Accumulation Of Reactive Oxidative Species Research Articles

Dolutegravir-induced growth and lifespan effects in Caenorhabditis elegans

BackgroundIntegrase strand transfer inhibitor (INSTIs)-based combination antiretroviral treatment in people living with HIV (PLWH) has been reportedly correlated with several adverse effects, such as weight gain, fetal defects or psychiatric disorders.MethodsTo comprehensively understand the adverse effect of INSTIs, our study utilized Caenorhabditis Elegans (C. elegans) as a model to investigate how dolutegravir (DTG) affected its life cycle, growth, reproduction and lifespan.ResultsOur results indicated that DTG enhanced body growth at the early stage of treatment, but no change was detected for long-term treatment. The treatment also influenced the reproductive system, decreased egg-hatching but had no effect on egg-laying. Besides, DTG resulted in lifespan reduction, which is dependent on increased levels of reactive oxidative species (ROS) accumulation. Treatment with N-acetyl-cysteine (NAC) in worms restrained intracellular ROS accumulation and improved DTG-induced lifespan reduction.ConclusionsOur study demonstrates for the first time the effect of DTG treatment on life cycle. DTG-induced adverse effects are potentially associated with intracellular ROS accumulation. Quenching ROS accumulation might provide a novel strategy for dealing with the adverse effects of INSTIs.

Abstract A020: Neutralizing oxidative damage at telomeres prevents T cell dysfunction and improves adoptive cell therapy

Abstract Background: Failure of adoptive cell therapies (ACTs) is associated with insufficient persistence within the patient, inability to infiltrate tumor sites, and cell-intrinsic loss of functionality. One of the key factors responsible for T cell dysfunction are metabolic barriers such as nutrient competition, low oxygen tension and damaging byproducts in the tumor microenvironment. This leads to a T cell metabolic deregulation affecting their ability to find and kill cancer cell targets properly and effectively. Therefore, elucidating the metabolic pressures experienced in the TME could provide new therapeutic targets to improve ACTs. Reactive Oxidative Species (ROS) accumulation in the TME have detrimental effects on T cell function and anti-tumor response, although the precise targets of ROS are unclear. There are accumulating data showing that mitochondrial ROS can have profound effects on the telomere status of cells. However, there is little evidence describing the role or oxidative stress on telomere health, or the importance of telomere function in immune cells. Our current study demonstrates that tumor infiltrating lymphocytes (TIL) accumulate DNA damage at telomeres, most prominently in terminally exhausted T cells. Furthermore, inducing ROS accumulation at telomeres alone drives T cell dysfunction. Importantly we discovered that alleviating ROS specifically at telomeres improves the response to adoptive cell therapies in a mouse tumor model. Methods: In this study we perform telomeric and centromeric FISH assays to analyze TIL for DNA damage accumulation. We used a chemo-optogenetic FAPS-TAPS to generate singlet oxygen and consequent 8-oxo-guanine lesions specifically at telomeres.1 We tethered the antioxidant protein GPX1 to TRF1, subunit of the Shelterin Complex at telomeres, to generate a telomere-guided ROS scavenger. Results: Telo-FISH analysis demonstrates an accumulation of telomeric DNA damage in TIL from B16 mouse tumors shown by the presence of 53BP1 and ƔH2AX at telomeres. Our data show that mitochondria and telomeric ROS cause the accumulation of DNA damage at telomeres, as well as the development of telomere fragility. These cells ultimately become dysfunctional showing a diminished capability for cytokine production. Importantly, localizing the ROS scavenger GPX1 directly to telomeres reduced telomere fragility and improved the function of therapeutic T cells in the B16 melanoma. Conclusions: Our data suggest that dysfunctional T cells in cancer are not classically senescent, bearing short telomeres, but rather harbor damaged telomeres due to exposure to oxidative stress. Telomeric damage is sufficient to drive a dysfunctional state in newly activated T cells. Protecting telomeres through expression of a telomere-targeted antioxidant protein may preserve T cell function in the tumor microenvironment and drive superior responses to adoptive cell therapies. Citation Format: Dayana Rivadeneira, Sanjana Thosar, Victoria Dean, William Gunn, Konstantinos Lontos, Marcel Bruchez, Patricia Opresko, Greg Delgoffe. Neutralizing oxidative damage at telomeres prevents T cell dysfunction and improves adoptive cell therapy [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor Immunology and Immunotherapy; 2023 Oct 1-4; Toronto, Ontario, Canada. Philadelphia (PA): AACR; Cancer Immunol Res 2023;11(12 Suppl):Abstract nr A020.

