Moderate Levels Of Reactive Oxygen Species Research Articles

LIVING LONGER WITH MODERATE AMOUNTS OF OXYGEN RADICALS IN FRUIT FLIES

Abstract The role of reactive oxygen species (ROS) in aging is complex and paradoxical. While the free radical theory of aging suggests that ROS cause cellular damage and senescence, some studies have shown that moderate ROS levels can enhance cellular adaptation and longevity. In this research, we used Drosophila as an in vivo model to investigate the optimal level of ROS that can promote healthy aging. We exposed flies to different concentrations of paraquat (PQ), a herbicide that generates superoxide anions, and measured their lifespan, stress resistance, and gene expression. We found that intermediate PQ levels (0.1 mM) extended lifespan and improved stress tolerance, while high PQ levels (0.5 mM) shortened lifespan and impaired stress response. We also observed an up-regulation of ROS scavenger genes, such as superoxide dismutase 1 and glutathione peroxidase, in the intermediate PQ group, suggesting a homeostatic mechanism to balance ROS levels. Our findings challenge the conventional view of ROS as harmful agents and provide novel insights into the molecular mechanisms of aging.

Effect of Antioxidant on Sperm Freezing

A major cause of male infertility, which includes 50 causes of infertility, is oxidative stress. Aerobic respiration in the sperm cell is associated with the production of reactive oxygen species (ROS). Some reactions, such as sperm capacitation and acrosome reactions require moderate levels of ROS. However, when the amount of ROS production surpasses the natural antioxidant defense of the cell, it causes a harmful effect called oxidative stress. This results in the oxidation of lipids, proteins, carbohydrates, and nucleotides. The production of ROS is increased by various factors, including the presence of leukocytes, sperms with abnormal and immature morphology, centrifugation of samples, and changes in oxygen concentration, pH, and temperature. Sperm freezing is a method widely used in assisted reproductive technology (ART) as well as infertility treatment. The process of freezing also contributes to increased oxidative stress because it alters the fluidity of the mitochondrial membrane, leading to increased production and release of ROS. Sperm have defense mechanisms against oxidative stress caused by increased ROS production, which includes the presence of enzymatic and non-enzymatic antioxidants in semen. The utilization of supplementary antioxidants in the freezing environment is a strategy to combat oxidative stress. This review aims to summarize the current evidence on the effect of antioxidants on sperm freezing.

Reactive Oxygen Species are Essential for Placental Angiogenesis During Early Gestation.

Background Preeclampsia (PE) is associated with insufficient placental perfusion attributed to maldevelopment of the placental vasculature. Reactive oxygen species (ROS) are implicated in angiogenesis, but their regulatory effects and mechanisms in placental vascular development remain unclear. Methods Placental oxidative stress was determined throughout gestation by measuring 4-hydroxynonenal (4HNE) and malondialdehyde (MDA). The antioxidant MitoQ was administered to pregnant mice from GDs 7.5 to 11.5; placental morphology and angiogenesis pathways were examined on GDs 11.5 and 18.5. Moreover, we established a mouse mFlt-1-induced PE model and assessed blood pressure, urine protein levels, and placental vascular development on GDs 11.5 and 18.5. Human umbilical vein endothelial cells (HUVECs) were treated with various H2O2 concentrations to evaluate cell viability, intracellular ROS levels, and tube formation capability. MitoQ, an AKT inhibitor and an ERK1/2 inhibitor were applied to validate the ROS-mediated mechanism regulating placental angiogenesis. Results First-trimester placentas presented significantly higher MDA and 4HNE levels. MitoQ significantly reduced the blood vessel density and angiogenesis pathway activity in the placenta on GDs 11.5 and 18.5. Serum sFlt-1 levels were elevated, and we observed poor placental angiogenesis and PE-like symptoms in cases with mFlt-1 overexpression. Moderate H2O2 treatment promoted HUVEC proliferation and angiogenesis, whereas these improvements were abolished by MitoQ, AKT inhibitor, or ERK1/2 inhibitor treatment. Conclusions Moderate ROS levels are essential for placental angiogenesis; diminishing ROS with potent antioxidants during placentation decreases placental angiogenesis and increases PE risk. Therefore, antioxidant therapy should be considered carefully for normal pregnant women during early gestation.

