Induced Reactive Oxygen Species Generation Research Articles

ASLdC3: A Derivative of Acidic Sophorolipid Disrupts Mitochondrial Function, Induces ROS Generation, and Inhibits Biofilm Formation in Candida albicans.

Fungal infections account for more than 140 million cases of severe and life-threatening conditions each year, causing approximately 1.7 million deaths annually. Candida albicans and related species are the most common human fungal pathogens, causing both superficial (mucosal and cutaneous) and life-threatening invasive infections (candidemia) with a 40-75% mortality rate. Among many virulence factors of Candida albicans, morphological transition from yeast to hyphae, secretion of hydrolytic enzymes, and formation of biofilms are considered to be crucial for pathogenicity. However, the arsenals for the treatment against these pathogens are restricted to only a few classes of approved drugs, the efficacy of which is being compromised by host toxicity, fungistatic activity, and the emergence of drug resistance. In this study, we have described the development of a molecule, exhibiting excellent antifungal activity (MIC 8 μg/mL), by tailoring acidic sophorolipids with aryl alcohols via enzyme catalysis. This novel derivative, ASLdC3, is a surface-active compound that lowers the surface tension of the air-water interface up to 2-fold before reaching the critical micelle concentration of 25 μg/mL. ASLdC3 exhibits excellent antibiofilm properties against Candida albicans and other nonalbicans Candida species. The molecule primarily exhibits its antifungal activity by perturbing mitochondrial function through the alteration of the mitochondrial membrane potential (MMP) and generation of reactive oxygen species (ROS). The ROS damages fungal cell membrane function and cell wall integrity, eventually leading to cell death. ASLdC3 was found to be nontoxic in in vitro assay and nonhemolytic. Besides, it does not cause toxicity in the C. elegans model. Our study provides a valuable foundation for the potential of acidic sophorolipid as a nontoxic, biodegradable precursor for the design and synthesis of novel molecules for use as antimicrobial drugs as well as for other clinical applications.

Reactive Oxygen Species (ROS)-Mediated Antibacterial Oxidative Therapies: Available Methods to Generate ROS and a Novel Option Proposal.

The increasing emergence of multidrug-resistant (MDR) pathogens causes difficult-to-treat infections with long-term hospitalizations and a high incidence of death, thus representing a global public health problem. To manage MDR bacteria bugs, new antimicrobial strategies are necessary, and their introduction in practice is a daily challenge for scientists in the field. An extensively studied approach to treating MDR infections consists of inducing high levels of reactive oxygen species (ROS) by several methods. Although further clinical investigations are mandatory on the possible toxic effects of ROS on mammalian cells, clinical evaluations are extremely promising, and their topical use to treat infected wounds and ulcers, also in presence of biofilm, is already clinically approved. Biochar (BC) is a carbonaceous material obtained by pyrolysis of different vegetable and animal biomass feedstocks at 200-1000 °C in the limited presence of O2. Recently, it has been demonstrated that BC's capability of removing organic and inorganic xenobiotics is mainly due to the presence of persistent free radicals (PFRs), which can activate oxygen, H2O2, or persulfate in the presence or absence of transition metals by electron transfer, thus generating ROS, which in turn degrade pollutants by advanced oxidation processes (AOPs). In this context, the antibacterial effects of BC-containing PFRs have been demonstrated by some authors against Escherichia coli and Staphylococcus aureus, thus giving birth to our idea of the possible use of BC-derived PFRs as a novel method capable of inducing ROS generation for antimicrobial oxidative therapy. Here, the general aspects concerning ROS physiological and pathological production and regulation and the mechanism by which they could exert antimicrobial effects have been reviewed. The methods currently adopted to induce ROS production for antimicrobial oxidative therapy have been discussed. Finally, for the first time, BC-related PFRs have been proposed as a new source of ROS for antimicrobial therapy via AOPs.

Visible and Red Light-Triggered Anticancer Profile of a Ferrocene-Re(I)-Tricarbonyl Conjugate: Experimental and Theoretical Studies.

