Activating Nicotinamide Adenine Dinucleotide Phosphate Research Articles

A homologous membrane-camouflaged self-assembled nanodrug for synergistic antitumor therapy

Limited success has been achieved in ferroptosis-induced cancer treatment due to the challenges related to low production of toxic reactive oxygen species (ROS) and inherent ROS resistance in cancer cells. To address this issue, a self-assembled nanodrug have been investigated that enhances ferroptosis therapy by increasing ROS production and reducing ROS inhibition. The nanodrug is constructed by allowing doxorubicin (DOX) to interact with Fe2+ through coordination interactions, forming a stable DOX-Fe2+ chelate, and this chelate further interacts with sorafenib (SRF), resulting in a stable and uniform nanoparticle. In tumor cells, overexpressed glutathione (GSH) triggers the disassembly of nanodrug, thereby activating the drug release. Interestingly, the released DOX not only activates nicotinamide adenine dinucleotide phosphate oxidase 4 (NOX4) to produce abundant H2O2 production for enhanced ROS production, but also acts as a chemotherapeutics agent, synergizing with ferroptosis. To enhance tumor selectivity and improve the blood clearance, the nanodrug is coated with a related cancer cell membrane, which enhances the selective inhibition of tumor growth and metastasis in a B16F10 mice model. Our findings provide valuable insights into the rational design of self-assembled nanodrug for enhanced ferroptosis therapy in cancer treatment. Statement of significanceFerroptosis is a non-apoptotic form of cell death induced by the iron-regulated lipid peroxides (LPOs), offering a promising potential for effective and safe anti-cancer treatment. However, two significant challenges hinder its clinical application: 1) The easily oxidized nature of Fe2+ and the low concentration of H2O2 leads to a low efficiency of intracellular Fenton reaction, resulting in poor therapeutic efficacy; 2) The instinctive ROS resistance of cancer cells induce drug resistance. Therefore, we developed a simple and high-efficiency nanodrug composed of self-assembling by Fe2+ sources, H2O2 inducer and ROS resistance inhibitors. This nanodrug can effectively deliver the Fe2+ sources into tumor tissue, enhance intracellular concentration of H2O2, and reduce ROS resistance, achieving a high-efficiency, precise and safe ferroptosis therapy.

Empagliflozin prevents oxidative stress in human coronary artery endothelial cells via the NHE/PKC/NOX axis

BackgroundEmpagliflozin (EMPA) ameliorates reactive oxygen species (ROS) generation in human endothelial cells (ECs) exposed to 10 % stretch, but the underlying mechanisms are still unclear. Pathological stretch is supposed to stimulate protein kinase C (PKC) by increasing intracellular calcium (Ca2+), therefore activating nicotinamide adenine dinucleotide phosphate oxidase (NOX) and promoting ROS production in human ECs. We hypothesized that EMPA inhibits stretch-induced NOX activation and ROS generation through preventing PKC activation. MethodsHuman coronary artery endothelial cells (HCAECs) were pre-incubated for 2 h before exposure to cyclic stretch (5 % or 10 %) with either vehicle, EMPA or the PKC inhibitor LY-333531 or PKC siRNA. PKC activity, NOX activity and ROS production were detected after 24 h. Furthermore, the Ca2+ chelator BAPTA-AM, NCX inhibitor ORM-10962 or NCX siRNA, sodium/potassium pump inhibitor ouabain and sodium hydrogen exchanger (NHE) inhibitor cariporide were applied to explore the involvement of the NHE/Na+/NCX/Ca2+ in the ROS inhibitory capacity of EMPA. ResultsCompared to 5 % stretch, 10 % significantly increased PKC activity, which was reduced by EMPA and PKC inhibitor LY-333531. EMPA and LY-333531 showed a similar inhibitory capacity on NOX activity and ROS generation induced by 10 % stretch, which was not augmented by combined treatment with both drugs. PKC-β knockdown inhibits the NOX activation induced by Ca2+ and 10 % stretch. BAPTA, pharmacologic or genetic NCX inhibition and cariporide reduced Ca2+ in static HCAECs and prevented the activation of PKC and NOX in 10%-stretched cells. Ouabain increased ROS generation in cells exposed to 5 % stretch. ConclusionEMPA reduced NOX activity via attenuation of the NHE/Na+/NCX/Ca2+/PKC axis, leading to less ROS generation in HCAECs exposed to 10 % stretch.