Oxidative stress‐mediated up‐regulation of ABC transporters in lung cancer cells

This paper aimed to evaluate the role of oxidative stress in the regulation of ABC transporters in human lung cancer (A549) cells facing substrate (doxorubicin, DOX) and nonsubstrate (ethanol, ETH and hydrogen peroxide, HP) chemicals. After 24-h treatment, all the chemicals caused significant cytotoxicity as reflected by the reduction in cell viability and the increase in reactive oxygen species (ROS) levels. Depending on the rescuing effects of ROS scavenger including glutathione (GSH) and Vitamin C, the toxicity dependence on oxidative stress were found to be HP > ETH > DOX. The addition of transporter inhibitors significantly enhanced the ROS levels and death-inducing effects of chemicals, indicating the universal detoxification function of ABC transporters. At moderate ROS levels (about 3-4-folds of control levels, caused by 10 μM DOX, 400 mM ETH, and 400 μM HP), all the three chemicals induced the gene expressions and activities of ABC transporters, but these values decreased at too high ROS levels (8.36-folds of control levels) caused by HP at LC50 (800 μM). Such induction could be attenuated by GSH and KCZ, and was completely abolished by 50 μM KCZ, indicating an important role of oxidative stress and pregnane X receptor (PXR) in the induction of ABC transporters. Finally, this paper revealed the critical role of oxidative stress in the modulation of ABC transporters by either substrate or nonsubstrate chemicals during 24-h treatment. Such information should be beneficial for overcoming ABC transporter-mediated multidrug resistance.

The interplay of ROS and the PI3K/Akt pathway in autophagy regulation.

Autophagy causes the breakdown of damaged proteins and organelles to their constituent components. The phosphatidylinositol 3-kinase (PI3K) pathway played an important role in regulating the autophagic response of cells in response to changing reactive oxygen species (ROS) levels. The PI3K α catalytic subunit inhibits autophagy, while its β catalytic subunit promotes autophagy in response to changes in ROS levels. The downstream Akt protein acts against autophagy initiation in response to increases in ROS levels under nutrient-rich conditions. Akt acts by activating a mechanistic target of the rapamycin complex 1 (mTORC1) and by arresting autophagic gene expression. The AMP-activated protein kinase (AMPK) protein counteracts the Akt actions. mTORC1 and mTORC2 inhibit autophagy under moderate ROS levels, but under high ROS levels, mTORC2 can promote cellular senescence via autophagy. Phosphatase and tensin homolog (PTEN) protein are the negative regulators of the PI3K pathway, and it has proautophagic activities. Studies conducted on cells treated with flavonoids and ionizing radiation showed that the moderate increase in ROS levels in the flavonoid-treated groups corresponded with higher PTEN levels and lowered Akt levels leading to a higher occurrence of autophagy. In contrast, higher ROS levels evoked by ionizing radiation caused a lowering of the incidence of autophagy.

Targeting Reactive Oxygen Species Capacity of Tumor Cells with Repurposed Drug as an Anticancer Therapy.

Accumulating evidence shows that elevated levels of reactive oxygen species (ROS) are associated with cancer initiation, growth, and response to therapies. As concentrations increase, ROS influence cancer development in a paradoxical way, either triggering tumorigenesis and supporting the proliferation of cancer cells at moderate levels of ROS or causing cancer cell death at high levels of ROS. Thus, ROS can be considered an attractive target for therapy of cancer and two apparently contradictory but virtually complementary therapeutic strategies for the regulation of ROS to treat cancer. Despite tremendous resources being invested in prevention and treatment for cancer, cancer remains a leading cause of human deaths and brings a heavy burden to humans worldwide. Chemotherapy remains the key treatment for cancer therapy, but it produces harmful side effects. Meanwhile, the process of de novo development of new anticancer drugs generally needs increasing cost, long development cycle, and high risk of failure. The use of ROS-based repurposed drugs may be one of the promising ways to overcome current cancer treatment challenges. In this review, we briefly introduce the source and regulation of ROS and then focus on the status of repurposed drugs based on ROS regulation for cancer therapy and propose the challenges and direction of ROS-mediated cancer treatment.

