Redox Signaling Research Articles

Metabolic profiling and combined therapeutic strategies unveil the cytotoxic potential of selenium-chrysin (SeChry) in NSCLC cells.

Lung cancer ranks as the predominant cause of cancer-related mortalities on a global scale. Despite progress in therapeutic interventions, encompassing surgical procedures, radiation, chemotherapy, targeted therapies and immunotherapy, the overall prognosis remains unfavorable. Imbalances in redox equilibrium and disrupted redox signaling, common traits in tumors, play crucial roles in malignant progression and treatment resistance. Cancer cells, often characterized by persistent high levels of reactive oxygen species (ROS) resulting from genetic, metabolic, and microenvironmental alterations, counterbalance this by enhancing their antioxidant capacity. Cysteine availability emerges as a critical factor in chemoresistance, shaping the survival dynamics of non-small cell lung cancer (NSCLC) cells. Selenium-chrysin (SeChry) was disclosed as a modulator of cysteine intracellular availability. This study comprehensively characterizes the metabolism of SeChry and investigates its cytotoxic effects in NSCLC. SeChry treatment induces notable metabolic shifts, particularly in selenocompound metabolism, impacting crucial pathways such as glycolysis, gluconeogenesis, the tricarboxylic acid (TCA) cycle, and amino acid metabolism. Additionally, SeChry affects the levels of key metabolites such as acetate, lactate, glucose, and amino acids, contributing to disruptions in redox homeostasis and cellular biosynthesis. The combination of SeChry with other treatments, such as glycolysis inhibition and chemotherapy, results in greater efficacy. Furthermore, by exploiting NSCLC's capacity to consume lactate, the use of lactic acid-conjugated dendrimer nanoparticles for SeChry delivery is investigated, showing specificity to cancer cells expressing monocarboxylate transporters.

Development of a Novel Amplifiable System to Quantify Hydrogen Peroxide in Living Cells.

Although many redox signaling molecules are present at low concentrations, typically ranging from micromolar to submicromolar levels, they often play essential roles in a wide range of biological pathways and disease mechanisms. However, accurately measuring low-abundant analytes has been a significant challenge due to the lack of sensitivity and quantitative capability of existing measurement methods. In this study, we introduced a novel chemically induced amplifiable system for quantifying low-abundance redox signaling molecules in living cells. We utilized H2O2 as a proof-of-concept analyte and developed a probe that quantifies cellular peroxide levels by combining the NanoBiT system with androgen receptor dimerization as a reporting mechanism. Our system demonstrated a highly sensitive response to cellular peroxide changes induced both endogenously and exogenously. Furthermore, the system can be adapted for the quantification of other signaling molecules if provided with suitable probing chemistry.

Furanodienone induces apoptosis via regulating the PRDX1/MAPKs/p53/caspases signaling axis through NOX4-derived mitochondrial ROS in colorectal cancer cells

Furanodienone, a biologically active constituent of sesquiterpenes isolated from Rhizome Curcumae, has been reported to induce apoptosis in human colorectal cancer (CRC) cells by promoting the generation of reactive oxygen species (ROS). However, the source of ROS and how it manipulates apoptosis in CRC cells remains to be elucidated. Herein, we assessed the potential role of the well-known sources of intracellular ROS—mitochondrial electron transport chain and the nicotinamide adenine dinucleotide phosphate oxidases (NOXs), on furanodienone-induced cell death. The results indicated that furanodienone substantially increased the levels of mitochondrial ROS, which were subsequently eliminated by the general NOX inhibitor. Specifically, the nuclear factor kappa-B (NF-κB) translocation triggered a significant rise in the expression of NOX4, an isoform of the NOXs family, upon furanodienone treatment. Nevertheless, the specific NOX4 inhibitor GLX351322 attenuated cell apoptosis and mitochondrial ROS production. As a result, ROS burst induced by furanodienone suppressed the expression of peroxiredoxin1 (PRDX1), a redox signaling protein overexpressed in CRC cells, through a nuclear factor-erythroid-2-related factor 2 (Nrf2)-dependent pathway, thus amplifying the mitogen-activated protein kinases (MAPKs)/p53-mediated apoptotic signaling by increasing the p-p38, p-JNK levels, as well as the cleaved caspases −3, −8 and −9. In vivo experiments further confirmed the anti-proliferative impact of PRDX1 following furanodienone treatment. In summary, the study demonstrated that furanodienone-induced apoptosis in CRC cells is initiated by mitochondrial ROS derived from NOX4, which targeted the PRDX1 and activated the downstream MAPKs/p53-mediated caspase-dependent signaling pathway. Our findings may provide novel insights into the development of adjuvant drugs for CRC treatment and therapeutic applications.