Epicardial Adipose Tissue-Derived Leptin Promotes Myocardial Injury in Metabolic Syndrome Rats Through PKC/NADPH Oxidase/ROS Pathway.

Background The epicardial adipose tissue (EAT) of metabolic syndrome (MetS) is abnormally accumulated with dysfunctional secretion of adipokines, closely relating to cardiac dysfunction. The current study was designed to identify the effects of EAT-derived leptin on the myocardium of MetS rats and explore the potential molecular mechanisms. Methods and Results A MetS rat model was established in 8-week-old Wistar rats by a 12-week high-fat diet. MetS rats exhibited increased leptin secretion from EAT, cardiac hypertrophy, and diastolic dysfunction with preserved systolic function. The myocardium of MetS rats had abnormal structure, increased oxidative stress injury, and higher inflammatory factor levels, especially the subepicardial myocardium, which was correlated with the EAT-derived leptin level but not the serum leptin. The EAT was separated from each group of rats to prepare EAT-conditioned medium. H9C2 rat cardiomyoblasts were treated with EAT-conditioned medium or leptin, plus various inhibitors. EAT-derived leptin from MetS rats promoted mitochondrial oxidative stress and dysfunction, induced mitochondrial pathway apoptosis, and inhibited cell viability in H9C2 cardiomyoblasts via the protein kinase C/reduced nicotinamide adenine dinucleotide phosphate oxidase/reactive oxygen species (PKC/NADPH oxidase/ROS) pathway. EAT-derived leptin from MetS rats stimulated inflammation in H9C2 cardiomyocytes by promoting activator protein 1 nuclear translocation via the PKC/NADPH oxidase/ROS pathway. Leptin promoted the interaction between p-p47phox and gp91phox in H9C2 cardiomyocytes via protein kinase C, activating nicotinamide adenine dinucleotide phosphate oxidase, increasing reactive oxygen species generation, and inhibiting cell viability. Conclusions EAT-derived leptin induces MetS-related myocardial injury through the following 2 cooperative ways via PKC/NADPH oxidase/ROS pathway: (1) inducing mitochondrial pathway apoptosis by promoting mitochondrial oxidative stress and dysfunction; and (2) stimulating inflammation by promoting activator protein 1 nuclear translocation.

Stretch-induced reactive oxygen species contribute to the Frank-Starling mechanism.

Myocardial stretch physiologically activates NADPH oxidase 2 (NOX2) to increase reactive oxygen species (ROS) production. Although physiological low-level ROS are known to be important as signalling molecules, the role of stretch-induced ROS in the intact myocardium remains unclear. To address this, we investigated the effects of stretch-induced ROS on myocardial cellular contractility and calcium transients in C57BL/6J and NOX2-/- mice. Axial stretch was applied to the isolated cardiomyocytes using a pair of carbon fibres attached to both cell ends to evaluate stretch-induced modulation in the time course of the contraction curve and calcium transient, as well as to evaluate maximum cellular elastance, an index of cellular contractility, which is obtained from the end-systolic force-length relationship. In NOX2-/- mice, the peak calcium transient was not altered by stretch, as that in wild-type mice, but the lack of stretch-induced ROS delayed the rise of calcium transients and reduced contractility. Our mathematical modelling studies suggest that the augmented activation of ryanodine receptors by stretch-induced ROS causes a rapid and large increase in the calcium release flux, resulting in a faster rise in the calcium transient. The slight increase in the magnitude of calcium transients is offset by a decrease in sarcoplasmic reticulum calcium content as a result of ROS-induced calcium leakage, but the faster rise in calcium transients still maintains higher contractility. In conclusion, a physiological role of stretch-induced ROS is to increase contractility to counteract a given preload, that is, it contributes to the Frank-Starling law of the heart. KEY POINTS: Myocardial stretch increases the production of reactive oxygen species by NADPH oxidase 2. We used NADPH oxidase 2 knockout mice to elucidate the physiological role of stretch-induced reactive oxygen species in the heart. We showed that stretch-induced reactive oxygen species modulate the rising phase of calcium transients and increase myocardial contractility. A mathematical model simulation study demonstrated that rapid activation of ryanodine receptors by reactive oxygen species is important for increased contractility. This response is advantageous for the myocardium, which must contract against a given preload.