Utility of Reactive Species Generation in Plasma Medicine for Neuronal Development.

Reactive oxygen species (ROS) are critical signaling molecules for neuronal physiology that stimulate growth and development and play vital roles in several pathways when in a balanced state, but they cause neurodegeneration when unbalanced. As ROS levels above a certain threshold cause the activation of the autophagy system, moderate levels of ROS can be used as treatment strategies. Currently, such treatments are used together with low-level laser or photodynamic therapies, photo-bio modulation, or infrared treatments, in different chronic diseases but not in the treatment of neurodegeneration. Recently, non-thermal plasma has been successfully used in biomedical applications and treatments, and beneficial effects such as differentiation, cell growth, and proliferation, stimulation of ROS based pathways have been observed. Besides the activation of a wide range of biological signaling pathways by generating ROS, plasma application can be an effective treatment in neuronal regeneration, as well as in neuronal diseases. In this review, we summarize the generation and role of ROS in neurons and provide critical insights into their potential benefits on neurons. We also discuss the underlying mechanisms of ROS on neuronal development. Regarding clinical applications, we focus on ROS-based neuronal growth and regeneration strategies and in the usage of non-thermal plasma in neuronal and CNS injury treatments.

Warburg effect and its role in tumourigenesis.

Glucose is a crucial molecule in energy production and produces different end products in non-tumourigenic- and tumourigenic tissue metabolism. Tumourigenic cells oxidise glucose by fermentation and generate lactate and adenosine triphosphate even in the presence of oxygen (Warburg effect). The Na+/H+-antiporter is upregulated in tumourigenic cells resulting in release of lactate- and H+ ions into the extracellular space. Accumulation of lactate- and proton ions in the extracellular space results in an acidic environment that promotes invasion and metastasis. Otto Warburg reported that tumourigenic cells have defective mitochondria that produce less energy. However, decades later it became evident that these mitochondria have adapted with alterations in mitochondrial content, structure, function and activity. Mitochondrial biogenesis and mitophagy regulate the formation of new mitochondria and degradation of defective mitochondria in order to combat accumulation of mutagenic mitochondrial deoxyribonucleic acid. Tumourigenic cells also produce increase reactive oxygen species (ROS) resulting from upregulated glycolysis leading to pathogenesis including cancer. Moderate ROS levels exert proliferative- and prosurvival signaling, while high ROS quantities induce cell death. Understanding the crosstalk between aberrant metabolism, redox regulation, mitochondrial adaptions and pH regulation provides scientific- and medical communities with new opportunities to explore cancer therapies.

Nanomedicines for Reactive Oxygen Species Mediated Approach: An Emerging Paradigm for Cancer Treatment.

Growth in the knowledge of cancer biology has led to the emergence and evolution of cancer nanomedicines by providing the rationale for leveraging nanotechnology to develop better treatment options. The discovery of nanometer-sized intercellular openings in the defective angiogenic tumor vasculature contributed to the development of an idea for the well-known cancer passive targeting regime, enhanced permeability and retention (EPR) effect, of the nanomedicines. Recently, reactive oxygen species (ROS) have been highlighted as one of the key players that underlie the acquisition of the various hallmarks of cancer. As ROS are associated with all stages of cancer, their applications in cancer treatment based on the following concentration-dependent implications have attracted much attention: (1) low to moderate levels of ROS as key signaling molecules, (2) elevated levels of ROS in cancer cells as one of the unique characteristics of cancer, and (3) excessive levels of ROS as cytotoxic agents. Considering ROS from a different point of view, various cancer nanomedicines have been designed to achieve spatiotemporal control of therapeutic action, the main research focus in this area. This Account includes our efforts and preclinical achievements in development of nanomedicines for a range of ROS-mediated cancer therapies. It begins with general background regarding cancer nanomedicines, the significance of ROS in cancer, and a brief overview of ROS-mediated approaches for cancer therapy. Then, this Account highlights the two key roles of ROS that define therapeutic purposes of cancer nanomedicines: (1) ROS as drug delivery enhancers and (2) ROS as cell death inducers. The former inspired us to develop nitric oxide-generating nanoparticles for improved EPR effect, endogenous ROS-responsive polymeric micelles for enhanced intracellular drug delivery, and exogenous ROS-activated micelles for subcellular localization via photochemical internalization. While refining conventional chemotherapy, recent researches also have focused on the latter, the cytotoxic ROS, to advance alternative treatment modalities such as oxidation therapy, photodynamic therapy (PDT), and sonodynamic therapy (SDT). In particular, we have been motivated to develop polymeric nanoreactors containing enzymes to produce H2O2 for oxidation therapy, photosensitizer-loaded gold-nanoclustered polymeric nanoassemblies for photothermally activated PDT overcoming the oxygen dependency of PDT, and hydrophilized TiO2 nanoparticles and Au-TiO2 nanocomposites as novel sonosensitizers for improved SDT efficiency. The integration of nanomedicine and ROS-mediated therapy has emerged as the new paradigm in the treatment of cancer, based on promising proof-of-concept demonstrations in preclinical studies. Further efforts to ensure clinical translation along with more sophisticated cancer nanomedicines to address relevant challenges are expected to be made in the coming years.