We have red-shifted the light absorbance property of a Re(I)-tricarbonyl complex via distant conjugation of a ferrocene moiety and developed a novel complex ReFctp, [Re(Fctp)(CO)3Cl], where Fctp = 4'-ferrocenyl-2,2':6',2″-terpyridine. ReFctp showed green to red light absorption ability and blue emission, indicating its potential for photodynamic therapy (PDT) application. The conjugation of ferrocene introduced ferrocene-based transitions, which lie at a higher wavelength within the PDT therapeutic window. The time-dependent density functional theory and excited state calculations revealed an efficient intersystem crossing for ReFctp, which is helpful for PDT. ReFctp elicited both PDT type I and type II pathways for reactive oxygen species (ROS) generation and facilitated NADH (1,4-dihydro-nicotinamide adenine dinucleotide) oxidation upon exposure to visible light. Importantly, ReFctp showed effective penetration through the layers of clinically relevant 3D multicellular tumor spheroids and localized primarily in mitochondria (Pearson's correlation coefficient, PCC = 0.65) of A549 cancer cells. ReFctp produced more than 20 times higher phototoxicity (IC50 ∼1.5 μM) by inducing ROS generation and altering mitochondrial membrane potential in A549 cancer cells than the nonferrocene analogue Retp, [Re(CO)3(tp)Cl], where tp = 2,2':6',2″-terpyridine. ReFctp induced apoptotic mode of cell death with a notable photocytotoxicity index (PI, PI = IC50dark/IC50light) and selectivity index (SI, SI = normal cell's IC50dark/cancer cell's IC50light) in the range of 25-33.

Glaesserella parasuis serotype 5 induces pyroptosis via the RIG-I/MAVS/NLRP3 pathway in swine tracheal epithelial cells

Glaesserella parasuis (G. parasuis) is a common Gram-negative commensal bacterium in the upper respiratory tract of swine that can cause Glässer's disease under stress conditions. Pyroptosis is an important immune defence mechanism of the body that plays a crucial role in clearing pathogen infections and endogenous danger signals. This study aimed to investigate the mechanism of G. parasuis serotype 5 SQ (GPS5-SQ)-induced pyroptosis in swine tracheal epithelial cells (STECs). The results of the present study demonstrated that GPS5-SQ infection induces pyroptosis in STECs by enhancing the protein level of the N-terminal domain of gasdermin D (GSDMD-N) and activating the NOD-like receptor protein 3 (NLRP3) inflammasome. Furthermore, the levels of pyroptosis-related proteins, including GSDMD-N and cleaved caspase-1 were considerably decreased in STECs after the knockdown of retinoic acid inducible gene-I (RIG-I) and mitochondrial antiviral signaling protein (MAVS). These results indicated that GPS5-SQ might trigger pyroptosis through the activation of the RIG-I/MAVS/NLRP3 signaling pathway. More importantly, the reactive oxygen species (ROS) scavenger N-acetylcysteine (NAC) repressed the activation of the RIG-I/MAVS/NLRP3 signaling and rescued the decrease in Occludin and zonula occludens-1 (ZO-1) after GPS5-SQ infection. Overall, our findings show that GPS5-SQ can activate RIG-I/MAVS/NLRP3 signaling and destroy the integrity of the epithelial barrier by inducing ROS generation in STECs, shedding new light on G. parasuis pathogenesis.

PAD4 Inhibitor-Functionalized Layered Double Hydroxide Nanosheets for Synergistic Sonodynamic Therapy/Immunotherapy Of Tumor Metastasis.

Sonodynamic therapy (SDT) is demonstrated to trigger the systemic immune response of the organism and facilitate the treatment of metastatic tumors. However, SDT-mediated neutrophil extracellular traps (NETs) formation can promote tumor cell spread, thus weakening the therapeutic effectiveness of metastatic tumors. Herein, the amorphous CoW-layered double hydroxide (a-CoW-LDH) nanosheets are functionalized with a peptidyl arginine deiminase 4 (PAD4) inhibitor, i.e., YW3-56, to construct a multifunctional nanoagent (a-LDH@356) for synergistic SDT/immunotherapy. Specifically, a-CoW-LDH nanosheets can act as a sonosensitizer to generate abundant reactive oxygen species (ROS) under US irradiation. After loading with YW3-56, a-LDH@356 plus US irradiation not only effectively induces ROS generation and immunogenic cell death, but also inhibits the elevation of citrullinated histone H3 (H3cit) and the release of NETs, enabling a synergistic enhancement of anti-tumor metastasis effect. Using 4T1 tumor model, it is demonstrated that combining a-CoW-LDH with YW3-56 stimulates an anti-tumor response by upregulating the proportion of immune-activated cells and inducing polarization of M1 macrophages, and inhibits immune escape by downregulating the expression of PD-1 on immune cells under US irradiation, which not only arrests primary tumor progression with a tumor inhibition rate of 69.5% but also prevents tumor metastasis with the least number of lung metastatic nodules.

