H2O2-mediated Oxidative Stress Research Articles

Moisturizing and antioxidant factors of skin barrier restoring cream with shea butter, silkflo and vitamin E in human keratinocyte cells.

Moisturizers are integral to daily skincare routines, reflecting the increasing trend among people towards cosmetic products, particularly for skin care. They significantly contribute to preserving skin health, particularly by regulating the epidermal barrier and moisture levels within the skin. This study aims to explore the moisturizing and antioxidant effect of skin barrier restoring cream Moiz MM (MZ) with shea butter, silkflo and vitamin E by investigating its protective effect against oxidative stress-induced cellular damage and therapeutic mechanisms in human keratinocytes cells (HaCaT). The invitro antioxidant activity of MZ was evaluated by DPPH, ABTS and NO assays. For the cell culture study, HaCaT cells were cultured and stimulated using H2O2 and then treated with different concentrations of MZ. Then, it was subjected to DCFH-DA staining, reverse transcriptase PCR and western blot analysis for the evaluation of various skin-moisture-related components in human keratinocyte cells. Type I procollagen was examined using ELISA technique. The results highlighted that oxidative stress in HaCaT cells decreased type I procollagen synthesis, while MZ treatment significantly increased the synthesis. Moreover, the viability of HaCaT cells was not affected in the presence of MZ, which demonstrates its non-toxic effect. Furthermore, MZ can counteract H2O2-mediated oxidative stress by enhancing the antioxidant enzyme activity such as superoxide dismutase and catalase, and decrease reactive oxygen species generation in skin cells. Additionally, MZ greatly promotes hyaluronic acid production by enhancing the expression of the hyaluronic acid synthase-1 gene and Aquaporin 3 protein. This study suggests that MZ has the potential to serve as a moisturizing and antioxidant skincare formula.

Differential Regulation of Wingless-Wnt/c-Jun N-Terminal Kinase Crosstalk via Oxidative Eustress in Primary and Metastatic Colorectal Cancer Cells.

In the tumor microenvironment (TME), ROS production affects survival, progression, and therapy resistance in colorectal cancer (CRC). H2O2-mediated oxidative stress can modulate Wnt/β-catenin signaling and metabolic reprogramming of the TME. Currently, it is unclear how mild/moderate oxidative stress (eustress) modulates Wnt/β-catenin/APC and JNK signaling relationships in primary and metastatic CRC cells. In this study, we determined the effects of the H2O2 concentration inducing eustress on isogenic SW480 and SW620 cells, also in combination with JNK inhibition. We assessed cell viability, mitochondrial respiration, glycolysis, and Wnt/β-catenin/APC/JNK gene and protein expression. Primary CRC cells were more sensitive to H2O2 eustress combined with JNK inhibition, showing a reduction in viability compared to metastatic cells. JNK inhibition under eustress reduced both glycolytic and respiratory capacity in SW620 cells, indicating a greater capacity to adapt to TME. In primary CRC cells, H2O2 alone significantly increased APC, LEF1, LRP6, cMYC and IL8 gene expression, whereas in metastatic CRC cells, this effect occurred after JNK inhibition. In metastatic but not in primary tumor cells, eustress and inhibition of JNK reduced APC, β-catenin, and pJNK protein. The results showed differential cross-regulation of Wnt/JNK in primary and metastatic tumor cells under environmental eustress conditions. Further studies would be useful to validate these findings and explore their therapeutic potential.

Thiostrepton induces oxidative stress, mitochondrial dysfunction and ferroptosis in HaCaT cells

TST has been mainly studied for its anti-tumor proliferation and antimicrobial effects, but not widely used in dermatological diseases. The mechanism of cellular damage by TST in response to H2O2-mediated oxidative stress was investigated in human skin immortalized keratinocytes (HaCaT) as an in vitro model. The findings reveal that TST treatment leads to increased oxidative stress in the cells by reducing levels of superoxide dismutase (SOD), glutathione (GSH), and catalase (CAT). This effect is further supported by an upsurge in the expression of malondialdehyde (MDA, a pivotal marker of lipid peroxidation). Additionally, dysregulation of FoxM1 at both gene and protein levels corroborates its involvement TST associated effects. Analysis of ferroptosis-related genes confirms dysregulation following TST treatment in HaCaT cells. Furthermore, TST treatment exhibits effects on mitochondrial morphology and function, affirming its induction of apoptosis in the cells through heightened oxidative stress due to mitochondrial damage and dysregulation of mitochondrial membrane potential.