Antioxidant response of Arabidopsis thaliana seedlings to oxidative stress induced by carbon ion beams irradiation

Due to the fact that carbon ion beams irradiation as an important type of ionizing radiation can potentially cause oxidative stress in plants, it is significant to evaluate the antioxidant response of plants to carbon ion beams radiation. Therefore, the objective of this study is to investigate the effects of carbon ion beams irradiation on oxidative stress induced by reactive oxygen species (ROS) and antioxidant response in Arabidopsis thaliana seedlings by irradiating the dry seeds at various doses of carbon ion beams (0, 50, 100, 150 and 200 Gy) and measuring the plant growth parameters, ROS and malondialdehyde (MDA) levels, activities of antioxidant systems and antioxidant-related gene expression. The results showed that 50-Gy carbon ion beam irradiation exhibited stimulatory effects on germination index, root length and fresh weight in Arabidopsis seedlings, while high-dose irradiation (100–200 Gy) inhibited plant growth. Moreover, the production rate of superoxide anion radical, hydroxyl radical generation activity, hydrogen peroxide and MDA contents in Arabidopsis seedlings were obviously increased with the irradiation dose. Additionally, the antioxidant enzyme activities (superoxide dismutase, catalase and peroxidase) and non-enzymatic antioxidant contents (ascorbate and glutathione) in 50-Gy irradiated seedlings were apparently higher than control. Notably, transcriptional analysis displayed that 50-Gy carbon ion beams irradiation could enhance the expression of antioxidant-related genes in Arabidopsis seedlings. These results suggest that the improved activities of antioxidant systems induced by moderate ROS levels play important roles in growth promotion of Arabidopsis seedlings caused by low-dose carbon ion beams irradiation.

Function and Regulation of Protein Kinase D in Oxidative Stress: A Tale of Isoforms.

Oxidative stress is a condition that arises when cells are faced with levels of reactive oxygen species (ROS) that destabilize the homeostatic redox balance. High levels of ROS can cause damage to macromolecules including DNA, lipids, and proteins, eventually resulting in cell death. Moderate levels of ROS however serve as signaling molecules that can drive and potentiate several cellular phenotypes. Increased levels of ROS are associated with a number of diseases including neurological disorders and cancer. In cancer, increased ROS levels can contribute to cancer cell survival and proliferation via the activation of several signaling pathways. One of the downstream effectors of increased ROS is the protein kinase D (PKD) family of kinases. In this review, we will discuss the regulation and function of this family of ROS-activated kinases and describe their unique isoform-specific features, in terms of both kinase regulation and signaling output.

Redox Homeostasis Plays Important Roles in the Maintenance of the Drosophila Testis Germline Stem Cells.