Marine-derived antimicrobial peptide piscidin-1 triggers extrinsic and intrinsic apoptosis in oral squamous cell carcinoma through reactive oxygen species production and inhibits angiogenesis

Cancer of the head and neck encompasses a wide range of cancers, including oral and oropharyngeal cancers. Oral cancer is often diagnosed at advanced stages and has a dismal prognosis. Piscidin-1, a marine antimicrobial peptide (AMP) containing approximately 22 amino acids, also exhibits significant anticancer properties. We investigated the possible anti-oral cancer effects of piscidin-1 and clarified the mechanisms underlying these effects. We treated the oral squamous cell carcinoma cell lines OC2 and SCC4 with piscidin-1. Cell viability and the expression of different hallmark apoptotic molecules, including reactive oxygen species (ROS), were tested using the appropriate MTT assay, flow cytometry and western blotting assays, and human umbilical vein endothelial cell (HUVEC) wound healing, migration, and tube formation (angiogenesis) assays. Piscidin-1 increases cleaved caspase 3 levels to induce apoptosis. Piscidin-1 also increases ROS levels and intensifies oxidative stress in the endoplasmic reticulum and mitochondria, causing mitochondrial dysfunction. Additionally, it decreases the oxygen consumption rates and activity of mitochondrial complexes I–V. As expected, the antioxidants MitoTEMPOL and N-acetylcysteine reduce piscidin-1–induced ROS generation and intracellular calcium accumulation. Piscidin-1 also inhibits matrix metalloproteinase (MMP)-2/-9 expression in HUVECs, affecting migration and tube formation angiogenesis. We demonstrated that piscidin-1 can promote apoptosis via both intrinsic and extrinsic apoptotic pathways and findings indicate that piscidin-1 has anti-proliferative and anti-angiogenic properties in oral cancer treatment. Our study on piscidin-1 thus provides a basis for future translational anti-oral cancer drug research and a new theoretical approach for anti-oral cancer clinical research.

Dendritic Polylysine with Paclitaxel and Triptolide Codelivery for Enhanced Cancer Ferroptosis through the Accumulation of ROS.

Recently, paclitaxel (PTX) was reported to increase intracellular lipid reactive oxygen species (ROS) levels, triggering cancer cell ferroptosis. Based on this, some efforts had been made to improve PTX treatment for non-small-cell lung cancer (NSCLC). Our previous studies demonstrated that triptolide (TPL) could improve the antitumor effect of PTX. Nevertheless, the poor solubility and side effects often limit the application of chemotherapy drugs. In this paper, we constructed a novel nanodrug delivery system (NDDS) chemosynthesis by PEGylated generation 3 (G3) dendritic polylysine coloaded with PTX and TPL (PTX-TPL-PEG-PLL, PTPP), which was endowed with the ability of tumor targeting and favorable solubility. In addition, we demonstrated that TPL could induce ROS generation by regulating the NF-κB signaling pathway to enhance the ferroptosis-induced effect of PTX. Besides, ferroptosis induced by PTPP could improve chemoresistance through inhibiting the level of P-gp, GPX4, and SLC7A11. Furthermore, we determined that ferroptosis may strengthen the immune response by increasing the expression of CD8+ T cells and IFN-γ+ cells while decreasing Treg cells. In general, PTPP may be a potential system for NSCLC treatment.

Effects of photo-oxidation and transition metals on the formation of reactive oxygen species from aromatic compounds using spectroscopic method

Particulate matter (PM) can cause adverse health effects by overproducing reactive oxygen species (ROS). Although the ability of PM to induce ROS generation depends on its composition and environmental factors. This study explores how photo-oxidation affects ROS generation from aromatic compounds (ACs, including catechol (CAT), phthalic acid (PA), and 4,4′-oxydibenzoic acid (4,4′-OBA)) and their mixtures with transition metals (TMs, including Fe(II), Mn(II), and Cu(II)) using Fourier-transform infrared (FTIR) and Ultraviolet-visible spectroscopy (UV–Vis). Results showed that photo-oxidation facilitated ROS generation from ACs. CAT-Fe(II)/Cu(II) showed synergistic effects, but 4,4′-OBA-Fe(II)/Cu(II) showed antagonistic effects. ACs-Mn(II) and PA-Fe(II)/Cu(II) exhibited synergistic effects first and then showed antagonistic effects. The different interactions were due to complexation between ACs and TMs. The photo-oxidized ACs-TMs significantly enhanced ROS generation compared with ACs-TMs. The study suggested the photo-oxidation mechanism involved that the transfer of π-electrons from the ground to an excited state in benzene rings and functional groups, leading to the breakage and formation of chemical bonds or easier π-electron transfer from ACs to TMs. The former could generate ROS directly or produce polymers that promoted ROS generation, while the latter promoted ROS generation by transferring π-electrons to dissolved oxygen quickly. Our study revealed that both interactions among components and photo-oxidation significantly influenced ROS generation. Future studies should integrate broader atmospheric factors and PM components to fully assess oxidative potential and health impacts.