Shiga toxin 2 A-subunit induces mitochondrial damage, mitophagy and apoptosis via the interaction of Tom20 in Caco-2 cells

Shiga toxin type 2 (Stx2) is the primary virulence factor produced by Shiga toxin-producing enterohemorrhagic Escherichia coli (STEC), which causes epidemic outbreaks of gastrointestinal sickness and potentially fatal sequela hemolytic uremic syndrome (HUS). Most studies on Stx2-induced apoptosis have been performed with holotoxins, but the mechanism of how the A and B subunits of Stx2 cause apoptosis in cells is not clear. Here, we found that Stx2 A-subunit (Stx2A) induced mitochondrial damage, PINK1/Parkin-dependent mitophagy and apoptosis in Caco-2 cells. PINK1/Parkin-dependent mitophagy caused by Stx2A reduced apoptosis by decreasing the accumulation of reactive oxidative species (ROS). Mechanistically, Stx2A interacts with Tom20 on mitochondria to initiate the translocation of Bax to mitochondria, leading to mitochondrial damage and apoptosis. Overall, these data suggested that Stx2A induces mitochondrial damage, mitophagy and apoptosis via the interaction of Tom20 in Caco-2 cells and that mitophagy caused by Stx2A ameliorates apoptosis by eliminating damaged mitochondria. These findings provide evidence for the potential use of Tom20 inhibition as an anti-Shiga toxin therapy.

Unraveling the Mechanisms of Virus-Induced Symptom Development in Plants.

Plant viruses, as obligate intracellular parasites, induce significant changes in the cellular physiology of host cells to facilitate their multiplication. These alterations often lead to the development of symptoms that interfere with normal growth and development, causing USD 60 billion worth of losses per year, worldwide, in both agricultural and horticultural crops. However, existing literature often lacks a clear and concise presentation of the key information regarding the mechanisms underlying plant virus-induced symptoms. To address this, we conducted a comprehensive review to highlight the crucial interactions between plant viruses and host factors, discussing key genes that increase viral virulence and their roles in influencing cellular processes such as dysfunction of chloroplast proteins, hormone manipulation, reactive oxidative species accumulation, and cell cycle control, which are critical for symptom development. Moreover, we explore the alterations in host metabolism and gene expression that are associated with virus-induced symptoms. In addition, the influence of environmental factors on virus-induced symptom development is discussed. By integrating these various aspects, this review provides valuable insights into the complex mechanisms underlying virus-induced symptoms in plants, and emphasizes the urgency of addressing viral diseases to ensure sustainable agriculture and food production.

Genome-wide characterization of ascorbate peroxidase gene family in pepper (Capsicum annuum L.) in response to multiple abiotic stresses.

Pepper is widely grown all over the world, so it faces many abiotic stresses, such as drought, high temperature, low temperature, salt damage, and so on. Stresses causing the accumulation of reactive oxidative species (ROS) in plants are removed by antioxidant defense systems, and ascorbate peroxidase (APX) is an important antioxidant enzyme. Therefore, the present study performed genome-wide identification of the APX gene family in pepper. We identified nine members of the APX gene family in the pepper genome according to the APX proteins' conserved domain in Arabidopsis thaliana. The physicochemical property analysis showed that CaAPX3 had the longest protein sequence and the largest molecular weight of all genes, while CaAPX9 had the shortest protein sequence and the smallest MW. The gene structure analysis showed that CaAPXs were composed of seven to 10 introns. The CaAPX genes were divided into four groups. The APX genes of groups I and IV were localized in the peroxisomes and chloroplasts, respectively; the group II genes were localized in the chloroplasts and mitochondria; and the group III genes were located in the cytoplasm and extracell. The conservative motif analysis showed that all APX genes in the pepper had motif 2, motif 3, and motif 5. The APX gene family members were distributed on five chromosomes (Chr. 2, 4, 6, 8, and 9). The cis-acting element analysis showed that most CaAPX genes contain a variety of cis-elements related to plant hormones and abiotic stress. RNA-seq expression analysis showed that the expression patterns of nine APXs were different in vegetative and reproductive organs at different growth and development stages. In addition, the qRT-PCR analysis of the CaAPX genes revealed significant differential expression in response to high temperature, low temperature, and salinity stresses in leaf tissue. In conclusion, our study identified the APX gene family members in the pepper and predicted the functions of this gene family, which would provide resources for further functional characterization of CaAPX genes.