SummaryOxidative stress influences stem cell behavior by promoting the differentiation, proliferation, or apoptosis of stem cells. Thus, characterizing the effects of reactive oxygen species (ROS) on stem cell behavior provides insights into the significance of redox homeostasis in stem cell-associated diseases and efficient stem cell expansion for cellular therapies. We utilized the Drosophila testis as an in vivo model to examine the effects of ROS on germline stem cell (GSC) maintenance. High levels of ROS induced by alteration in Keap1/Nrf2 activity decreased GSC number by promoting precocious GSC differentiation. Notably, high ROS enhanced the transcription of the EGFR ligand spitz and the expression of phospho-Erk1/2, suggesting that high ROS-mediated GSC differentiation is through EGFR signaling. By contrast, testes with low ROS caused by Keap1 inhibition or antioxidant treatment showed an overgrowth of GSC-like cells. These findings suggest that redox homeostasis regulated by Keap1/Nrf2 signaling plays important roles in GSC maintenance.

The Importance of NADPH Oxidases and Redox Signaling in Angiogenesis.

Eukaryotic cells have to cope with the constant generation of reactive oxygen species (ROS). Although the excessive production of ROS might be deleterious for cell biology, there is a plethora of evidence showing that moderate levels of ROS are important for the control of cell signaling and gene expression. The family of the nicotinamide adenine dinucleotide phosphate oxidases (NADPH oxidases or Nox) has evolved to produce ROS in response to different signals; therefore, they fulfil a central role in the control of redox signaling. The role of NADPH oxidases in vascular physiology has been a field of intense study over the last two decades. In this review we will briefly analyze how ROS can regulate signaling and gene expression. We will address the implication of NADPH oxidases and redox signaling in angiogenesis, and finally, the therapeutic possibilities derived from this knowledge will be discussed.

The Role of Reactive Oxygen Species in Epilepsy

Excess of reactive oxygen species (ROS) is increasingly recognized as a key factor in seizure-induced neuronal damage. Thus, targeting ROS is a priority to prevent seizures and epilepsy comorbidities such as cognitive decline, which are largely driven by neuronal damage. One drawback of this approach is that moderate levels of ROS are important in physiological cellular function and adaptation. This may also explain why antioxidant strategies targeting the brain have largely been unsuccessful. To overcome this difficulty, more fine-tuned ROS suppression in epilepsy is warranted. To achieve this goal, it is necessary to identify the key producers of ROS in seizures and epilepsy. Recent studies show that NADPH oxidase and xanthine oxidase are enzymes generating ROS in settings where energy metabolism is high, such as seen during seizure activity. An alternative approach is by targeting inducible networks of cellular antioxidant defences such as the Keap1–Nrf2 system. Nrf2 has emerged as a powerful regulator of endogenous antioxidant defences, and one drug (dimethyl fumarate) activating the Nrf2 pathway is already in use in inflammatory central nervous system (CNS) diseases. Both targeting key producers of ROS during seizure activity and Nrf2-enhancing strategies represent exciting new avenues of drug discovery in epilepsy.

Abstract B31: NOX2 generated reactive oxygen species promotes glioblastoma growth through inhibition of PTEN

