used to evaluate cell apoptosis and western blot to detect the apoptotic proteins and brain-derived neurotrophic factor (BDNF) expression. Result The incubation with 100 μM, 50 μM, and 25 μM TPPU significantly increased PC-12 cell viability. Epoxyeicosatrienoic acid (EET) pretreatment also protected PC-12 cells from oxidative stress. In addition, TPPU reduced caspase-3 and Bax expression and induced Bcl-2 expression, and EETs exerted the same effect on caspase-3 expression as TPPU. A positive relationship was found between TPPU or EET incubation and BDNF expression. Conclusion These results revealed that TPPU reduced PC-12 cell oxidative stress injury induced by H2O2 and promoted BDNF expression.