Influence of Redox and Dopamine Regulation in Cocaine-Induced Phenotypes Using Drosophila.

Reactive Oxidative Species (ROS) are produced during cellular metabolism and their amount is finely regulated because of negative consequences that ROS accumulation has on cellular functioning and survival. However, ROS play an important role in maintaining a healthy brain by participating in cellular signaling and regulating neuronal plasticity, which led to a shift in our understanding of ROS from being solely detrimental to having a more complex role in the brain. Here we use Drosophila melanogaster to investigate the influence of ROS on behavioral phenotypes induced by single or double exposure to volatilized cocaine (vCOC), sensitivity and locomotor sensitization (LS). Sensitivity and LS depend on glutathione antioxidant defense. Catalase activity and hydrogen peroxide (H2O2) accumulation play a minor role, but their presence is necessary in dopaminergic and serotonergic neurons for LS. Feeding flies the antioxidant quercetin completely abolishes LS confirming the permissive role of H2O2 in the development of LS. This can only partially be rescued by co-feeding H2O2 or the dopamine precursor 3,4-dihydroxy-L-phenylalanine (L-DA) showing coordinate and similar contribution of dopamine and H2O2. Genetic versatility of Drosophila can be used as a tool for more precise dissection of temporal, spatial and transcriptional events that regulate behaviors induced by vCOC.

Comparison of oxidative stress response of in vitro retinal cells exposed to blue light from emissive versus reflective displays

AbstractDuring COVID‐19, there was increased use of handheld displays in educational settings. There is growing concern that eye health may be affected by prolonged exposure to the light‐emitting diodes used as frontlights or backlights in handheld displays. The potential impact of light exposure from tablet‐sized devices with different display technologies and various spectral outputs was assessed in an in vitro model using human retinal epithelial (ARPE‐19) cells. Cellular response was quantified by measuring reactive oxidative species (ROS) and by analyzing mitochondrial morphology. Control experiments established a baseline ROS response to hazardous blue light exposure and also that red light resulted in no detectable ROS response. Under identical conditions, ROS response increased with time for all devices. However, different device spectra caused ROS to accumulate at different rates. When operating the devices in the same mode (day or night), cells accumulated ROS two to three times more slowly on exposure to frontlit electronic paper displays compared to backlit liquid crystal displays. With increasing ROS accumulation, mitochondrial morphology shifted from elongate interconnected features typically observed under normal conditions to rounded disconnected features associated with oxidative stress response.

Cellular mechanism of action of forsythiaside for the treatment of diabetic kidney disease.

Background: Diabetic kidney disease (DKD) becomes the leading cause of death for end-stage renal disease, whereas the potential mechanism is unclear and effective therapy is still rare. Our study was designed to investigate the cellular mechanism of Forsythiaside against DKD. Materials and Methods: The targets of Forsythiaside and the DKD-related targets were obtained from databases. The overlapping targets in these two sets were regarded as potential targets for alleviation of DKD by Forsythiaside. The targets of diabetic podocytopathy and tubulopathy were also detected to clarify the mechanism of Forsythiaside ameliorating DKD from the cellular level. Results: Our results explored that PRKCA and RHOA were regarded as key therapeutic targets of Forsythiaside with excellent binding affinity for treating DKD podocytopathy. Enrichment analysis suggested the underlying mechanism was mainly focused on the oxidative stress and mTOR signaling pathway. The alleviated effects of Forsythiaside on the reactive oxidative species accumulation and PRKCA and RHOA proteins upregulation in podocytes were also confirmed. Conclusion: The present study elucidates that Forsythiaside exerts potential treatment against DKD which may act directly RHOA and PRKCA target by suppressing the oxidative stress pathway in podocytes. And Forsythiaside could be regarded as one of the candidate drugs dealing with DKD in future experimental or clinical researches.