Supersulfide formation in the sinus mucosa of chronic rhinosinusitis.

Disruption of the oxidative stress defense system is involved in developing various diseases. Sulfur compounds such as glutathione (GSH) and cysteine (CysSH) are representative antioxidants in the body. Recently, supersulfides, including reactive persulfide and polysulfide species, have gained attention as potent antioxidants regulating oxidative stress and redox signaling. However, their involvement in the pathogenesis of chronic rhinosinusitis (CRS) remains unclear. To clarify the changes in sulfur compounds within the sinus mucosa of each CRS subtype, we measured sulfur compound levels in the sinus mucosa of control individuals (n = 9), patients with eosinophilic CRS (ECRS) (n = 13), and those with non-ECRS (nECRS) (n = 11) who underwent sinus surgery using mass spectrometry. GSH and CysSH levels were significantly reduced, and the glutathione disulfide (GSSG)/GSH ratio, an oxidative stress indicator, was increased in patients with ECRS. Despite the absence of notable variations in supersulfides, patients with ECRS and nECRS exhibited a significant reduction in glutathione trisulfide (GSSSG), which serves as the precursor for supersulfides. This study is the first quantitative assessment of supersulfides in normal and inflamed sinus mucosa, suggesting that sulfur compounds contribute to the pathogenesis of CRS. N/A.

Design and synthesis of visible light-activatable photocaged peroxides for optical control of ROS-mediated cellular signaling

We designed and synthesized two novel photocaged peroxide compounds, N5TBHP and N6TBHP, featuring nitrogen-containing fused ring coumarin skeletons. Notably, a tetrahydroquinoline fused coumarin derivative, N6TBHP demonstrated significantly higher photocleavage efficiency under visible light at 455 nm compared to N5TBHP, which contains an indoline fused coumarin. This process effectively releases the oxidative stress inducer tert-butylhydroperoxide (TBHP). Additionally, N6TBHP exhibits high resistance to glutathione (GSH), and its UV spectral analysis suggests enhanced intracellular stability due to reduced reactivity with GSH through self-assembly. Furthermore, N6TBHP can release an optimal amount of TBHP into cells under visible light irradiation with minimal cell damage. These properties position N6TBHP as a promising tool for advancing research in intracellular redox signaling.

Elucidating PAR1 as a therapeutic target for delayed traumatic brain injury: Unveiling the PPAR-γ/Nrf2/HO-1/GPX4 axis to suppress ferroptosis and alleviate NLRP3 inflammasome activation in rats

Repetitive traumatic brain injury (RTBI) is acknowledged as a silent overlooked public health crisis, with an incomplete understanding of its pathomechanistic signaling pathways. Mounting evidence suggests the involvement of thrombin and its receptor, the protease-activated receptor (PAR)1, in the development of secondary injury in TBI; however, the consequences of PAR1 modulation and its impact on ferroptosis-redox signaling, and NLRP3 inflammasome activation in RTBI, remain unclear. Further, the utilitarian function of PAR1 as a therapeutic target in RTBI has not been elucidated. To study this crosstalk, RTBI was induced in Wistar rats by daily weight drops on the right frontal region for five days. Three groups were included: normal control, untreated RTBI, and RTBI+SCH79797 (a PAR1 inhibitor administered post-trauma at 25 μg/kg/day). The concomitant treatment of PAR1 antagonism improved altered behavior function, cortical histoarchitecture, and neuronal cell survival. Moreover, the receptor blockade downregulated mRNA expression of PAR1 but upregulatedthat of the neuroprotective receptor PPAR-γ. The anti-inflammatory impact of SCH79797 was signified by the low immune expression/levels of NF-κB p65,TNF-α, IL-1β, and IL-18. Consequently, the PAR1 blocker hindered the formation of inflammasome components NLRP3, ASC, and activated caspase-1. Ultimately, SCH79797 treatment abated ferroptosis-dependent iron redox signaling through the activation of the antioxidant Nrf2/HO-1 axis and its subsequent antioxidant machinery (GPX4, SOD) to limit lipid peroxidation, iron accumulation, and transferrin serum increment. Collectively, SCH79797 offered putative preventive mechanisms against secondary RTBI consequences in rats by impeding ferroptosis and NLRP3 inflammasome through activating the PPAR-γ/Nrf2 antioxidant cue.