Role of long non-coding RNA Zeb2os in protection of the injured heart

Abstract Heart failure due to ischemic heart disease is one of the leading causes of death worldwide. Terminated vasculature and cardiomyocyte apoptosis due to myocardial infarction are critical events in infarct healing. Therefore, identifying factors that can enhance cardiac repair by restoring the capillary network and inhibiting cardiomyocyte death is of great significance. Accumulating evidence implies that long non-coding RNAs (lncRNAs) are potent regulators of cardiac development and disease. Our preliminary data show the potential cardioprotective function of the long non-coding RNA Zeb2 opposite strand (Zeb2os). This lncRNA overlaps and is antisense to ZEB2 at the 5' splice site of an intron within the 5' UTR. Zeb2os was shown to prevent splicing of the intron containing an internal ribosome entry site (IRES) that increases translation of the ZEB2 transcript. In line with the known function of ZEB2, Zeb2os has pro-angiogenic and anti-apoptotic functions in different organs. However, its role in the heart still has to be determined. Our results show that Zeb2os expression is decreased in cultured cardiomyocytes subjected to H2O2-mediated oxidative stress, which correlates with increased apoptosis and reduced antioxidant defence of cardiac muscle cells. AAV6-mediated overexpression of Zeb2os in hiPSC-derived cardiomyocytes protects from H2O2-induced apoptosis, suggesting it may have cardioprotective activity, possibly through regulating Zeb2 translation. Additionally, systemic AAV9-mediated delivery of Zeb2os to the mouse heart enhanced angiogenesis by increasing the number of proliferating endothelial cells. Moreover, RNA-sequencing of Zeb2os overexpressing hearts at baseline implicated Zeb2os in the regulation of numerous genes related to response to hypoxia. In conclusion, we uncovered/identified Zeb2os as a new potential cardioprotective factor with pro-angiogenic and anti-apoptotic properties. We are currently investigating the effect of AAV9-mediated delivery of Zeb2os in a mouse model of cardiac ischemia to understand further its protective role in the injured heart, which is an essential step towards the development of improved therapies for ischemic heart disease.

Cell-Permeable Stimuli-Responsive Ubiquitin Probe for Time-Resolved Monitoring of Substrate Ubiquitination in Live Cells.

Dynamic monitoring of intracellular ubiquitin (Ub) conjugates is instrumental to understanding the Ub regulatory machinery. Although many biochemical approaches have been developed to characterize protein ubiquitination, chemical tools capable of temporal resolution probing of ubiquitination events remain to be developed. Here, we report the development of the first cell-permeable and stimuli-responsive Ub probe and its application for the temporal resolution profiling of ubiquitinated substrates in live cells. The probe carrying the photolabile group N-(2-nitrobenzyl)-Gly (Nbg) on the amide bond between Ub Gly75 and Gly76 is readily prepared through chemical synthesis and can be delivered to live cells by conjugation via a disulfide bond with the cyclic cell-penetrating peptide cR10D (i.e., 4-((4-(dimethylamino)phenyl)-azo)-benzoic acid-modified cyclic deca-arginine). Both in vitro and in vivo experiments showed that Ub-modifying enzymes (E1, E2s, and E3s) could not install the Ub probe onto substrate proteins prior to removal of the nitrobenzyl group, which was easily accomplished via photoirradiation. The utility and practicality of this probe were exemplified by the time-resolved biochemical and proteomic investigation of ubiquitination events in live cells during a H2O2-mediated oxidative stress response. This work shows a conceptually new family of chemical Ub tools for the time-resolved studies on dynamic protein ubiquitination in different biological processes and highlights the utility of modern chemical protein synthesis in obtaining custom-designed tools for biological studies.

Metabolic proteins with crucial roles in Edwardsiella tarda antioxidative adaptation and intracellular proliferation.