A Cell Selective Fluoride-Activated MOF Biomimetic Platform for Prodrug Synthesis and Enhanced Synergistic Cancer Therapy.

As a burgeoning bioorthogonal reaction, the fluoride-mediated desilylation is capable of prodrug activation. However, due to the reactions lack of cell selectivity and unitary therapy modality, this strongly impedes their biomedical applications. Herein, we construct a cancer cell-selective biomimetic metal-organic framework (MOF)-F platform for prodrug activation and enhanced synergistic chemodynamic therapy (CDT). With cancer cell membranes camouflage, the designed biomimetic nanocatalyst displays preferential accumulation to homotypic cancer cells. Then, pH-responsive nanocatalyst releases fluoride ions and ferric ions. For activation of our designed prodrug tert-butyldimethyl silyl (TBS)-hydroxycamptothecin (TBSO-CPT), fluoride ions can desilylate TBS and cleave the designed silyl ether linker to synthesize the OH-CPT (10-hydroxycamptothecin) drug molecule, which effectively kills cancer cells. Intriguingly, the bioorthogonal-synthesized OH-CPT drug upregulates intracellular H2O2 by activating nicotinamide adenine dinucleotide phosphate oxidase (NOX), amplifying the released iron induced Fenton reaction for synergistic CDT. Both in vitro and in vivo studies demonstrate our strategy presents a versatile fluoride-activated bioorthogonal catalyst for cancer cell-selective drug synthesis. Our work may accelerate the biomedical applications of fluoride-activated bioorthogonal chemistry.

Cancer Cell Membrane Biomimetic Mesoporous Nanozyme System with Efficient ROS Generation for Antitumor Chemoresistance.

Single-atom nanozymes (SAZs) with reaction specificity and optimized catalytic properties have great application prospects in tumor therapy. But the complex tumor microenvironment (low content of H2O2) limits its therapeutic effect. In this study, we developed a bionic mesoporous Fe SAZs/DDP nanosystem (CSD) for enhanced nanocatalytic therapy (NCT)/chemotherapy by simultaneously encapsulating the chemotherapeutic drugs cisplatin (DDP) and Fe SAZs with high peroxidase (POD) activity into the cancer cell membrane. CSD could evade immune recognition and actively targets tumor sites, and DDP upregulates endogenous H2O2 levels by activating nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, thereby enhancing SAZs-mediated hydroxyl radical (·OH) production, which subsequently leads to mitochondrial damage and intolerance to chemotherapy drug. We used the HGC27/DDP cell line for in vitro and in vivo experiments. The results showed that CSD achieved good therapeutic benefits, without any side effects such as inflammatory reaction. This system can induce multiple antitumor effect with H2O2 self-supply, mitochondrial damage, and ATP downregulation and eventually lead to chemosensitization.

CuS nanoparticles and camptothecin co-loaded thermosensitive injectable hydrogel with self-supplied H2O2 for enhanced chemodynamic therapy.