Electrochemical classification of benzodiazepines: A comprehensive approach combining insights from voltammetry and liquid chromatography − mass spectrometry

The growing non-medical use of benzodiazepines (BZs) has led to the emergence of counterfeit BZ pills and new psycho-active substances (NPS) in the BZ class on the illicit market. Comprehensive analytical methods for BZ identification are required to allow law enforcement, first aid responders and drug-checking services to analyze a variety of sample types and contents to make timely decisions on the spot. In this work, the electrochemical behavior of diazepam (DZ), clonazepam (CZ) and alprazolam (AP) is studied on graphite screen-printed electrodes, both with and without dissolved oxygen in the solution, to link their redox signals to their chemical structure. After elucidation of their reduction mechanisms using liquid chromatography coupled to high-resolution mass spectrometry, three structural classes (Class 1, Class 2 and Class 3) were defined, each with different redox centers and electrochemical behavior. Subsequently, 22 confiscated pills containing 14 different BZs were correctly assigned to these three structural classes, with the deoxygenated conditions displaying the highest class selectivity. Finally, the three classes were successfully detected after being spiked into five alcoholic beverages in the context of drug-facilitated sexual assault. For analysis in red wine, which complicated the analysis by interfering with Class 1, a dual test strategy in pH 2 and pH 7 was proposed for accurate detection. Its rapid measurements, broad scope and lack of interference from diluents or colors makes this method a promising approach for aiding various services in combating problematic BZ use.

Reactive oxygen species in hypertension.

Hypertension is a leading risk factor for stroke, heart disease and chronic kidney disease. Multiple interacting factors and organ systems increase blood pressure and cause target-organ damage. Among the many molecular elements involved in the development of hypertension are reactive oxygen species (ROS), which influence cellular processes in systems that contribute to blood pressure elevation (such as the cardiovascular, renal, immune and central nervous systems, or the renin-angiotensin-aldosterone system). Dysregulated ROS production (oxidative stress) is a hallmark of hypertension in humans and experimental models. Of the many ROS-generating enzymes, NADPH oxidases are the most important in the development of hypertension. At the cellular level, ROS influence signalling pathways that define cell fate and function. Oxidative stress promotes aberrant redox signalling and cell injury, causing endothelial dysfunction, vascular damage, cardiovascular remodelling, inflammation and renal injury, which are all important in both the causes and consequences of hypertension. ROS scavengers reduce blood pressure in almost all experimental models of hypertension; however, clinical trials of antioxidants have yielded mixed results. In this Review, we highlight the latest advances in the understanding of the role and the clinical implications of ROS in hypertension. We focus on cellular sources of ROS, molecular mechanisms of oxidative stress and alterations in redox signalling in organ systems, and their contributions to hypertension.

Mitochondria to plasma membrane redox signaling is essential for fatty acid β-oxidation-driven insulin secretion

We asked whether acute redox signaling from mitochondria exists concomitantly to fatty acid- (FA-) stimulated insulin secretion (FASIS) at low glucose by pancreatic β-cells. We show that FA β-oxidation produces superoxide/H2O2, providing: i) mitochondria-to-plasma-membrane redox signaling, closing KATP-channels synergically with elevated ATP (substituting NADPH-oxidase-4-mediated H2O2-signaling upon glucose-stimulated insulin secretion); ii) activation of redox-sensitive phospholipase iPLA2γ/PNPLA8, cleaving mitochondrial FAs, enabling metabotropic GPR40 receptors to amplify insulin secretion (IS). At fasting glucose, palmitic acid stimulated IS in wt mice; palmitic, stearic, lauric, oleic, linoleic, and hexanoic acids also in perifused pancreatic islets (PIs), with suppressed 1st phases in iPLA2γ/PNPLA8-knockout mice/PIs. Extracellular/cytosolic H2O2-monitoring indicated knockout-independent redox signals, blocked by mitochondrial antioxidant SkQ1, etomoxir, CPT1 silencing, and catalase overexpression, all inhibiting FASIS, keeping ATP-sensitive K+-channels open, and diminishing cytosolic [Ca2+]-oscillations. FASIS in mice was a postprandially delayed physiological event. Redox signals of FA β-oxidation are thus documented, reaching the plasma membrane, essentially co-stimulating IS.