Edwardsiella tarda is a severe fish pathogen, featured by its capacity to live inside host phagocytes. For intracellular survival, it is crucial for E. tarda to neutralize the deleterious effect of host reactive oxygen species (ROS). Accumulating evidence suggests that bacterial metabolism is closely connected to oxidative resistance. However, the roles of E. tarda metabolic proteins in antioxidative adaptation and intracellular proliferation remain elusive. In this study, we performed a proteomic analysis on E. tarda and identified 111 proteins responsive to H2O2-mediated oxidative stress. Based on this data, we further obtained eight crucial proteins, including seven metabolic proteins, for E. tarda antioxidation and intracellular infection. Among them, two C4-dicarboxylate transporters were found necessary for E. tarda to disseminate in fish tissues. Furthermore, the substrate of the two transporters was identified as L-aspartate, which was proven to be essential for the full antioxidative capacity of E. tarda. Our results indicate that reprogramming the metabolic flux to the production of pyruvate, a ketoacid capable of neutralizing ROS, was likely a pivotal strategy of E. tarda to survive the oxidative environments inside host cells. Together, the findings of this study highlight the significance of metabolic reprogramming for bacterial redox homeostasis and intracellular infection. IMPORTANCE Edwardsiella tarda is a significant fish pathogen that can live in challenging environments of reactive oxygen species (ROS), such as inside the phagocytes. Metabolic reconfiguration has been increasingly associated with bacterial oxidative tolerance and virulence. However, the metabolic proteins of E. tarda involved in such processes remain elusive. By proteomic analysis and functional characterization of protein null mutants, the present study identified eight crucial proteins for bacterial oxidative resistance and intracellular infection. Seven of them are metabolic proteins dictating the metabolic flux toward the generation of pyruvate, a key metabolite capable of scavenging ROS molecules. Furthermore, L-aspartate uptake, which can fuel the pyruvate generation, was found essential for the full antioxidative capacity of E. tarda. These findings identified seven metabolic proteins involved in bacterial oxidative adaptation and indicate that metabolic reprogramming toward pyruvate was likely a pivotal strategy of bacteria for antioxidative adaptation and intracellular survival.

Protection of H9c2 Myocardial Cells from Oxidative Stress by Crocetin via PINK1/Parkin Pathway-Mediated Mitophagy.

This study aimed to explore the oxidative stress-protective effect of crocetin on H2O2-mediated H9c2 myocardial cells through in vitro experiments, and further explore whether its mechanism is related to the impact of mitophagy. This study also aimed to demonstrate the therapeutic effect of safflower acid on oxidative stress in cardiomyocytes and explore whether its mechanism is related to the effect of mitophagy. Here, an H2O2-based oxidative stress model was constructed and assessed the degree of oxidative stress injury of cardiomyocytes by detecting the levels of lactate dehydrogenase (LDH), creatine kinase (CK), malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH Px). Reactive oxygen species (ROS)-detecting fluorescent dye DCFH-DA, JC-1 dye, and TUNEL dye were employed to assess mitochondrial damage and apoptosis. Autophagic flux was measured by transfecting Ad-mCherry-GFP-LC3B adenovirus. Mitophagy-related proteins were then detected via western blotting and immunofluorescence. However, crocetin (0.1-10 µM) could significantly improve cell viability and reduce apoptosis and oxidative stress damage caused by H2O2. In cells with excessive autophagic activation, crocetin could also reduce autophagy flow and the expression of mitophagy-related proteins PINK1 and Parkin, and reverse the transfer of Parkin to mitochondria. Crocetin could reduce H2O2-mediated oxidative stress damage and the apoptosis of H9c2 cells, and its mechanism was closely related to mitophagy.

Pseudomonas aeruginosa Dps (PA0962) Functions in H2O2 Mediated Oxidative Stress Defense and Exhibits In Vitro DNA Cleaving Activity.

We report the structural, biochemical, and functional characterization of the product of gene PA0962 from Pseudomonas aeruginosa PAO1. The protein, termed Pa Dps, adopts the Dps subunit fold and oligomerizes into a nearly spherical 12-mer quaternary structure at pH 6.0 or in the presence of divalent cations at neutral pH and above. The 12-Mer Pa Dps contains two di-iron centers at the interface of each subunit dimer, coordinated by conserved His, Glu, and Asp residues. In vitro, the di-iron centers catalyze the oxidation of Fe2+ utilizing H2O2 (not O2) as an oxidant, suggesting Pa Dps functions to aid P. aeruginosa to survive H2O2-mediated oxidative stress. In agreement, a P. aeruginosa Δdps mutant is significantly more susceptible to H2O2 than the parent strain. The Pa Dps structure harbors a novel network of Tyr residues at the interface of each subunit dimer between the two di-iron centers, which captures radicals generated during Fe2+ oxidation at the ferroxidase centers and forms di-tyrosine linkages, thus effectively trapping the radicals within the Dps shell. Surprisingly, incubating Pa Dps and DNA revealed unprecedented DNA cleaving activity that is independent of H2O2 or O2 but requires divalent cations and 12-mer Pa Dps.