Chemodynamic therapy (CDT) is a kind of anti-tumor strategy emerging in recent years, but the concentration of hydrogen peroxide (H2O2) in the tumor microenvironment is insufficient, and it is difficult for a single CDT to completely inhibit tumor growth. Here, we designed a CuS nanoparticles (NPs) and camptothecin (CPT) co-loaded thermosensitive injectable hydrogel (SCH) with self-supplied H2O2 for enhanced CDT. SCH is composed of CuS NPs and CPT loaded into agarose hydrogel according to a certain ratio. We injected SCH into the tumor tissue of mice, and under the irradiation of near-infrared region (NIR) laser at 808 nm, CuS NPs converted the NIR laser into heat to realize photothermal therapy (PTT), and at the same time, the agarose hydrogel was changed into a sol state and CPT was released. CPT activates nicotinamide adenine dinucleotide phosphate oxidase, increases the level of H2O2 inside the tumor, and realizes the self-supply of H2O2. At the same time, CuS can accelerate the release of Cu2+ in an acidic environment and light, combined with H2O2 generated by CPT for CDT treatment, and consume glutathione in tumor and generate hydroxyl radical, thus inducing tumor cell apoptosis. The SCH system we constructed achieved an extremely high tumor inhibition rate in vitro and in vivo, presenting a new idea for designing future chemical kinetic systems.

706 Ceramide-1-phosphate accelerates premature skin aging in skin fibroblasts exposed to diesel particulate extract

Extrinsic factors, such as air pollutants and ultraviolet irradiation (UVR), accelerate premature aging, and a correlation between levels of pollutants and skin aging symptoms has been reported. Diesel particulate matter, a major source of air pollution, helps initiate some diseases, including skin diseases. We recently demonstrated that diesel particulate extract (DPE) activates nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX), followed by activation of sphingomyelinase (SMase), leading to ceramide (Cer) production in cultured normal keratinocytes (KC).

Endoplasmic Reticulum Stress and Reactive Oxygen Species in Plants.

The endoplasmic reticulum (ER) is a key compartment responsible for protein processing and folding, and it also participates in many signal transduction and metabolic processes. Reactive oxygen species (ROS) are important signaling messengers involved in the redox equilibrium and stress response. A number of abiotic and biotic stresses can trigger the accumulation of unfolded or misfolded proteins and lead to ER stress. In recent years, a number of studies have reported that redox metabolism and ROS are closely related to ER stress. ER stress can benefit ROS generation and even cause oxidative burden in plants, finally leading to oxidative stress depending on the degree of ER stress. Moreover, ER stress activates nicotinamide adenine dinucleotide phosphate (NADPH) oxidase-mediated ROS signaling, increases antioxidant defense mechanisms, and alters the glutathione (GSH) redox state. Meanwhile, the accumulation of ROS plays a special role in inducing the ER stress response. Given these factors, plants have evolved a series of complex regulatory mechanisms to interact with ROS in response to ER stress. In this review, we summarize the perceptions and responses of plant ER stress and oxidative protein folding in the ER. In addition, we analyze the production and signaling of ROS under ER stress in detail in order to provide a theoretical basis for reducing ER stress to improve the crop survival rate in agricultural applications.

Near-infrared light reduces β-amyloid-stimulated microglial toxicity and enhances survival of neurons: mechanisms of light therapy for Alzheimer’s disease

BackgroundLow-intensity light can decelerate neurodegenerative disease progression and reduce amyloid β (Aβ) levels in the cortex, though the cellular and molecular mechanisms by which photobiomodulation (PBM) protects against neurodegeneration are still in the early stages. Microglia cells play a key role in the pathology of Alzheimer’s disease by causing chronic inflammation. We present new results concerning the PBM of both oxidative stress and microglia metabolism associated with the activation of metabolic processes by 808 nm near-infrared light.MethodsThe studies were carried out using healthy male mice to obtain the microglial cell suspension from the hippocampus. Oligomeric β-amyloid (1-42) was prepared and used to treat microglia cells. Light irradiation of cells was performed using diode lasers emitting at 808 nm (30 mW/cm2 for 5 min, resulting in a dose of 10 J/cm2). Mitochondrial membrane potential, ROS level studies, cell viability, apoptosis, and necrosis assays were performed using epifluorescence microscopy. Phagocytosis, nitric oxide and H2O2 production, arginase, and glucose 6-phosphate dehydrogenase activities were measured using standard assays. Cytokines, glucose, lactate, and ATP were measurements with ELISA. As our data were normally distributed, two-way ANOVA test was used.ResultsThe light induces a metabolic shift from glycolysis to mitochondrial activity in pro-inflammatory microglia affected by oligomeric Aβ. Thereby, the level of anti-inflammatory microglia increases. This process is accompanied by a decrease in pro-inflammatory cytokines and an activation of phagocytosis. Light exposure decreases the Aβ-induced activity of glucose-6-phosphate dehydrogenase, an enzyme that regulates the rate of the pentose phosphate pathway, which activates nicotinamide adenine dinucleotide phosphate oxidases to further produce ROS. During co-cultivation of neurons with microglia, light prevents the death of neurons, which is caused by ROS produced by Aβ-altered microglia.ConclusionsThese original data clarify reasons for how PBM protects against neurodegeneration and support the use of light for therapeutic research in the treatment of Alzheimer’s disease.Graphical