Oxidative Cysteine Post Translational Modifications Drive the Redox Code Underlying Neurodegeneration and Amyotrophic Lateral Sclerosis.

Redox dysregulation, an imbalance between oxidants and antioxidants, is crucial in the pathogenesis of various neurodegenerative diseases. Within this context, the "redoxome" encompasses the network of redox molecules collaborating to maintain cellular redox balance and signaling. Among these, cysteine-sensitive proteins are fundamental for this homeostasis. Due to their reactive thiol groups, cysteine (Cys) residues are particularly susceptible to oxidative post-translational modifications (PTMs) induced by free radicals (reactive oxygen, nitrogen, and sulfur species) which profoundly affect protein functions. Cys-PTMs, forming what is referred to as "cysteinet" in the redox proteome, are essential for redox signaling in both physiological and pathological conditions, including neurodegeneration. Such modifications significantly influence protein misfolding and aggregation, key hallmarks of neurodegenerative diseases such as Alzheimer's, Parkinson's, and notably, amyotrophic lateral sclerosis (ALS). This review aims to explore the complex landscape of cysteine PTMs in the cellular redox environment, elucidating their impact on neurodegeneration at protein level. By investigating specific cysteine-sensitive proteins and the regulatory networks involved, particular emphasis is placed on the link between redox dysregulation and ALS, highlighting this pathology as a prime example of a neurodegenerative disease wherein such redox dysregulation is a distinct hallmark.

Phylogenetic Profiling Analysis of the Phycobilisome Revealed a Novel State-Transition Regulator Gene in Synechocystis sp. PCC 6803.

Phycobilisomes play a crucial role in the light-harvesting mechanisms of cyanobacteria, red algae and glaucophytes, but the molecular mechanism of their regulation is largely unknown. In the cyanobacterium, Synechocystis sp. PCC 6803, we identified slr0244 as a phycobilisome-related gene using phylogenetic profiling analysis, a method used to predict gene function based on comparative genomics. To investigate the physiological function of the slr0244 gene, we characterized slr0244 mutants spectroscopically. Disruption of the slr0244 gene impaired state transition, a process by which the distribution of light energy absorbed by the phycobilisomes between two photosystems is regulated in response to the changes in light conditions. The Slr0244 protein seems to act in the process of state transition, somewhere at or downstream of the sensing step of the redox state of the plastoquinone (PQ) pool. These findings, together with past reports describing the interaction of this gene product with thioredoxin and glutaredoxin, suggest that the slr0244 gene is a novel state-transition regulator that integrates the redox signal of PQ pools with that of the photosystem I-reducing side. The protein has two universal stress protein (USP) motifs in tandem. The second motif has two conserved cysteine residues found in USPs of other cyanobacteria and land plants. These redox-type USPs with conserved cysteines may function as redox regulators in various photosynthetic organisms. Our study also shows the efficacy of phylogenetic profiling analysis in predicting the function of cyanobacterial genes that have not been annotated so far.

Culture of Bovine Aortic Endothelial Cells in Galactose Media Enhances Mitochondrial Plasticity and Changes Redox Sensing, Altering Nrf2 and FOXO3 Levels.

Understanding the complex biological processes of cells in culture, particularly those related to metabolism, can be biased by culture conditions, since the choice of energy substrate impacts all of the main metabolic pathways. When glucose is replaced by galactose, cells decrease their glycolytic flux, working as an in vitro model of limited nutrient availability. However, the effect of these changes on related physiological processes such as redox control is not well documented, particularly in endothelial cells, where mitochondrial oxidation is considered to be low. We evaluated the differences in mitochondrial dynamics and function in endothelial cells exposed to galactose or glucose culture medium. We observed that cells maintained in galactose-containing medium show a higher mitochondrial oxidative capacity, a more fused mitochondrial network, and higher intercellular coupling. These factors are documented to impact the cellular response to oxidative stress. Therefore, we analyzed the levels of two main redox regulators and found that bovine aortic endothelial cells (BAEC) in galactose media had higher levels of FOXO3 and lower levels of Nrf2 than those in glucose-containing media. Thus, cultures of endothelial cells in a galactose-containing medium may provide a more suitable target for the study of in vitro mitochondrial-related processes than those in glucose-containing media; the medium deeply influences redox signaling in these cells.