Molecular characterization and expression profiling of caveolin-1 from Amphiprion clarkii and elucidation of its involvement in antiviral response and redox homeostasis.

Caveolin-1 (Cav-1), a major structural component of caveolae, is involved in various biological functions, such as endocytosis, cholesterol trafficking, transcytosis, signal transduction, and immunity. To date, three caveolin members have been identified in mammals: Cav-1, Cav-2, and Cav-3. In this study, we identified the Cav-1 sequence from Amphiprion clarkii (AcCav-1). The protein is 181 amino acids long, with a molecular weight of 20.73kDa and a predicted isoelectric point of 5.48. The phylogenetic tree disclosed that AcCav-1 is closely related to teleost fish orthologs and clusters together with vertebrates. It shares the highest identity (99.4%) and similarity (100%) with Amphiprion ocellaris. Subcellular localization assays showed that Cav-1 expressed in the endoplasmic reticulum and cytoplasm. Further, AcCav-1 was ubiquitously expressed in all examined tissues, but most highly in the skin and the spleen. The up and downregulation of AcCav-1 was observed throughout the testing period after in-vivo immunostimulation with lipopolysaccharides (LPS), polyinosinic:polycytidylic acid (poly (I:C), and Vibrio harveyi (V. harveyi). The antiviral assay showed that AcCav-1 overexpression suppresses the replication of the viral hemorrhagic septicemia virus (VHSV) in Fathead minnow cells by activating antiviral genes. Further, LPS induced NO production and H2O2-mediated oxidative stress assays showed that AcCav-1 is involved in the regulation of oxidative stress. Collectively, these findings suggest the indispensable role of Cav-1 in the immune system of A.clarkii.

Number 2 Feibi Recipe Ameliorates Pulmonary Fibrosis by Inducing Autophagy Through the GSK-3β/mTOR Pathway.

Number 2 Feibi Recipe (N2FBR) is a traditional Chinese medicine formula for treating idiopathic pulmonary fibrosis. N2FBR inhibits H2O2-mediated oxidative stress damage in alveolar epithelial cells by increasing autophagy, as we previously demonstrated. However, it is unknown if similar mechanisms occur in vivo. We established a pulmonary fibrosis model by instilling bleomycin (BLM) from the airway to examine the effects of N2FBR on pulmonary fibrosis and investigate its probable mechanism in this work. We discovered that N2FBR treatment effectively alleviated interstitial fibrosis as well as collagen deposition, primarily in upregulating SOD, GSH-Px, T-AOC and downregulating MDA content. N2FBR also increased the expression of LC3B, Beclin-1, LAMP1, TFEB and downregulated the expression of p62, legumain. N2FBR treatment boosted the production of autophagosomes, according to the results of the TEM observation. Furthermore, we explored that N2FBR exerted its anti-oxidative stress and pro-autophagy effects via GSK-3β/mTOR signalling pathway. Therefore, these results provide further evidence for the protective effect of N2FBR in pulmonary fibrosis. Our findings could have ramifications for the development of antifibrosis therapies.

Role of SPAK\u2013NKCC1 signaling cascade in the choroid plexus blood\u2013CSF barrier damage after stroke

BackgroundThe mechanisms underlying dysfunction of choroid plexus (ChP) blood–cerebrospinal fluid (CSF) barrier and lymphocyte invasion in neuroinflammatory responses to stroke are not well understood. In this study, we investigated whether stroke damaged the blood–CSF barrier integrity due to dysregulation of major ChP ion transport system, Na+–K+–Cl− cotransporter 1 (NKCC1), and regulatory Ste20-related proline-alanine-rich kinase (SPAK).MethodsSham or ischemic stroke was induced in C57Bl/6J mice. Changes on the SPAK–NKCC1 complex and tight junction proteins (TJs) in the ChP were quantified by immunofluorescence staining and immunoblotting. Immune cell infiltration in the ChP was assessed by flow cytometry and immunostaining. Cultured ChP epithelium cells (CPECs) and cortical neurons were used to evaluate H2O2-mediated oxidative stress in stimulating the SPAK–NKCC1 complex and cellular damage. In vivo or in vitro pharmacological blockade of the ChP SPAK–NKCC1 cascade with SPAK inhibitor ZT-1a or NKCC1 inhibitor bumetanide were examined.ResultsIschemic stroke stimulated activation of the CPECs apical membrane SPAK–NKCC1 complex, NF-κB, and MMP9, which was associated with loss of the blood–CSF barrier integrity and increased immune cell infiltration into the ChP. Oxidative stress directly activated the SPAK–NKCC1 pathway and resulted in apoptosis, neurodegeneration, and NKCC1-mediated ion influx. Pharmacological blockade of the SPAK–NKCC1 pathway protected the ChP barrier integrity, attenuated ChP immune cell infiltration or neuronal death.ConclusionStroke-induced pathological stimulation of the SPAK–NKCC1 cascade caused CPECs damage and disruption of TJs at the blood–CSF barrier. The ChP SPAK–NKCC1 complex emerged as a therapeutic target for attenuating ChP dysfunction and lymphocyte invasion after stroke.