Forced expression of microRNA-221-3p exerts protective effects against manganese-induced cytotoxicity in human lung epithelial cells

Manganese (Mn)-induced pulmonary toxicity and the underlying molecular mechanisms remain largely enigmatic. Further, in recent years, microRNAs (miRNAs) have emerged as regulators of several pollutants-mediated toxicity. In this context, our study aimed at elucidating whether miRNAs are involved in manganese (II) chloride (MnCl2) (Mn2+)-induced cytotoxicity in lung epithelial cells. Growth inhibition of Mn2+ towards normal human bronchial epithelial (BEAS-2B) and adenocarcinomic human alveolar basal epithelial (A549) cells was analyzed by MTT assay following 24 or 48 h treatment. Reactive oxygen species (ROS) generation, mitochondrial membrane potential (ΔΨm), cell cycle arrest, and apoptosis were evaluated by flow cytometry. RT-qPCR and Western blot were performed to analyze the expression of cyclins, anti-oxidant genes, and miRNAs. We used small RNA sequencing to investigate Mn2+-induced changes in miRNA expression patterns. In both cell lines, Mn2+ treatment inhibited growth in a dose-dependent manner. Further, compared with vehicle-treated cells, Mn2+ (250 μM) treatment induced ROS generation, cell cycle arrest, apoptosis, and decreased ΔΨm as well as altered the expression of cyclins and anti-oxidant genes. Sequencing data revealed that totally 296 miRNAs were differentially expressed in Mn2+-treated cells. Among them, miR-221-3p was one of the topmost down-regulated miRNAs in Mn2+-treated cells. We further confirmed this association in A549 cells. In addition, transient transfection was performed to study gain-of-function experiments. Forced expression of miR-221-3p significantly improved cell viability and reduced Mn2+-induced cell cycle arrest and apoptosis in BEAS-2B cells. In conclusion, miR-221-3p may be the most likely target that accounts for the cytotoxicity of Mn2+−exposed lung epithelial cells.

Microplastics induced apoptosis in macrophages by promoting ROS generation and altering metabolic profiles

The ubiquitous presence of Microplastics (MPs) in various environments documented in recent years has recently raised significant concerns about their toxic effects. While macrophages serve as the first line of defense against toxic substances and pathogens, the impact and mechanisms of microplastics on these immune cells remain unclear. This study aims to explore whether MPs induce macrophage apoptosis through the promotion of reactive oxygen species (ROS) generation and alterations in metabolic profiles. The viability of RAW264.7 cells decreased as the concentration of 0.5 µm or 5 µm MPs ranged from 0.2 to 1.5 mg/mL, with a more pronounced effect observed in the 0.5 µm MPs group. Zebrafish exposed to 0.5 µm or 5 µm MPs at a concentration of 0.5 mg/mL exhibited decreased macrophage abundance and increased apoptosis, accompanied by alterations in the expression of inflammatory and apoptosis-related genes. While 0.5 µm MPs were observed to enter macrophages, 5 µm MPs only adhered to the cell membrane surface. Both particle sizes induced ROS generation and disrupted cellular metabolism in RAW264.7 cells. Notably, macrophages exhibited a more pronounced response to 0.5 µm MPs, characterized by heightened ROS generation, increased secretion of pro-inflammatory mediators, and a significant decrease in sphingolipid metabolism. These findings suggest that the adverse effects on macrophages are greater with 0.5 µm MPs compared to 5 µm MPs, possibly attributed to particle size effects. This study contributes additional evidence on the impact of MPs on human immune cells.

Antitumor effects of a novel photosensitizer-mediated photodynamic therapy and its influence on the cell transcriptome.