Euphorbia supina Extracts Block NADPH Oxidase-Mediated, Ceramide-Induced Apoptosis Initiated by Diesel Particulate Matter.

Air pollutants contribute to the development of diseases such as asthma, chronic obstructive pulmonary disease (COPD), pulmonary cancer, cardiovascular problems, and some skin diseases. We recently found that a major air pollutant, diesel particulate matter (DPM), induces apoptosis in human keratinocytes by increasing a proapoptotic lipid mediator, ceramide. DPM activates nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX), which stimulates sphingomyelinase, leading to an increased conversion of sphingomyelin to ceramide. Interestingly, we characterized that although NOX is a reactive oxygen species (ROS) generator, the activation of sphingomyelinases by NOX is an ROS-independent mechanism. A Korean weed, prostrate spurge Euphorbia supina Rafin (ESR), has been used for centuries as a folk medicine to treat bronchitis, hepatitis, hemorrhage, and skin inflammation. Flavonoids, terpenes and tannins are enriched in ESR, and although ESR has proven antioxidative activity, its biological activities are largely unknown. Here, we investigate whether and how ESR protects keratinocytes against DPM-mediated apoptosis. We found that ESR-extracts (ESR-Ex) protect keratinocytes from DPM-induced apoptosis by inhibiting NOX activation in keratinocytes in response to DPM. We also demonstrated that ESR-Ex suppresses NOX activation via a blockage of the aryl hydrocarbon receptor (AhR) activation-mediated transcription of neutrophil cytosolic factor 1 (NCF1)/p47phox, a subunit of NOX. Our study reveals previously uncharacterized biological activity of ESR-Ex; i.e., its inhibition of Ahr and NOX activation. Thus, because the inhibition of NOX has already been developed to treat NOX-mediated diseases, including various types of cardiovascular diseases and cancers, initiated by air pollutants and because AhR activation contributes to the development of chronic inflammatory diseases, our study provides further advantages for the medical use of ESR.

H2O2 Self‐Producing Single‐Atom Nanozyme Hydrogels as Light‐Controlled Oxidative Stress Amplifier for Enhanced Synergistic Therapy by Transforming “Cold” Tumors

AbstractSingle‐atom nanozyme (SAzyme) with peroxidase‐like activity can alter cellular redox balance and shows promising potential for tumor therapy. However, the “cold” immune microenvironment and limited amount of hydrogen peroxide (H2O2) in solid tumors severely restrict its efficacy. Herein, a light‐controlled oxidative stress amplifier system is designed by co‐encapsulating Pd‐C SAzymes and camptothecin in agarose hydrogel, which exhibits enhanced synergistic antitumor activity by self‐producing H2O2 and transforming “cold” tumors. In this nanozyme hydrogel system, the Pd‐C SAzyme converts near‐infrared laser into heat, resulting in agarose degradation and consequent camptothecin release. The camptothecin increases H2O2 level in tumors by activating nicotinamide adenine dinucleotide phosphate oxidase, improving the catalytic performance of SAzymes with peroxidase‐like activity. Moreover, the combination of photothermal therapy, chemotherapy, and nanozyme‐based catalytic therapy further facilitates tumor immunogenic death and enhanced antitumor immunity. The results reveal the synergistic antitumor potential of the novel SAzyme/chemotherapeutics‐based hydrogel system.