Molecular characterization of heat shock protein 20 (hsp20) in goldfish (Carassius auratus) and expression analysis in response to environmental stresses

Heat shock proteins (Hsps) are described as molecular chaperones that were synthesized rapidly in response to various stress conditions in many organisms. However, information of small heat shock protein (sHsp) in response to environmental stresses is lacking in fish. In this study, the full length cDNA of hsp20 in goldfish, Carassius auratus, with a 65 bp 5′-untranslated region (UTR), a 563 bp 3′-UTR, and a 450 bp open reading frame encoding 149 amino acids, was obtained. gfhsp20 contained a highly conserved α-crystallin domain. Consistent with humans and zebrafish, goldfish HSP20 gene had three exons and two introns. gfhsp20 mRNA were detected by quantitative real-time PCR in all of the tissues examined, with the highest level in the muscle, followed by the skin, heart, intestine, liver, spleen, brain, kidney, gill and head kidney. gfhsp20 mRNA was significantly induced with heat shock in liver and spleen, but decreased in brain. Under H2O2-mediated oxidative stress, the expression level of gfhsp20 was significantly up-regulated in brain, liver and spleen. After LPS and PolyI:C infection, the expression level of gfhsp20 in spleen was significantly up-regulated with a higher increase under PolyI:C infection than that of LPS injection. These results revealed the potential function of gfHsp20 in response to heat shock, oxidative stress and infectious pathogens.

Number 2 Feibi Recipe Inhibits H2O2-Mediated Oxidative Stress Damage of Alveolar Epithelial Cells by Regulating the Balance of Mitophagy/Apoptosis

Reactive oxygen species (ROS)-mediated alveolar epithelial cell (AEC) injury and apoptosis are considered to be the initiating link of idiopathic pulmonary fibrosis (IPF), and protecting AECs can alleviate IPF. This study aimed to explore the protective effect of number 2 Feibi recipe (FBR-2) medicated serum on H2O2-mediated oxidative stress injury in AECs and further explore its mechanism. We found that FBR-2 can regulate downstream antioxidant enzymes expression by activating nuclear factor erythroid 2-related factor 2 (Nrf2), reducing the level of intracellular ROS, protecting mitochondrial function and improving cell survival. FBR-2 can also activate mitophagy through the PINK1/Parkin pathway. Moreover, FBR-2 can inhibit apoptosis by blocking the mitochondrial apoptosis mechanism. In summary, these data indicate that FBR-2 medicated serum can inhibit H2O2-mediated oxidative stress damage in AECs by regulating the balance of mitophagy/apoptosis. This study provides new evidence for the antifibrotic effect of FBR-2 and provides new drug candidates for the clinical treatment of IPF.

Protection of Human Lens Epithelial Cells from Oxidative Stress Damage and Cell Apoptosis by KGF-2 through the Akt/Nrf2/HO-1 Pathway.