Photodynamic therapy (PDT) is a promising cancer treatment. This study investigated the antitumor effects and mechanisms of a novel photosensitizer meso-5-[ρ-diethylene triamine pentaacetic acid-aminophenyl]-10,15,20-triphenyl-porphyrin (DTP) mediated PDT (DTP-PDT). Cell viability, reactive oxygen species (ROS), and apoptosis were measured with a Cell Counting Kit-8 assay, DCFH-DA fluorescent probe, and Hoechst staining, respectively. Cell apoptosis- and autophagy-related proteins were examined using western blotting. RNA sequencing was used to screen differentially expressed mRNAs (DERs), and bioinformatic analysis was performed to identify the major biological events after DTP-PDT. Our results show that DTP-PDT inhibited cell growth and induced ROS generation in MCF-7 and SGC7901 cells. The ROS scavenger N-acetyl-L-cysteine (NAC) and the P38 MAPK inhibitor SB203580 alleviated DTP-PDT-induced cytotoxicity. DTP-PDT induced cell apoptosis together with upregulated Bax and downregulated Bcl-2, which could also be inhibited by NAC or SB203580. The level of LC3B-II, a marker of autophagy, was increased by DTP-PDT. A total of 3496 DERs were obtained after DTP-PDT. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses indicated that DERs included those involved in cytosolic ribosomes, the nuclear lumen, protein binding, cell cycle, protein targeting to the endoplasmic reticulum, and ribosomal DNA replication. Disease Ontology and Reactome enrichment analyses indicated that DERs were associated with a variety of cancers and cell cycle checkpoints. Protein-protein interaction results demonstrated that cdk1 and rps27a ranked in the top 10 interacting genes. Therefore, DTP-PDT could inhibit cell growth and induce cell apoptosis and autophagy, partly through ROS and the P38 MAPK signaling pathway. Genes associated with the cell cycle, ribosomes, DNA replication, and protein binding may be the key changes in DTP-PDT-mediated cytotoxicity.

Heavy-atom-free BODIPY dendrimer: utilizing the spin-vibronic coupling mechanism for two-photon photodynamic therapy in zebrafish.

In this study, the heavy-atom-free BODIPY dendrimer TM4-BDP was synthesized for near-infrared photodynamic therapy, and was composed of a triphenylamine-BODIPY dimer and four 1-(2-morpholinoethyl)-1H-indole-3-ethenyl groups. The TM4-BDP could achieve near-infrared photodynamic therapy through two different photosensitive pathways, which include one-photon excitation at 660 nm and two-photon excitation at 1000 nm. In the one-photon excitation pathway, the TM4-BDP could generate singlet oxygen and superoxide radicals under 660 nm illumination. In addition, the one-photon PDT experiment in human nasopharyngeal carcinoma (CNE-2) cells also indicated that the TM4-BDP could specifically accumulate in lysosomes and show great cell phototoxicity with an IC50 of 22.1 μM. In the two-photon excitation pathway, the two-photon absorption cross-section at 1030 nm of TM4-BDP was determined to be 383 GM, which means that it could generate reactive oxygen species (ROS) under 1000 nm femtosecond laser excitation. Moreover, the two-photon PDT experiment in zebrafish also indicated the TM4-BDP could be used for two-photon fluorescence imaging and two-photon induced ROS generation in biological environments. Furthermore, in terms of the ROS generation mechanism, the TM4-BDP employed a novel spin-vibronic coupling intersystem crossing (SV-ISC) process for the mechanism of ROS generation and the femtosecond transient absorption spectra indicated that this novel SV-ISC mechanism was closely related to its charge transfer state lifetime. These above experiments of TM4-BDP demonstrate that the dendrimer design is an effective strategy for constructing heavy-atom-free BODIPY photosensitizers in the near-infrared region and lay the foundation for two-photon photodynamic therapy in future clinical trials.