Superoxide signal orchestrates tetrathiomolybdate-induced longevity via ARGK-1 in Caenorhabditis elegans

PurposeWhile reactive oxygen species (ROS) have been identified as key redox signaling agents contributing to aging process, which and how specific oxidants trigger healthy longevity remain unclear. This paper aimed to explore the precise role and signaling mechanism of superoxide (O2•−) in health and longevity. MethodsA tool for precise regulation of O2•− levels in vivo was developed based on the inhibition of superoxide dismutase 1 (SOD1) by tetrathiomolybdate (TM) in Caenorhabditis elegans (C. elegans). Then, we examined the effects of TM on lifespan, reproduction, lipofuscin accumulation, mobility, and stress resistance. Finally, the signaling mechanism for longevity induced by TM-O2•− was screened by transcriptome analysis and tested in sod-1 and argk-1 RNAi strains, sod-2, sod-3, and daf-16 mutants. ResultsTM promoted longevity in C. elegans with a concomitant extension of healthy lifespan as indicated by increasing fertility and mobility and reducing lipofuscin accumulation, as well as enhanced resistance to different abiotic stresses. Mechanically, TM could precisely regulate O2•− levels in nematodes via modulating SOD1 activity. An O2•− scavenger Mn(III)TBAP abolished TM-induced lifespan extension, while an O2•− generator paraquat at low concentration mimicked the life prolongation effects. The longevity in TM-treated worms was abolished by sod-1 RNAi but was not affected in sod-2 or sod-3 mutants. Further transcriptome analysis revealed arginine kinase ARGK-1 and its downstream insulin/insulin-like growth factor 1 signaling (IIS) as potential effectors for TM-O2•‾-induced longevity, and argk-1 RNAi or daf-16 mutant nullified the longevity. ConclusionsThese findings indicate that it is feasible to precisely control specific oxidant in vivo and O2•− orchestrates TM-induced health and longevity in C. elegans via ARGK-1-IIS axis.

Superoxide is an Intrinsic Signaling Molecule Triggering Muscle Hypertrophy.

Aims: Redox signaling plays a key role in skeletal muscle remodeling induced by exercise and prolonged inactivity, but it is unclear which oxidant triggers myofiber hypertrophy due to the lack of strategies to precisely regulate individual oxidants invivo. In this study, we used tetrathiomolybdate (TM) to dissociate the link between superoxide (O2•-) and hydrogen peroxide and thereby to specifically explore the role of O2•- in muscle hypertrophy in C2C12 cells and mice. Results: TM can linearly regulate intracellular O2•- levels by inhibition of superoxide dismutase 1 (SOD1). A 70% increase in O2•- levels in C2C12 myoblast cells and mice is necessary and sufficient for triggering hypertrophy of differentiated myotubes and can enhance exercise performance by more than 50% in mice. SOD1 knockout blocks TM-induced O2•- increments and thereby prevents hypertrophy, whereas SOD1 restoration rescues all these effects. Scavenging O2•- with antioxidants abolishes TM-induced hypertrophy and the enhancement of exercise performance, whereas the restoration of O2•- levels with a O2•- generator promotes muscle hypertrophy independent of SOD1 activity. Innovation and Conclusion: These findings suggest that O2•- is an endogenous initiator of myofiber hypertrophy and that TM may be used to treat muscle wasting diseases. Our work not only suggests a novel druggable mechanism to increase muscle mass but also provides a tool for precisely regulating O2•- levels invivo.

The Emergence of Tumor-Initiating Cells in an Advanced Hypopharyngeal Tumor Model Exhibits Enhanced Angiogenesis and Nuclear Factor Erythroid 2-Related Factor 2-Associated Antioxidant Effects.