Oxidative stress exerts a significant influence on the pathogenesis of various cataracts by inducing degradation and aggregation of lens proteins and apoptosis of lens epithelial cells. Keratinocyte growth factor−2 (KGF-2) exerts a favorable cytoprotective effect against oxidative stress in vivo and in vitro. In this work, we investigated the molecular mechanisms of KGF-2 against hydrogen peroxide- (H2O2-) induced oxidative stress and apoptosis in human lens epithelial cells (HLECs) and rat lenses. KGF-2 pretreatment could reduce H2O2-induced cytotoxicity as well as reactive oxygen species (ROS) accumulation. KGF-2 also increases B-cell lymphoma-2 (Bcl-2), quinine oxidoreductase-1 (NQO-1), superoxide dismutase (SOD2), and catalase (CAT) levels while decreasing the expression level of Bcl2-associated X (Bax) and cleaved caspase-3 in H2O2-stimulated HLECs. LY294002, the phosphatidylinositol-3-kinase (PI3K)/Akt inhibitor, abolished KGF-2's effect to some extent, demonstrating that KGF-2 protected HLECs via the PI3K/Akt pathway. On the other hand, KGF-2 activated the Nrf2/HO-1 pathway by regulating the PI3K/Akt pathway. Silencing nuclear factor erythroid 2-related factor 2 (Nrf2) by targeted-siRNA and inhibiting heme oxygenase-1 (HO-1) through zinc protoporphyrin IX (ZnPP) significantly decreased cytoprotection of KGF-2. Furthermore, as revealed by lens organ culture assays, KGF-2 treatment decreased H2O2-induced lens opacity in a concentration-dependent manner. As demonstrated by these data, KGF-2 resisted H2O2-mediated apoptosis and oxidative stress in HLECs through Nrf2/HO-1 and PI3K/Akt pathways, suggesting a potential protective effect against the formation of cataracts.

Disruption of metabolic function and redox homeostasis as antibacterial mechanism of Lindera glauca fruit essential oil against Shigella flexneri

Current trend in food safety and public health is focusing on the use of natural antimicrobials. The Lindera glauca fruit with rich resource and essential oil has emerged as potential application in China. This work aimed to detect chemical compounds of L. glauca fruit essential oil (LGFEO) and to evaluate antimicrobial activity and mechanism of LGFEO for develop natural antimicrobials. A total of 70 volatile compounds were identified, and the major groups were monoterpene (56.62%) and sesquiterpene (39.71%). The analysis of antimicrobial activity indicated that LGFEO could effectively inhibit the growth of all tested 13 food-borne pathogens, of which Shigella flexneri had the highest inhibition zone diameter (DIZ, 25.45 nm), and the lowest minimum inhibitory concentration (MIC, 0.156 μg/mL) and bactericidal concentration (MBC, 0.312 μg/mL). To unravel antibacterial mechanism of LGFEO on S. flexneri , the dynamic changes for bacterial growth curve, material release, and the levels of intracellular mallondialdehyde (MDA), hydrogen peroxide (H 2 O 2 ), and ROS-detoxifying enzymes were analyzed in S. flexneri cells exposed to LGFEO at different concentrations (1/2 × MIC, 1 × MIC and 2 × MIC) and times (0–24 h), revealing that LGFEO could cause cell material leakage, antioxidant enzyme inactivation, H 2 O 2 and MDA accumulation, and bacterial growth inhibition. Also, antibacterial action of LGFEO was conducted on key enzymes of respiratory metabolic (glycolysis, oxidative pentose phosphate pathway, tricarboxylic acid cycle, and respiratory chain), and the levels of intracellular ATP, ADP, nicotinamide adenine dinucleotide (NAD + and NADH), nicotinamide adenine dinucleotide phosphate (NADP + and NADPH), and oxidized and reduced glutathione (GSSG and GSH) in LGFEO-treated S. flexneri cells, indicating a role of LGFEO in repressing respiratory pathway and electron transport with low redox potential. All these revealed that LGFEO could effectively induce H 2 O 2 accumulation and lipid peroxidation, resulting in cell membrane damage and metabolic function disruption, giving rise to a disturbance of redox homeostasis, which may be the main cause of growth inhibition or cell death for S. flexneri . • Essential oil contains 70 volatile compounds with good antimicrobial activity. • The first time for antibacterial mechanism from energy and respiratory metabolism. • Antibacterial action of essential oil involves in H2O2-mediated oxidative stress. •Essential oil-induced disruption of redox homeostasis as main antibacterial action.

H2O2-Mediated Oxidative Stress Enhances Cystathionine γ-Lyase-Derived H2S Synthesis via a Sulfenic Acid Intermediate.