CD36 promotes tubular ferroptosis by regulating the ubiquitination of FSP1 in acute kidney injury

Reactive oxidative species (ROS) production-driven ferroptosis plays a role in acute kidney injury (AKI). However, its exact molecular mechanism is poorly understood. Scavenger receptor CD36 has important roles in oxidizing lipids, lipid accumulation, metabolic syndrome, and insulin resistance in chronic kidney disease, but its roles remain unexplored in AKI. The present study investigated the role and mechanism of CD36 in regulating proximal tubular cell ferroptosis and AKI. The expression of CD36 was found to be significantly up-regulated in AKI renal tissues and correlated with renal function, which might serve as an independent biomarker for AKI patients. Moreover, in adult mice subjected to AKI, deletion of CD36 (CD36−/−) induced tubular cell ROS accumulation, ferroptosis activation, and renal injury. Mechanistically, combining LC-MS/MS, co-IP, and ubiquitination analyses revealed that CD36 could specifically bind to ferroptosis suppressor protein 1 (FSP1) and regulate its ubiquitination at sites K16 and K24, leading to FSP1 degradation and progression of ferroptosis in AKI. The present results emphasize a novel mechanism of CD36 in cisplatin-induced AKI. The discovery of the special CD36 roles in promoting ferroptosis and AKI development by regulating the ubiquitination of FSP1 in proximal tubular cells may be potential therapeutic targets for AKI. Moreover, CD36 may play a key role in the progression of AKI. Therefore, targeting CD36 may provide a promising treatment option for AKI.

Quercetin alleviates nanoparticle-induced osteolysis via deactivating pyroptosis.

As one of the most successful stories in modern medicine, total joint arthroplasty (TJA) is performed several million times worldwide every year. However, more than 20% of patients will suffer from aseptic loosening (AL) following periprosthetic osteolysis (PPO) in the next few years. Unfortunately, the only effective treatment for PPO, i.e., revision surgery, can cause great surgical trauma. It has been reported that the accumulation of reactive oxidative species (ROS) generated by exposure to wear particles could activate NLRP3 inflammasome in macrophages and accelerate the progress of osteolysis. Given that the conservative treatment is not effective and can be accompanied by apparent side effects, we, therefore, investigated the therapeutic effect of the natural compound quercetin (Que) on wear particle-induced osteolysis. Our results showed that Que could activate nuclear factor erythroid 2-related factor 2 (Nrf2) to remove ROS and deactivate inflammasome activation. Besides, the imbalance between osteoclastogenesis and osteogenesis induced by inflammatory cytokines was also rescued by Que. Collectively, our work demonstrates that Que may be a qualified candidate for the conservative treatment of wear particle-induced osteolysis.

Mucosal-associated invariant T cells restrict reactive oxidative damage and preserve meningeal barrier integrity and cognitive function.

Increasing evidence indicates close interaction between immune cells and the brain, revising the traditional view of the immune privilege of the brain. However, the specific mechanisms by which immune cells promote normal neural function are not entirely understood. Mucosal-associated invariant T cells (MAIT cells) are a unique type of innate-like T cell with molecular and functional properties that remain to be better characterized. In the present study, we report that MAIT cells are present in the meninges and express high levels of antioxidant molecules. MAIT cell deficiency in mice results in the accumulation of reactive oxidative species in the meninges, leading to reduced expression of junctional protein and meningeal barrier leakage. The presence of MAIT cells restricts neuroinflammation in the brain and preserves learning and memory. Together, our work reveals a new functional role for MAIT cells in the meninges and suggests that meningeal immune cells can help maintain normal neural function by preserving meningeal barrier homeostasis and integrity.

THERMODYNAMIC STUDY OF CERIUM OXIDE NANOPARTICLES AND THEIR EFFECTS ON CELLULAR METABOLISM

Oxidative stress is associated with a large range of health conditions. It is caused by the accumulation of reactive oxidative species above cellular neutralization capability. Cells generally defend against oxidative stress with ROS decomposing enzymes such as superoxide dismutase and catalase. Cerium oxide nanoparticles display activity that mimics superoxide dismutase and catalase antioxidative properties, allowing them to combat oxidative stress. Oxidative stress can impair the mitochondrial function and energy output of cells, which can be measured as heat with closed ampoule isothermal microcalorimetry. Thermodynamic analysis of the cell’s response to nanoceria treatment can help improve understanding of its general medical applications. Murine macrophages of the RAW264.7 cell line were cultured in 10% FBS supplemented DMEM media with 1% AA until it reached 90-95% confluency monitored by the EVOS M5000 microscope. After establishing a baseline heat output for healthy cells using a TAM-III isothermal microcalorimeter, the heat flow was measured in cells under induced oxidative stress. MTT assay was performed with various concentrations of nanoceria to identify the optimal dosage at which its antioxidative properties are most effective. In this study, we described the effect of nanoceria on mitochondrial activity.