Aims: Hypopharyngeal cancer (HPC) is associated with the worst prognosis of all head and neck cancers and is typically identified in an advanced stage at the time of diagnosis. While oxidative stress might contribute to the onset of HPC in patients using tobacco or alcohol, the extent of this influence and the characteristics of HPC cells in advanced stage remain to be investigated. In this study, we explored whether HPC cells survived from necrotic xenograft tumors at late stage would display increased tumor resistance along with altered tolerance to oxidative stress. Results: The remnant living HPC cells isolated from a late-stage xenograft tumor, named FaDu ex vivo cells, showed stronger chemo- and radioresistance, tumorigenesis, and invasiveness compared with parental FaDu cells. FaDu ex vivo cells also displayed increased angiogenic ability after re-transplantation in mice visualized by invivo near infrared-II fluorescence imaging modality. Moreover, FaDu ex vivo cells exhibited significant tumor-initiating cell (TIC)-related properties accompanied by a reduction of the level of reactive oxygen species, which was associated with the upregulation of transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2). Interestingly, inhibition of Nrf2 by the RNA interference and the chemical inhibitor could reduce the TIC-related properties of FaDu ex vivo cells. Innovation: Oxidative stress potentially initiates HPC, but elevation of Nrf2-associated antioxidant mechanisms would be essential to mitigate this effect for promoting and sustaining the stemness of HPC at the advanced stage. Conclusion: Present data suggest that the antioxidant potency of advanced HPC would be a therapeutic target for the design of adjuvant treatment. Antioxid. Redox Signal. 41, 505-521.

Ferroptosis-inducing compounds synergize with docetaxel to overcome chemoresistance in docetaxel-resistant non-small cell lung cancer cells

Development of resistance to therapy-induced cell death is a major hurdle in the effective treatment of advanced solid tumors. Erastin and RSL3 were originally found to induce synthetic lethality by induction of a novel form of cell death termed ferroptosis. Emerging evidence suggests that ferroptosis inducers enhance chemosensitivity of classic therapeutic agents by triggering ferroptotic cell death. In this study we evaluated the effects of erastin and RSL3 on the resistance of docetaxel, doxorubicin, and cisplatin, and revealed a mechanism whereby these ferroptosis inducers augment docetaxel efficacy in non-small cell lung cancer by regulating redox signaling to promote ferroptosis. Transcriptome analysis revealed that combination treatment modulated not only p53 signaling pathway but also immune responses and several signaling pathways including MAPK, NF-κB and PI3K/Akt. Considering that glutathione peroxidase 4 (GPX4) serves as the main effector to protect cells from ferroptosis, this study identified three novel non-covalent GPX4 inhibitors with the aid of pharmacophore-based virtual screening. The new ferroptosis-inducing compounds synergized with docetaxel to increase the cytotoxicity by promoting ferroptotic cell death in docetaxel-resistant A549/DTX cells. Collectively, the induction of ferroptosis contributed to docetaxel-induced cytotoxic effects and overcame drug resistance in A549/DTX cells. Ferroptosis has a great potential to become a new approach to attenuate resistance to some classic therapeutic drugs in cancer patients.

Targeting ROS in cancer: rationale and strategies.

Reactive oxygen species (ROS) in biological systems are transient but essential molecules that are generated and eliminated by a complex set of delicately balanced molecular machineries. Disruption of redox homeostasis has been associated with various human diseases, especially cancer, in which increased ROS levels are thought to have a major role in tumour development and progression. As such, modulation of cellular redox status by targeting ROS and their regulatory machineries is considered a promising therapeutic strategy for cancer treatment. Recently, there has been major progress in this field, including the discovery of novel redox signalling pathways that affect the metabolism of tumour cells as well as immune cells in the tumour microenvironment, and the intriguing ROS regulation of biomolecular phase separation. Progress has also been made in exploring redox regulation in cancer stem cells, the role of ROS in determining cell fate and new anticancer agents that target ROS. This Review discusses these research developments and their implications for cancer therapy and drug discovery, as well as emerging concepts, paradoxes and future perspectives.