Melatonin prevents overproduction of reactive oxygen species and vascular dysfunction induced by cyclophosphamide

AimsOverproduction of reactive oxygen species (ROS) is a pathologic hallmark of cyclophosphamide toxicity. For this reason, antioxidant compounds emerge as promising tools for preventing tissue damage induced by cyclophosphamide. We hypothesized that melatonin would display cytoprotective action in the vasculature by preventing cyclophosphamide-induced oxidative stress. Materials and methodsMale C57BL/6 mice (22–25 g) were injected with a single dose of cyclophosphamide (300 mg/kg; i.p.). Mice were pretreated or not with melatonin (10 mg/kg/day, i.p.), given during 4 days before cyclophosphamide injection. Functional (vascular reactivity) and oxidative/inflammatory patterns were evaluated at 24 h in resistance arteries. The antioxidant action of melatonin was assessed in vitro in cultured vascular smooth muscle cells (VSMCs) of mesenteric arteries. Key findingsCyclophosphamide induced ROS generation in both mesenteric arterial bed (MAB) and cultured VSMCs, and this was normalized by melatonin. Cyclophosphamide-induced ROS generation and lipoperoxidation in the bladder and kidney was also prevented by melatonin. Increased levels of tumor necrosis factor (TNF)-α and interleukin (IL)-6 were detected in the MAB of cyclophosphamide-treated mice, all of which were prevented by melatonin. Functional assays using second-order mesenteric arteries of cyclophosphamide-treated mice revealed a decrease in vascular contractility. Melatonin prevented vascular hypocontractility in the cyclophosphamide group. Melatonin partially prevented the decrease in myeloperoxidase (MPO) and N-acetyl-beta-D-glucosaminidase (NAG) activities in the MAB of the cyclophosphamide group. SignificanceMelatonin may constitute a novel and promising therapeutic approach for management of the toxic effects induced by cyclophosphamide in the vasculature.

MicroRNA-195-5p mediates arsenic-induced cytotoxicity in human lung epithelial cells: Beneficial role of plant-derived tannic acid

Arsenic (As), a highly toxic metalloid, which causes environmental lung diseases and affects millions of people worldwide. Respiratory epithelial cells are essential for maintaining lung homeostasis, aberrant epithelial damage and death due to exposure to a wide range of environmental pollutants, which are considered to be the initial trigger for many pulmonary diseases. Accumulating evidence has shown that microRNAs (miRNAs) appear to be important players in various normal physiological and pathological processes. Therefore, the present study was carried out to examine the cytotoxic effects of a trivalent form of As (As3+) in normal human bronchial (BEAS-2B) and adenocarcinoma alveolar basal (A549) epithelial cells and the role of miR-195-5p. Further, we also explored the protective effects of a natural dietary polyphenol tannic acid (TA). As3+ (1 μM) treatment in BEAS-2B cells for 24 h induced cytotoxicity by decreasing the cell viability, mitochondrial membrane potential (ΔΨm) and inducing reactive oxygen species (ROS) generation, lipid peroxidation (LPO), cell cycle arrest, and apoptosis, which was associated with a significantly higher level of miR-195-5p expression compared with vehicle control. Forced expression of miR-195-5p alone suppressed cell survival, ΔΨm, regulated cell cycle distribution and induced ROS generation in BEAS-2B cells. As expected, miR-195-5p inhibition effectively rescued BEAS-2B cells from As3+-mediated toxicity, confirming the involvement of miR-195-5p in the cytotoxic effects of As3+. Further, TA pre-treatment expressively alleviated As3+-induced toxicity by suppressing ROS production, miR-195-5p expression, and increasing ΔΨm. These in vitro results indicate that miR-195-5p may be useful as a therapeutic target for treating As3+ toxicity.

Synergistic effects of CuO/TiO2 -chitosan-farnesol nanocomposites: Synthesis, characterization, antimicrobial, and anticancer activities on melanoma cells SK-MEL-3.

The current investigation focuses on synthesizing copper oxide (CuO)-titanium oxide (TiO2 )-chitosan-farnesol nanocomposites with potential antibacterial, antifungal, and anticancer properties against Melanoma cells (melanoma cells [SK-MEL-3]). The nanocomposites were synthesized using the standard acetic acid method and subsequently characterized using an X-ray diffractometer, scanning electron microscope, transmission electron microscopy, and Fourier transform infrared spectroscopy. The results from the antibacterial tests against Streptococcus pneumoniae and Stapylococcus aureus demonstrated significant antibacterial efficacy. Additionally, the antifungal studies using Candida albicans through the agar diffusion method displayed a considerable antifungal effect. For evaluating the anticancer activity, various assays such as MTT assay, acridine orange/ethidium bromide dual staining assay, reactive oxygen species (ROS) generation assay, and mitochondrial membrane potential (MMP) analysis were conducted on SK-MEL-3 cells. The nanocomposites exhibited the ability to induce ROS generation, decrease MMP levels, and trigger apoptosis in SK-MEL-3 cells. Collectively, the findings demonstrated a distinct pattern for the synthesized bimetallic nanocomposites. Furthermore, these nanocomposites also displayed significant (p < 0.05) antibacterial, antifungal, and anticancer effects when tested on the SK-MEL-3 cell line.