Reactive oxygen species accumulation is synchronised with growth inhibition of temperature-sensitive recAts polA Escherichia coli

When combined with recombinase defects, chromosome breakage and double-strand break repair deficiencies render cells inviable. However, cells are viable when an SOS response occurs in recAts polA cells in Escherichia coli. Here, we aimed to elucidate the underlying mechanisms of this process. Transposon mutagenesis revealed that the hslO gene, a redox chaperone Hsp33 involved in reactive oxidative species (ROS) metabolism, was required for the suppression of recAts polA lethality at a restricted temperature. Recently, it has been reported that lethal treatments trigger ROS accumulation. We also found that recAts polA cells accumulated ROS at the restricted temperature. A catalase addition to the medium alleviates the temperature sensitivity of recAts polA cells and decreases ROS accumulation. These results suggest that the SOS response and hslO manage oxidative insult to an acceptable level in cells with oxidative damage and rescue cell growth. Overall, ROS might regulate several cellular processes.

Characterization of 24-epibrassinolide-mediated modulation of the drought stress responses: Morphophysiology, antioxidant metabolism and hormones in grapevine (Vitis vinifera L.)

Drought stress is one of the major abiotic stresses that limit grape growth and yield. Brassinosteroids (BRs) are a class of phytohormones essential for plant growth, development, and adaptation to environmental stress. This study aimed to reveal the physiological and biochemical mechanisms of exogenous BRs in alleviating the drought stress in grapevines. Two-year-old grape seedlings (Vitis vinifera L.) were sprayed with 24-epibrassinolide (EBR), a synthetic analog of BRs, and then subjected to drought treatment. The results showed that exogenous EBR significantly mitigated the reduction of photosynthetic pigment contents and photosystem II efficiency and decreased the damage to chloroplasts when grape seedlings were subjected to drought stress. Drought stress resulted in the accumulation of reactive oxidative species (ROS) and an increase in lipid peroxidation. A reduction in oxidative damage was observed in EBR-pretreated plants, which was probably due to the elevated antioxidant system. Exogenous EBR improved the activities of superoxide dismutase (14%), catalase (18%), peroxidase (17%), and ascorbate peroxidase (9%), and promoted the accumulation of ascorbic acid (10%) and glutathione (7%) under drought stress. EBR pretreatment also promoted autophagic activity, which contributed to the degradation of damaged chloroplasts. Moreover, EBR pretreatment increased the concentrations of abscisic acid, jasmonic acid, auxin, and gibberellic acid. Taken together, exogenous EBR could ameliorate the deleterious effects of drought stress by up-regulating photosynthetic capacity, antioxidant system, autophagic activity, and hormone concentrations.

Characterization of SOD and GPX Gene Families in the Soybeans in Response to Drought and Salinity Stresses.

Plant stresses causing accumulation of reactive oxidative species (ROS) are scavenged by effective antioxidant defense systems. Therefore, the present study performed genome-wide identification of superoxide dismutase (SOD) and glutathione peroxidase (GPX) gene families in cultivated and wild soybeans, and 11 other legume species. We identified a total of 101 and 95 genes of SOD and GPX, respectively, across thirteen legume species. The highest numbers of SODs and GPXs were identified in cultivated (Glycine max) and wild (Glycine soja). A comparative phylogenetic study revealed highest homology among the SODs and GPXs of cultivated and wild soybeans relative to other legumes. The exon/intron structure, motif and synteny blocks were conserved in both soybean species. According to Ka/Ks, purifying the selection played the major evolutionary role in these gene families, and segmental duplication are major driving force for SODs and GPXs expansion. In addition, the qRT-PCR analysis of the G. max and G. soja SOD and GPX genes revealed significant differential expression of these genes in response to oxidative, drought and salinity stresses in root tissue. In conclusion, our study provides new insights for the evolution of SOD and GPX gene families in legumes, and provides resources for further functional characterization of these genes for multiple stresses.