Abstract The tumor suppressor protein Phosphatase and Tensin Homolog (PTEN) is a negative regulator of the Akt pathway which promotes proliferation, migration, and self-renewal. PTEN is mutated or deleted in ~40% of glioblastoma (GBM) resulting in increased Akt activity; however almost all GBM's have increased Akt activity, regardless of PTEN status. This raises the possibility that GBM may inactivate PTEN through means other than mutation or deletion. Treatment with exogenous reactive oxygen species (ROS), like hydrogen peroxide, inactivates PTEN through oxidation in the phosphatase domain. The low oxygen environment of tumors (1%O2) increases ROS levels as compared to normal body tissue (4% O2), and tissue culture conditions (20% O2). Recent data indicate that low to moderate levels of ROS are required for cell maintenance and stem cell renewal while moderate increases in ROS may promote migration, invasion, and proliferation. Indeed, our lab recently found that normal neural stem cells have higher ROS levels, which is required for stem cell renewal. Moreover, data from our lab and others, have found that growing cells under physiological oxygen levels increases ROS and promotes renewal and proliferation. This suggests that the low-oxygen environment of tumors may promote proliferation and tumorigenesis through post-translational modification of proteins, like PTEN. NADPH oxidase (NOX) is a membrane bound complex of proteins that produces a significant amount of endogenous intracellular ROS. While this protein was originally identified in macrophages as being important for immune surveillance, new data suggests that the family of NOX proteins (1-4) has many other biological functions in a variety of different cell types. New data has found that the constitutively active NOX4 acts as an oncogene in several different cancer types by promoting migration, invasion, and proliferation. Within the brain, NOX2 promotes ROS-induced stem cell renewal in normal neural stem cells. Additionally, increased NOX2 expression in GBM patients correlates with shorter survival, while patients with low NOX2 expression generally live longer. Together these data suggest that, through NOX2, the low-oxygen environment of GBM increases ROS levels that oxidize and inhibit PTEN. This novel post-translational modification may increase proliferation and promote tumorigenicity through activation of Akt in cells with intact PTEN. We examined endogenous ROS levels in patient-derived GBM samples and found that increased ROS correlated with faster proliferating cells. Furthermore, low levels of O2 (1%) increased proliferation and Akt activation in a PTEN dependent manner. Both PTEN null and knockdown cells showed no increase in either proliferation or Akt activation over cells grown at room air (20%). Moreover, pharmacological inhibition of Akt abrogated ROS-induced proliferation, suggesting this mechanism functions through the PTEN/Akt pathway. This response was due directly to ROS production, as treatment with antioxidants decreased both ROS production and proliferation at low oxygen and room air. Additionally, we found that growth at 1% O2 led to the oxidation and inhibition of PTEN as seen by Akt phosphorylation. Treatment with apocynin, a NOX2 inhibitor decreased ROS production, proliferation, and Akt phosphorylation, suggesting that ROS-induced proliferation may be mediated through the NOX2 protein. This data not only show PTEN oxidation and inhibition under physiological conditions for the first time, it suggests that the low-oxygen tumor microenvironment may alter PTEN positive GBM to up-regulate the Akt pathway and promote tumorigenesis. Citation Format: Kirsten Ludwig, Harley Kornblum. NOX2 generated reactive oxygen species promotes glioblastoma growth through inhibition of PTEN. [abstract]. In: Proceedings of the AACR Special Conference: Advances in Brain Cancer Research; May 27-30, 2015; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2015;75(23 Suppl):Abstract nr B31.

ZNF32 protects against oxidative stress-induced apoptosis by modulating C1QBP transcription.

Reactive oxygen species (ROS)-driven oxidative stress has been recognized as a critical inducer of cancer cell death in response to therapeutic agents. Our previous studies have demonstrated that zinc finger protein (ZNF)32 is key to cell survival upon oxidant stimulation. However, the mechanisms by which ZNF32 mediates cell death remain unclear. Here, we show that at moderate levels of ROS, Sp1 directly binds to two GC boxes within the ZNF32 promoter to activate ZNF32 transcription. Alternatively, at cytotoxic ROS concentrations, ZNF32 expression is repressed due to decreased binding activity of Sp1. ZNF32 overexpression maintains mitochondrial membrane potential and enhances the antioxidant capacity of cells to detoxify ROS, and these effects promote cell survival upon pro-oxidant agent treatment. Alternatively, ZNF32-deficient cells are more sensitive and vulnerable to oxidative stress-induced cell injury. Mechanistically, we demonstrate that complement 1q-binding protein (C1QBP) is a direct target gene of ZNF32 that inactivates the p38 MAPK pathway, thereby exerting the protective effects of ZNF32 on oxidative stress-induced apoptosis. Taken together, our findings indicate a novel mechanism by which the Sp1-ZNF32-C1QBP axis protects against oxidative stress and implicate a promising strategy that ZNF32 inhibition combined with pro-oxidant anticancer agents for hepatocellular carcinoma treatment.

Cadmium-induced autophagy is mediated by oxidative signaling in PC-12 cells and is associated with cytoprotection.