Fructose-Induced Insulin Resistance: Prospective Biochemical Mechanisms

Increased intake of dietary fructose is markedly associated with multiple negative health outcomes and burdens. Insulin resistance (IR) and type 2 diabetes mellitus (T2DM) are the most common complications that present with conjugated cellular-biochemical abnormalities. This article explains the involvement of increased dietary fructose intake in the occurrence of IR and T2DM and addresses basic metabolic mechanisms. PubMed, Medline, Science Direct, ADI, and WHO databases were searched through June 2021. Current research predicts that over 350 million people may have diabetes by 2030. IR acts as an influencer promoter of T2DM development. IR can occur as a result of high fructose intake. Fructose metabolism results in de novo lipogenesis, while its decreasing effect of peroxisome proliferator-activated receptor (PPAR) activity elevates the levels of inflammatory cytokines, resulting in down-regulation of insulin receptor substrate-1 phosphorylation. Fructose stimulates oxidative stress by activating nicotinamide adenine dinucleotide phosphate oxidase and synthesis of advanced glycation end-products. Fructose also stimulates purine-induced uric acid synthesis and leptin resistance, which contributes to abnormal insulin action. It is crucial to understand the mechanisms of fructose-induced IR via induction of oxidative stress, inflammation, leptin resistance, and uric acid production. This helps prevent and control variable diseases, T2DM being the most.

The Protective Effect of Natural Compounds on Doxorubicin-Induced Cardiotoxicity via Nicotinamide Adenine Dinucleotide Phosphate Oxidase Inhibition

Doxorubicin (DOX) is widely prescribed for the treatment of several human cancers. Unfortunately, cumulative doses of DOX are the main cause of myocardial dysfunction. Although preclinical and pharmaceutical studies were performed to investigate the potential of natural compounds in minimizing DOX toxicity, a comprehensive review of them is not available. This review can help the researchers for an effective search strategy. Oxidative stress and p53 play an important role in DOX-associated cardiotoxicity. DOX activates nicotinamide adenine dinucleotide phosphate NADPH oxidase (NOX) in the heart, resulting in excessive reactive oxygen species that can induce cardiomyocyte apoptosis through phosphorylation of p53, DNA damage and/or mitogen-activated protein kinases-mediated cardiomyocyte apoptosis. Although a few chemical drugs with high efficacy are administered along with DOX to prevent or more likely to reduce cardiovascular toxicity, their use is often limited by additional side effects. Recently, attention has been drawn to natural compounds that prevent DOX cardiotoxicity. This review focuses on some of the natural bioactive compounds with potential therapeutic efficacy against DOX-induced cardiotoxicity (DIC). Some natural compounds, especially flavonols, flavonoids and proanthocyanidins, have the most protective effects against DIC by forming stable radicals and preventing the assembly of the NOX subunits.

Mitochondrial Dysfunction and Alzheimer's Disease: Role of Microglia.

In 1907, Alois Alzheimer observed, as he quoted, development of “numerous fibers” and “adipose saccules” in the brain of his diseased patient Auguste Deter. The neurodegenerative disease became known as Alzheimer’s disease (AD) and is the most common cause of dementia worldwide. AD normally develops with aging and is mostly initiated because of the imbalance between the formation and clearance of amyloid-β (Aβ). Formation of neurofibrillary tangles (NFTs) of hyperphosphorylated tau is another hallmark of AD. Neuroinflammation plays a significant role in the development and pathology of AD. This chapter explores the role of mitochondrial dysfunction in microglia in case of AD. Mitochondrial oxidative stress in microglia has been linked to the development of AD. Elevated generation of reactive oxygen species (ROS) and loss of mitochondrial membrane potential through various mechanisms have been observed in AD. Aβ interacts with microglial receptors, such as triggering receptor expressed in myeloid cells 2 (TREM2), activating downstream pathways causing mitochondrial damage and aggravating inflammation and cytotoxicity. Fibrillar Aβ activates nicotinamide adenine dinucleotide phosphate (NADPH) oxidase in microglia leading to elevated induction of mitochondrial ROS which further causes neurotoxicity. Elevated ROS in microglia causes activation of inflammatory and cell death pathways. Production of ATP, regulation of mitochondrial health, autophagy, and mitophagy in microglia play significant roles in the AD pathology. Understanding microglial physiology and mitochondrial dysfunction will enable better therapeutic interventions.