Hydrogen sulfide (H2S), which is generated mainly by cystathionine γ-lyase (CSE) in the cardiovascular system, plays a pivotal role in a wide range of physiological and pathological processes. However, the regulatory mechanism of the CSE/H2S system is poorly understood. Herein, we show that oxidation induces the disulfide bond formation between Cys252 and Cys255 in the CXXC motif, thus stimulating the H2S-producing activity of CSE. The activity of oxidized CSE is approximately 2.5 fold greater than that of the reduced enzyme. Molecular dynamics and molecular docking suggest that the disulfide bond formation induces the conformational change in the active site of CSE and consequently increases the affinity of the enzyme for the substrate L-cysteine. Mass spectrometry and mutagenesis studies further established that the residue Cys255 is crucial for oxidation sensing. Oxidative stress-mediated sulfenylation of Cys255 leads to a sulfenic acid intermediate that spontaneously forms an intramolecular disulfide bond with the vicinal thiol group of Cys252. Moreover, we demonstrate that exogenous hydrogen peroxide (H2O2) and endogenous H2O2 triggered by vascular endothelial growth factor (VEGF) promote cellular H2S production through the enhancement of CSE activity under oxidative stress conditions. By contrast, incubation with H2O2 or VEGF did not significantly enhance cellular H2S production in the presence of PEG-catalase, an enzymatic cell-permeable H2O2 scavenger with high H2O2 specificity. Taken together, we report a new posttranslational modification of CSE that provides a molecular mechanism for H2O2/H2S crosstalk in cells under oxidative stress.

Tamarix articulata Extracts Exhibit Antioxidant Activity and Offer Protection against Hydrogen Peroxide-Mediated Toxicity to Human Skin Fibroblasts.

Tamarix articulata (TA) is a wild halophytic plant growing in extremely harsh environmental conditions in the deserts of Saudi Arabia. Evaluating the protective effect of the methanolic extract of different parts (fresh and dry leaves, stem, and root) of TA was determined by MTT assay using Hs27 skin fibroblasts as the cellular model. The study was designed and conducted in two sets. The first set assesses the toxicity profile of TA extracts in both concentration- and time-dependent ways on Hs27 cells. Our MTT results showed that methanolic extracts from all four parts of TA at varying doses (27.5, 55, 110, and 220 μg/mL) display negligible toxicity when exposed for 4 h. However, exposure of Hs27 cells to varying doses of all four TA extracts for 24 and 48 h promotes significant 23%, 24%, 26%, and 25% (p < 0.05) and 35%, 36%, 39%, and 41% (p < 0.05) cell toxicity at 220 μg/mL of all four TA extracts compared to untreated control cells. To evaluate the protection offered by TA extracts against H₂O₂, we perform a second set of experiments to preincubate Hs27 cells with the TA extracts in both dose- and time-dependent way. This is followed by 300 μM hydrogen peroxide- (H₂O₂-) mediated oxidative insult for 1 h. Using MTT assay, we found that methanolic extracts of TA at different time points (4, 24, and 48 h) and higher doses (220 μg/mL) provide significant protection in cell viability when challenged with H2O2-induced oxidative stress in Hs27 cells. The protective effect was more pronounced at 48 h and 220 μg/mL and the amounts were 39%, 41%, 41%, and 44% for stem, root, fresh leaf, and dry leaf TA extracts (p < 0.05), respectively, compared to untreated cells (2–4%). Collectively, the current study demonstrates that methanolic extracts of TA contain potential bioactive compounds and offer significant protection against H2O2-mediated oxidative stress in Hs27 skin fibroblasts.

Autophagy Contributes to Oxidative Stress-Induced Apoptosis in Porcine Granulosa Cells.

Oxidative stress-induced granulosa cell (GC) death is a major cause of follicular atresia. As the major types of programmed cell death, autophagy and apoptosis have been observed in response to H2O2-mediated oxidative stress and have been demonstrated to be responsible for porcine GC death. To date, however, the cellular reactions linking autophagy to the apoptosis of porcine GC under oxidative stress are still poorly understood. Porcine GC were treated with H2O2, and autophagic flux was examined by western blotting. Cell viability and cell death assays were performed after cotreatment of porcine GC with autophagy activator (rapamycin) or inhibitor (3-methyladenine, 3-MA) together with H2O2. We revealed that short exposure (1-3h) of porcine GC to H2O2 dramatically increased autophagic flux (1.8- to 2.5-fold over that in the control), whereas 6-12h prolonged treatment decreased autophagy but elevated the caspase-3 activity and GC apoptotic rate. Furthermore, we showed that pretreatment with rapamycin exacerbated H2O2-mediated cytotoxicity and caspase-3 activation but that 3-MA or siRNAs specific for Beclin 1 and Atg7 genes ameliorated H2O2-mediated GC apoptosis. Together, our results indicate that autophagy plays a pivotal role in H2O2-mediated porcine GC apoptosis. Importantly, we show that the early induction of autophagic flux contributes to oxidative stress-induced apoptosis in porcine GC. The results also suggest that regulating the autophagy response in porcine GC under oxidative stress might be a new strategy for abnormal follicular atresia.