Decreased Expression and Uncoupling of Endothelial Nitric Oxide Synthase in the Cerebral Cortex of Rats with Thioacetamide-Induced Acute Liver Failure.

Acute liver failure (ALF) is associated with deregulated nitric oxide (NO) signaling in the brain, which is one of the key molecular abnormalities leading to the neuropsychiatric disorder called hepatic encephalopathy (HE). This study focuses on the effect of ALF on the relatively unexplored endothelial NOS isoform (eNOS). The cerebral prefrontal cortices of rats with thioacetamide (TAA)-induced ALF showed decreased eNOS expression, which resulted in an overall reduction of NOS activity. ALF also decreased the content of the NOS cofactor, tetrahydro-L-biopterin (BH4), and evoked eNOS uncoupling (reduction of the eNOS dimer/monomer ratio). The addition of the NO precursor L-arginine in the absence of BH4 potentiated ROS accumulation, whereas nonspecific NOS inhibitor L-NAME or EDTA attenuated ROS increase. The ALF-induced decrease of eNOS content and its uncoupling concurred with, and was likely causally related to, both increased brain content of reactive oxidative species (ROS) and decreased cerebral cortical blood flow (CBF) in the same model.

The Role of Ceramide Synthases Response to the Heavy Metal Cadmium Chloride

Cadmium is an extremely toxic heavy metal that is hazardous to human health. In the United States, the primary source of cadmium exposure is in the food supply. Ingestion results in a buildup of cadmium in the body, specifically in the kidneys, which can lead to organ damage and cancer. This toxin has been known to induce stress in organisms by causing an accumulation of reactive oxidative species (ROS). To counter stress, cells respond with a variety of signaling pathways. We are interested in studying the sphingolipid pathway, which is known to mediate stress responses, but its role in defending against cadmium toxicity is unclear. We hypothesized that ceramide synthase, a sphingolipid metabolism enzyme, is required for survival to cadmium stress and decreased accumulation of ROS in animals. To test this, we utilized the model organism Caenorhabditis elegans. Ceramide synthase converts sphingosine to ceramide in the cells. We examined animals with mutations in ceramide synthase (hyl‐2 mutants) by treating worms to acute and chronic cadmium exposure and examining changes in survival, reproduction, and locomotion. We present data showing that cadmium does not affect animal reproduction, but it does affect survival upon exposure. Cadmium exposure resulted in decreased survival, however, and this result was less prominent in organisms possessing hyl‐2/ceramide synthase. We also found that the hyl‐2 gene is vital for oxidative stress response mechanisms. In addition, we present data on the effects of cadmium exposure on gene expression of sphingolipid metabolism enzymes using qPCR. These results are important because stress plays a role in aging, and if we better understand how stress is regulated via sphingolipids, it could become a potential pharmaceutical target to slow the degradative aging process.Support or Funding InformationNIH‐ 2R15AG052933

Role of mitochondrial damage in cadmium-induced cell apoptosis and DNA damage of hepatocytes

To investigate the role of mitochondrial damage mediated by reactive oxidative species(ROS) in cadmium-induced cell apoptosis and DNA damage of L02 hepatocytes, so as to provide experimental basis for the subsequent study and protection of people exposed to Cd. The L02 hepatocytes were cultured in vitro treated with 0-90 μmol/L Cd for 24 h, and the methylthiazolyldiphenyltetrazoliumbromide assay was used to detect the cell viability. The colony formation assay, flow cytometry, comet assay, 2', 7'-dichlorofluorescein diacetate, MitoTracker Red CMXRos and 10-N-nonyl-acridine-orange, mitochondrial membrane potential detection kit(JC-1) and adenosine triphosphate(ATP) assay kits and Western Blot were used to investigate cell growth and proliferation, cell apoptosis, DNA damage, ROS levels, mitochondrial morphology, mitochondrial membrane potential, mitochondrial mass, ATP content and related proteins after the cells exposed to 0, 20, 40 μmol/L Cd for 24 h. The cells were pretreated with vitamin C before adding Cd exposure, and ROS levels, mitochondrial function, cell apoptosis, DNA damage and proteins were measured. The cell viability was significantly inhibited with the increase of Cd concentration and treatment time. The cells were treated with Cd for 24 h for further study according to the result of MTT assay. Compared with control group, the colony formation rate were 8. 23% and 6. 17% respectively in 20 and 40 μmol/L Cd treatment and the apoptosis rate were 15. 85% and 26. 26%, respectively. We also found that the B cell lymphoma/leukemia(Bcl-2) gene protein was significantly reduced, while the levels of Bcl-2 associated X protein(Bax) and cleaved cysteine aspastic acid-specific protease 3(cleaved-caspase-3) were increased in a dose-dependent manner. Cd treatment also induced DNA damage and accumulation of intracellular ROS, accompanied by a mitochondrial morphological change, significant decrease in Δψm, mitochondrial mass, ATP content, mitochondrial cytochrome C(cyt c) and an increase in cytoplasmic cyt c expression(P<0. 05). In addition, pretreatment with antioxidant vitamin C not only significantly increased cyt c, mitochondrial mass, ATP content and mitochondrial cyt c, but also reduced the expression of cytoplasmic cyt c(P<0. 05), cell apoptosis and DNA damage induced by Cd. Cd exposure could induce ROS accumulation in L02 hepatocytes, which can lead to mitochondrial damage, and ultimately lead to cell apoptosis and DNA damage.