Modulation of oxidative-nitrosative stress and inflammatory response by rapamycin in target and distant organs in rats exposed to hindlimb ischemia-reperfusion: the role of mammalian target of rapamycin.

Mammalian target of rapamycin (mTOR) has been recognized with potential immunomodulatory properties playing an important role in various physiopathological processes including ischemia-reperfusion (I/R) injury. I/R injury stimulate reactive oxygen and nitrogen species by activating nicotinamide adenine dinucleotide phosphate oxidase and inducible nitric oxide synthase, respectively. Controversial results have been obtained in different I/R models following localized I/R; however, the precise role of the mTOR signaling pathway remains undefined. The objective of the current study was to evaluate the role of the mTOR in oxidative-nitrosative stress and inflammation in hindlimb I/R-induced injury in target and remote organ injuries. In rats subjected to I/R, an increased expression of ribosomal protein S6 (rpS6), inhibitor κB (IκB)-α, nuclear factor-κB (NF-κB) p65, inducible nitric oxide synthase, cyclooxygenase 2, gp91phox, and levels of tumor necrosis factor α, nitrite, nitrotyrosine, malondialdehyde and the activities of myeloperoxidase and catalase in the tissues and (or) sera were detected. Treatment with rapamycin, a selective inhibitor of mTOR, reversed all the I/R-induced changes as manifested by its anti-inflammatory and antioxidant effects in kidney and gastrocnemius muscle of rats. Collectively, these findings suggest that rapamycin protects against I/R-induced oxidative-nitrosative stress and inflammation leading to organ injuries via suppression of mTOR/IκB-α/NF-κB signaling pathway.

2,3′4,4′,5-Pentachlorobiphenyl induces hepatocellular carcinoma cell proliferation through pyruvate kinase M2-dependent glycolysis

Polychlorinated biphenyls (PCBs) are classic persistent organic pollutants (POPs) and are associated with the progression of many cancers, including liver cancer. The present study investigated the effect of 2,3’4,4′,5-pentachlorobiphenyl (PCB118) on hepatocellular carcinoma cell proliferation and its underlying mechanisms. The results indicated that PCB118 exposure promotes the proliferation and glycolysis of hepatocellular carcinoma SMMC-7721 cells. Moreover, PCB118 exposure increased the expression level of pyruvate kinase M2 (PKM2) and its nuclear translocation, whereas treatment with PKM2 shRNA suppressed the induction of cell proliferation and glycolysis by PCB118. PCB118 stimulated reactive oxygen species (ROS) production by activating nicotinamide adenine dinucleotide phosphate (NADPH) oxidase. Treatment with the antioxidants N-acetyl-L-cysteine (NAC) and superoxide dismutase (SOD) prevented PCB118-induced effects on PKM2, cell proliferation and glycolysis. Furthermore, we found that PCB118 activated NADPH oxidase through the aryl hydrocarbon receptor (AhR) in SMMC-7721 cells. Consistently, treatment with AhR shRNA suppressed PCB118-induced effects on PKM2, cell proliferation and glycolysis. Overall, these results indicated that PCB118 promotes HCC cell proliferation via PKM2-dependent upregulation of glycolysis, which is mediated by AhR/NADPH oxidase-induced ROS production.

Calcium Release from Endoplasmic Reticulum Involves Calmodulin-Mediated NADPH Oxidase-Derived Reactive Oxygen Species Production in Endothelial Cells.