Diselenium-linked dimeric prodrug nanomedicine breaking the intracellular redox balance for triple-negative breast cancer targeted therapy

Triple-negative breast cancer (TNBC) has been regarded as the strongest malignancy in cases of breast cancer with a poor prognosis. The development of effective treatment strategies for TNBC has always been an urgent and unmet need. The intracellular redox balance is essential for maintaining TNBC cell malignancy. Disrupting intracellular redox balance by enlarging reactive oxygen species (ROS) generation and facilitating glutathione (GSH) depletion to amplify intracellular oxidative stress may be an alternative strategy to eliminate TNBC cells. However, inducing ROS generation and GSH depletion concurrently may be challenging. Herein, a diselenium linked-dimeric prodrug nanomedicine FA-SeSe-NPs was developed to break the intracellular redox homeostasis for TNBC targeted therapy. The dimeric prodrug was synthesized by conjugating two cucurbitacin B (CuB) molecules via one diselenium bond, which was subsequently assembled with FA-PEG-DSPE to form the final nanomedicine FA-SeSe-NPs. Using the active targeting potential of folic acid (FA), FA-SeSe-NPs could accumulate in tumor tissue with elevated levels and then be specifically internalized by cancer cells. In the high ROS and GSH conditions of TNBC cells, the diselenium bond can specifically respond to ROS to produce selenium free radicals to increase ROS and react with GSH to generate S-Se bond to deplete GSH. The released CuB further induced ROS production in TNBC cells. The diselenium bond and CuB functioned synergistically to amplify oxidative stress to kill the TNBC cells. Here, we provide a promising strategy to disrupt the intracellular redox balance of cancer cells for effective TNBC therapy.

Zinc Overload Induces Damage to H9c2 Cardiomyocyte Through Mitochondrial Dysfunction and ROS-Mediated Mitophagy.

Zinc homeostasis is essential for maintaining redox balance, cell proliferation, and apoptosis. However, excessive zinc exposure is toxic and leads to mitochondrial dysfunction. In this study, we established a zinc overload model by treating rat cardiomyocyte H9c2 cells with Zn2+ at different concentrations. Our results showed that zinc overload increased LDH and reactive oxygen species (ROS) levels, leading to cell death, mitochondrial membrane potential decrease and impaired mitochondrial function and dynamics. Furthermore, zinc overload activated the PINK1/Parkin signaling pathway and induced mitochondrial autophagy via ROS, while NAC inhibited mitophagy and weakened the activation of PINK1/Parkin pathway, thereby preserving mitochondrial biogenesis. In addition, our data also showed that Mfn2 deletion increased ROS production and exacerbated cytotoxicity induced by zinc overload. Our results therefore suggest that Zn2+-induced ROS generation causes mitochondrial autophagy and mitochondrial dysfunction, damaging H9c2 cardiomyocytes. Additionally, Mfn2 may play a key role in zinc ion-mediated endoplasmic reticulum and mitochondrial interactions. Our results provide a new perspective on zinc-induced toxicology.

HDGF stimulates liver tumorigenesis by enhancing reactive oxygen species generation in mitochondria

Hepatoma-derived growth factor (HDGF) overexpression and uncontrolled reactive oxygen species (ROS) accumulation are involved in malignant transformation and poor prognosis in various types of cancer. However, the interplay between HDGF and ROS generation has not been elucidated in hepatocellular carcinoma. Here, we first analyzed the profile of HDGF expression and ROS production in newly generated orthotopic hepatomas by ultrasound-guided implantation. In situ superoxide detection showed that HDGF-overexpressing hepatomas had significantly elevated ROS levels compared with adjacent nontumor tissues. Consistently, liver tissues from HDGF-deficient mice exhibited lower ROS fluorescence than those from age- and sex-matched WT mice. ROS-detecting fluorescent dyes and flow cytometry revealed that recombinant HDGF (rHDGF) stimulated the production of superoxide anion, hydrogen peroxide, and mitochondrial ROS generation in cultured hepatoma cells in a dose-dependent manner. In contrast, the inactive Ser103Ala rHDGF mutant failed to promote ROS generation or oncogenic behaviors. Seahorse metabolic flux assays revealed that rHDGF dose dependently upregulated bioenergetics through enhanced basal and total oxygen consumption rate, extracellular acidification rate, and oxidative phosphorylation in hepatoma cells. Moreover, antioxidants of N-acetyl cysteine and MitoQ treatment significantly inhibited HDGF-mediated cell proliferation and invasive capacity. Genetic silencing of superoxide dismutase 2 augmented the HDGF-induced ROS generation and oncogenic behaviors of hepatoma cells. Finally, genetic knockdown nucleolin (NCL) and antibody neutralization of surface NCL, the HDGF receptor, abolished the HDGF-induced increase in ROS and mitochondrial energetics. In conclusion, this study has demonstrated for the first time that the HDGF/NCL signaling axis induces ROS generation by elevating ROS generation in mitochondria, thereby stimulating liver carcinogenesis.