α-Mangostin reduced the viability of A594 cells in vitro by provoking ROS production through downregulation of NAMPT/NAD.

α-Mangostin (MAN) is a bioactive compound isolated from the inedible pericarp of a tropical fruit mangosteen (Garcinia mangostana Linn). It exhibits notable therapeutic potentials on lung cancers, but the underlying mechanisms are still largely unknown. This study was designed to further explore the mechanisms involved in cytotoxicity of MAN on A549 cells. Apoptosis and cell cycle distribution were analyzed by flow cytometry methods. The fluorescent probes DCFH-DA and JC-1 were used to assess the intracellular reactive oxidative species (ROS) and mitochondrial membrane potential statuses, respectively. The regulation of MAN on relevant pathways was investigated by immunoblotting assays. The results obtained indicated that MAN caused significant apoptosis and cell cycle arrest in A549 cells, which eventually resulted in inhibition on cell proliferation in vitro. All these phenomena were synchronized with escalated oxidative stress and downregulation of nicotinamide phosphoribosyltransferase/nicotinamide adenine dinucleotide (NAMPT/NAD). Supplementation with nicotinamide mononucleotide (NMN) and N-acetylcysteine (NAC) efficiently eased MAN-induced ROS accumulation, and potently antagonized MAN-elicited apoptosis and cell cycle arrest. The pro-apoptotic effect of MAN was further confirmed by increased expressions of cleaved caspase 3, 6, 7, and 9, and its effect on cell cycle progression was validated by the altered expressions of p-p38, p-p53, CDK4, and cyclin D1. The immunoblotting assays also demonstrated that NAC/NMN effectively restored these molecular changes elicited by MAN treatment. Collectively, this study revealed a unique anti-tumor mechanism of MAN by provoking ROS production through downregulation of NAMPT/NAD signaling and further validated MAN as a potential therapeutic reagent for lung cancer treatment.

Antitumor effects of seleno-short-chain chitosan (SSCC) against human gastric cancer BGC-823 cells.

Seleno-short-chain chitosan (SSCC) is a derivative of chitosan. In the present study, we sought to investigate the underlying antitumor mechanism of SSCC on human gastric cancer BGC-823 cells in vitro. MTT assay suggested that SSCC exhibited a dose-dependent inhibitory effect on the proliferation of BGC-823 cells. We found the SSCC-treated cells showed typical morphological characteristics of apoptosis in a dose dependent manner by observing on microscope. Annexin V-FITC/PI double staining and cell cycle assay identified that SSCC could induce BGC-823 cells apoptosis by triggering G2/M phase arrest. Our research provided the first evidence that SSCC could effectively induce the apoptosis of BGC-823 cells via an intrinsic mitochondrial pathway, as indicated by inducing the disruption of mitochondrial membrane potential (MMP), the excessive accumulation of reactive oxidative species (ROS), the increase of Bax/Bcl-2 ratio and the activation of caspase 3, caspase 9 and cytochrome C (Cyt-C) in BGC-823 cells. These combined results clearly indicated that SSCC could induce BGC-823 cells apoptosis by the involvement of mitochondrial signaling pathway, which provided precise experimental evidence for SSCC as a potential agent in the prevention and treatment of human gastric cancer.