Oxidative stress induced by cadmium (Cd) is a common phenomenon that has been observed in numerous studies. However, the underlying mechanism remains unknown. Recently, exposure of PC-12 cells to Cd has been shown to activate autophagy, which acts as a temporary survival pathway under stressful conditions by delaying the occurrence of apoptosis. The present study investigated the impact of oxidative stress on Cd‑induced autophagy in PC-12 cells. The results demonstrated that Cd‑induced autophagy (following treatment with Cd for 4 h), increased the levels of intracellular reactive oxygen species (ROS), decreased the mitochondrial membrane potential and resulted in apoptosis. A treatment with chloroquine (CQ; an autophagic inhibitor) sensitized the PC‑12 cells to Cd, due to the increased production of ROS, which was associated with the incapacity to reduce mitochondrial and cell death. N-acetyl-L-cysteine, an antioxidant agent, decreased Cd-induced autophagy and reduced intracellular ROS levels, but enhanced CQ‑induced apoptotic cell death. These findings indicate that moderate levels of ROS are essential in the regulation of Cd-induced autophagy, which subsequently enhances cell survival. Thus, the results of the present study provide an insight for future investigation of Cd-induced neurotoxicity.

ROS-Generating Oxidase Nox3 Regulates the Self-Renewal of Mouse Spermatogonial Stem Cells1

Spermatogonial stem cells (SSCs) represent a unique population of germ cells with self-renewal potential. Although reactive oxygen species (ROS) are considered toxic to germ cells, we recently showed that moderate levels of ROS are required for SSC self-renewal and that Nox1 is involved in ROS generation. In this study, we showed that self-renewal factor treatment induces Nox3 to trigger SSC self-renewal. Nox3 was transiently expressed in cultured spermatogonia by FGF2 and GDNF stimulation, whereas Nox1 was expressed predominantly during the stable phase of proliferation. Nox3 inhibition by short hairpin RNA reduced cytokine-induced ROS generation and limited the proliferation of cultured spermatogonia. Although Nox3 overexpression revealed no apparent effect, depletion of Nox3 decreased the number of SSCs in both cultured spermatogonia and freshly isolated testis cells. Our results suggest that self-renewal of SSCs is regulated by sequential activation of different Nox genes, and underscore the complexity of ROS regulation in the self-renewal division of SSCs.

The iron chelator deferasirox affects redox signalling in haematopoietic stem/progenitor cells.

The iron chelator deferasirox (DFX) prevents complications related to transfusional iron overload in several haematological disorders characterized by marrow failure. It is also able to induce haematological responses in a percentage of treated patients, particularly in those affected by myelodysplastic syndromes. The underlying mechanisms responsible for this feature, however, are still poorly understood. In this study, we investigated the effect of DFX-treatment in human haematopoietic/progenitor stem cells, focussing on its impact on the redox balance, which proved to control the interplay between stemness maintenance, self-renewal and differentiation priming. Here we show, for the first time, that DFX treatment induces a significant diphenyleneiodonium-sensitive reactive oxygen species (ROS) production that leads to the activation of POU5F1 (OCT4), SOX2 and SOX17 gene expression, relevant in reprogramming processes, and the reduction of the haematopoietic regulatory proteins CTNNB1 (β-Catenin) and BMI1. These DFX-mediated events were accompanied by decreased CD34 expression, increased mitochondrial mass and up-regulation of the erythropoietic marker CD71 (TFRC) and were compound-specific, dissimilar to deferoxamine. Our findings would suggest a novel mechanism by which DFX, probably independently on its iron-chelating property but through ROS signalling activation, may influence key factors involved in self-renewal/differentiation of haematopoietic stem cells.

Dynamic Changes in Intracellular ROS Levels Regulate Airway Basal Stem Cell Homeostasis through Nrf2-Dependent Notch Signaling

Airways are exposed to myriad environmental and damaging agents such as reactive oxygen species (ROS), which also have physiological roles as signaling molecules that regulate stem cell function. However, the functional significance of both steady and dynamically changing ROS levels in different stem cell populations, as well as downstream mechanisms that integrate ROS sensing into decisions regarding stem cell homeostasis, are unclear. Here, we show in mouse and human airway basal stem cells (ABSCs) that intracellular flux from low to moderate ROS levels is required for stem cell self-renewal and proliferation. Changing ROS levels activate Nrf2, which activates the Notch pathway to stimulate ABSC self-renewal and an antioxidant program that scavenges intracellular ROS, returning overall ROS levels to a low state to maintain homeostatic balance. This redox-mediated regulation of lung stem cell function has significant implications for stem cell biology, repair of lung injuries, and diseases such as cancer.