MicroRNAs as Regulators of Radiation-Induced Oxidative Stress.

microRNAs (miRNAs) represent small RNA molecules involved in the regulation of gene expression. They are implicated in the regulation of diverse cellular processes ranging from cellular homeostasis to stress responses. Unintended irradiation of the cells and tissues, e.g., during medical uses, induces various pathological conditions, including oxidative stress. miRNAs may regulate the expression of transcription factors (e.g., nuclear factor erythroid 2 related factor 2 (Nrf2), nuclear factor kappa B (NF-κB), tumor suppressor protein p53) and other redox-sensitive genes (e.g., mitogen-activated protein kinase (MAPKs), sirtuins (SIRTs)), which trigger and modulate cellular redox signaling. During irradiation, miRNAs mainly act with reactive oxygen species (ROS) to regulate the cell fate. Depending on the pathway involved and the extent of oxidative stress, this may lead to cell survival or cell death. In the context of radiation-induced oxidative stress, miRNA-21 and miRNA-34a are among the best-studied miRNAs. miRNA-21 has been shown to directly target superoxide dismutase (SOD), or NF-κB, whereas miRNA-34a is a direct regulator of NADPH oxidase (NOX), SIRT1, or p53. Understanding the mechanisms underlying radiation-induced injury including the involvement of redox-responsive miRNAs may help to develop novel approaches for modulating the cellular response to radiation exposure.

Co-crosslinking strategy for dual functionalization of small magnetic nanoparticles with redox probes and biological probes

Surface functionalization strategy is becoming a crucial bridge from magnetic nanoparticles (MNPs) to their broad bio-application. To realize the multiple functions of MNPs such as magnetic manipulation, target capture, and signal amplification in their use of electrochemical biosensing, co-crosslinking strategy was proposed here to construct dual-functionalized MNPs by combining ultra-sensitive redox moieties and specific biological probes. In this work, MNPs with a TEM size of 10 nm were synthesized by co-precipitation for amination and PEGylation to maintain colloid stability once dispersed in high-ionic-strength buffer (such as phosphate-buffered saline). Then, MNPs@IgG were prepared via the bis(sulfosuccinimidyl) suberate (BS3) cross-linker to conjugate these IgG onto the MNP surface, with a binding efficiency of 73%. To construct dual-functionalized MNPs, these redox probes of ferrocene-NHS (Fc) were co-crosslinked onto the MNP surface, together with IgG, by using BS3. The developed MNPs@Redox@IgG were characterized by SDS‒PAGE to identify IgG binding and by square wave voltammetry (SWV) to validate the redox signal. Additionally, the anti-CD63 antibodies were selected for the development of MNPs@anti-CD63 for use in the bio-testing of exosome sample capture. Therefore, co-crosslinking strategy paved a way to develop dual-functionalized MNPs that can be an aid of their potential utilization in diagnostic assay or electrochemical methods.Graphical

Unveiling novel targets in melanoma under melanogenesis stimulation and photodynamic therapy by redox proteomics.

Melanogenesis-stimulated B16-F10 cells enter in a quiescent state, present inhibited mitochondrial respiration and increased reactive oxygen species levels. These alterations suggest that these cells may be under redox signaling, allowing tumor survival. The aim of this study was to evaluate redox-modified proteins in B16-F10 cells after melanogenesis stimulation and rose bengal-photodynamic therapy (RB-PDT). A redox proteomics label-free approach based on the biotin switch assay technique with biotin-HPDP and N-ethylmaleimide was used to assess the thiol-oxidized protein profile. Aconitase was oxidized at Cys-448 and Cys-451, citrate synthase was oxidized at Cys-202 and aspartate aminotransferase (Got2) was oxidized at Cys-272 and Cys-274, exclusively after melanogenesis stimulation. After RB-PDT, only guanine nucleotide-binding protein subunit beta-2-like 1 (Gnb2l1) was oxidized (Cys-168). In contrast, melanogenesis stimulation followed by RB-PDT led to the oxidation of different cysteines in Gnb2l1 (Cys-153 and Cys-249). Besides that, glyceraldehyde-3-phosphate dehydrogenase (Gapdh) presented oxidation at Cys-245, peptidyl-prolyl cis-trans isomerase A (Ppia) was oxidized at Cys-161 and 5,6-dihydroxyindole-2-carboxylic acid oxidase (Tyrp1) was oxidized at Cys-65, Cys-30, and Cys-336 after melanogenesis stimulation followed by RB-PDT. The redox alterations observed in murine melanoma cells and identification of possible target proteins are of great importance to further understand tumor resistance mechanisms.