Caffeine reduces viability, induces apoptosis, inhibits migration and modulates the CD39/CD73 axis in metastatic cutaneous melanoma cells.

We aimed to evaluate the effect of caffeine on viability, apoptosis, migration, redox profile and modulatory effect of the purinergic system of cutaneous melanoma cells. The melanoma cells SK-MEL-28 and non-tumoural CCD-1059sk cells were treated for 24 h with different concentrations of caffeine. Cell viability was evaluated by a biochemical assay and fluorescence microscopy, and flow cytometry assessed apoptosis induction. A wound-healing assay assessed cell migration. The redox profile was evaluated by the levels of markers of reactive oxygen species (ROS), nitric oxide (NOx), total thiols (PSH) and non-protein thiols (NPSH). RT-qPCR and flow cytometry assessed the expression of CD39 and CD73. ATPase/ADPase and AMPase enzyme activities were evaluated by hydrolysis of ATP, ADP and AMP nucleotides. A bioluminescent assay assessed extracellular ATP levels. Caffeine significantly reduced melanoma cell viability and migration and did not affect non-tumoural cells. Caffeine increased ROS levels and improved PSH levels in melanoma cells. Furthermore, caffeine reduced CD39 and CD73 expression, decreased ATP, ADP and AMP nucleotide hydrolysis and increased extracellular ATP levels. We have shown that caffeine reduces metastatic cutaneous melanoma cell viability and migration, induces ROS generation and improves PSH levels. In an unprecedented manner, we also showed that caffeine reduces the expression of CD39 and CD73 and, consequently, ATPase/ADPase/AMPase hydrolytic activity of ectonucleotidases, thus displacing the CD39/CD73 axis and increasing extracellular ATP levels. Therefore, caffeine may be an interesting compound for clinical trials with the CD39/CD73 axis as a therapeutic target.

Toxoplasma gondii Induces Pyroptosis in Human Placental Trophoblast and Amniotic Cells by Inducing ROS Production and Activation of Cathepsin B and NLRP1/NLRP3/NLRC4/AIM2 Inflammasome

Toxoplasma gondii infection in pregnant women may cause fetal anomalies; however, the underlying mechanisms remain unclear. The current study investigated whether T. gondii induces pyroptosis in human placental cells and the underlying mechanisms. Human placental trophoblast (BeWo and HTR-8/SVneo) and amniotic (WISH) cells were infected with T. gondii, and then reactive oxygen species (ROS) production, cathepsin B (CatB) release, inflammasome activation, and pyroptosis induction were evaluated. The molecular mechanisms of these effects were investigated by treating the cells with ROS scavengers, a CatB inhibitor, or inflammasome-specific siRNA. T. gondii infection induced ROS generation and CatB release into the cytosol in placental cells but decreased mitochondrial membrane potential. T. gondii-infected human placental cells and villi exhibited NLRP1, NLRP3, NLRC4, and AIM2 inflammasome activation and subsequent pyroptosis induction, as evidenced by increased expression of ASC, cleaved caspase-1, and mature IL-1β and gasdermin D cleavage. In addition to inflammasome activation and pyroptosis induction, adverse pregnancy outcome was shown in a T. gondii-infected pregnant mouse model. Administration of ROS scavengers, CatB inhibitor, or inflammasome-specific siRNA into T. gondii-infected cells reversed these effects. Collectively, these findings show that T. gondii induces NLRP1/NLRP3/NLRC4/AIM2 inflammasome-dependent caspase-1-mediated pyroptosis via induction of ROS production and CatB activation in placental cells. This mechanism may play an important role in inducing cell injury in congenital toxoplasmosis.