Background: Previous studies demonstrated that calcium/calmodulin (Ca2+/CaM) activates nicotinamide adenine dinucleotide phosphate oxidases (NOX). In endothelial cells, the elevation of intracellular Ca2+ level consists of two components: Ca2+ mobilization from the endoplasmic reticulum (ER) and the subsequent store-operated Ca2+ entry. However, little is known about which component of Ca2+ increase is required to activate NOX in endothelial cells. Here, we investigated the mechanism that regulates NOX-derived reactive oxygen species (ROS) production via a Ca2+/CaM-dependent pathway. Methods: We measured ROS production using a fluorescent indicator in endothelial cells and performed phosphorylation assays. Results: Bradykinin (BK) increased NOX-derived cytosolic ROS. When cells were exposed to BK with either a nominal Ca2+-free or 1 mM of extracellular Ca2+ concentration modified Tyrode’s solution, no difference in BK-induced ROS production was observed; however, chelating of cytosolic Ca2+ by BAPTA/AM or the depletion of ER Ca2+ contents by thapsigargin eliminated BK-induced ROS production. BK-induced ROS production was inhibited by a CaM inhibitor; however, a Ca2+/CaM-dependent protein kinase II (CaMKII) inhibitor did not affect BK-induced ROS production. Furthermore, BK stimulation did not increase phosphorylation of NOX2, NOX4, and NOX5. Conclusions: BK-induced NOX-derived ROS production was mediated via a Ca2+/CaM-dependent pathway; however, it was independent from NOX phosphorylation. This was strictly regulated by ER Ca2+ contents.

Hyaluronidase2 (Hyal2) modulates low shear stress-induced glycocalyx impairment via the LKB1/AMPK/NADPH oxidase-dependent pathway.

The endothelium glycocalyx layer (ECL), presents on the apical surface of endothelial cells, creates a barrier between circulating blood and the vessel wall. Low shear stress (LSS) may accelerate the degradation of the glycocalyx via hyaluronidase2 (Hyal2) and then alter the cell polarity. Yet the liver kinase B1 (LKB1) signaling pathway plays an important role in regulating cell polarity. However, the relationship between LKB1 and glycocalyx during LSS is not clear. In the current study, we demonstrate that LSS attenuates LKB1 and AMP-activated protein kinase activation as well as activated nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (p47phox ) and Hyal2 in the human umbilical vein endothelial cell (HUVEC). Pretreatment with 5-Aminoimidazole-4-carboxamide1-β-D-ribofuranoside(AICAR), or diphenyleneiodonium(DPI chloride) and transfection with LKB1 overexpression vector and p47phox small interfering RNA downregulated LSS-induced Hyal2 activation. By coimmunoprecipitation, we discovered the existence of p47phox /Hyal2 complex. LSS induced the dissociation of p47phox /Hyal2 complex, which was inhibited by LKB1 overexpression and AICAR. Furthermore, knockdown of Hyal2 performed a positive feedback on LKB1 activity. In addition, we also show that LSS enhanced LKB1 translocation from the cytosol to the nucleus. Taken together, these data indicate that Hyal2 regulates LSS-induced injury of the glycocalyx via LKB1/AMPK/NADPH oxidase signaling cascades.

Advanced oxidation protein products induce pre-osteoblast apoptosis through a nicotinamide adenine dinucleotide phosphate oxidase-dependent, mitogen-activated protein kinases-mediated intrinsic apoptosis pathway.

SummaryOsteoblast apoptosis contributes to age‐related bone loss. Advanced oxidation protein products (AOPPs) are recognized as the markers of oxidative stress and potent inducers of apoptosis. We have demonstrated that AOPP accumulation was correlated with age‐related bone loss. However, the effect of AOPPs on the osteoblast apoptosis still remains unknown. Exposure of osteoblastic MC3T3‐E1 cells to AOPPs caused the excessive generation of reactive oxygen species (ROS) by activating nicotinamide adenine dinucleotide phosphate (NADPH) oxidases. Increased ROS induced phosphorylation of mitogen‐activated protein kinases (MAPKs), which subsequently triggered intrinsic apoptosis pathway by inducing mitochondrial dysfunction, endoplasmic reticulum stress, and Ca2+ overload and eventually leads to apoptosis. Chronic AOPP loading in aged Sprague‐Dawley rats induced osteoblast apoptosis and activated NADPH oxidase signaling cascade, in combination with accelerated bone loss and deteriorated bone microstructure. Our study suggests that AOPPs induce osteoblast apoptosis by the NADPH oxidase‐dependent, MAPK‐mediated intrinsic apoptosis pathway.