DL0410 attenuates oxidative stress and neuroinflammation via BDNF/TrkB/ERK/CREB and Nrf2/HO-1 activation

Oxidative stress and neuroinflammation have been deeply associated with Alzheimer’s disease. DL0410 is a novel acetylcholinesterase inhibitor with potential anti-oxidative effects in AD-related animal models, while the specific mechanism has not been fully clarified. In this study, DL0410 was predicted to be related to the modification of cell apoptosis, oxidation-reduction process, inflammatory response and ERK1/ERK2 cascade by in silico target fishing and GO enrichment analysis. Then the possible protective effects of DL0410 were evaluated by hydrogen peroxide (H2O2)-induced oxidative stress model and lipopolysaccharides (LPS)-induced neuroinflammation model H2O2 decreased the viability of SH-SY5Y cells, induced malondialdehyde (MDA) accumulation, mitochondrial membrane potential (Δψm) loss and cell apoptosis, which could be reversed by DL0410 dose-dependently, indicating that DL0410 protected SH-SY5Y cells against H2O2-mediated oxidative stress. Western blot analysis showed that DL0410 increased the H2O2-triggered down-regulated TrkB, ERK and CREB phosphorylation and the expression of BDNF. In addition, TrkB inhibitor ANA-12, ERK inhibitor SCH772984 and CREB inhibitor 666-15 eliminated the inhibition of DL0410 on MDA accumulation and Δψm loss. Furthermore, DL0410 attenuates inflammatory responses and ROS production in LPS-treated BV2 cells, which is responsible for Nrf2 and HO-1 up-regulation. The present study demonstrates that DL0410 is a potential activator of the BDNF/TrkB/ERK/CREB and Nrf2/HO-1 pathway and may be a potential candidate for regulating oxidative stress and neuroinflammatory response in the brain. Together, the results showed that DL0410 is a promising drug candidate for treating AD and possibly other nervous system diseases associated with oxidative stress and neuroinflammation.

Influence of Tooth Pigmentation on H2O2 Diffusion and Its Cytotoxicity After In-office Tooth Bleaching.

Pigments in tooth structures affect the diffusion of H2O2 through enamel and dentin. The bleaching methodology can be impacted. Objective: The aim of this study was to evaluate the influence of the presence of pigments in tooth structures on the trans-enamel and trans-dentin diffusion of hydrogen peroxide (H2O2) and its cytotoxicity after carrying out an in-office bleaching therapy.Methods and Materials: A bleaching gel with 35% H2O2 was applied for 45 minutes (three times for 15 minutes) on enamel and dentin discs (n=6), either previously submitted to the intrinsic pigmentation protocol with a concentrated solution of black tea, or not, defining the following groups: G1, unbleached untreated discs (control 1); G2, unbleached pigmented discs (control 2); G3, bleached untreated discs; G4, bleached pigmented discs. The discs were adapted to artificial pulp chambers, which were placed in wells of 24-well plates containing 1 mL culture medium (Dulbecco's modified Eagle's medium [DMEM]). After applying the bleaching gel on enamel, the extracts (DMEM + components of bleaching gel that diffused through the discs) were collected and then applied on the cultured MDPC-23 odontoblast-like cells. Cell viability (methyl tetrazolium assay and Live & Dead, Calcein AM, and ethidium homodimer-1 [EthD-1] probes), the amount of H2O2 that diffused through enamel and dentin (leuco-crystal violet product), and the H2O2-mediated oxidative cell stress (SOx) and components of degradation were assessed (analysis of variance/Tukey; α=0.05).Results: There was no significant difference between the groups G1 and G2 for all the parameters tested (p>0.05). Reduction in the trans-enamel and trans-dentin diffusion of H2O2 occurred for G4 in comparison with G3. Significantly lower cell viability associated with greater oxidative stress was observed for G3 (p<0.05). Therefore, in-office tooth bleaching therapy performed in pigmented samples caused lower cytotoxic effects compared with untreated samples submitted to the same esthetic procedure (p<0.05).Conclusion: According to the methodology used in this investigation, the authors concluded that the presence of pigments in hard tooth structures decreases the trans-enamel and trans-dentin diffusion of H2O2 and the toxicity to pulp cells of an in-office bleaching gel with 35% H2O2.