Increased Reactive Oxygen Species Levels Research Articles

Gaseous ozone regulates reactive oxygen species metabolism and ascorbate–glutathione cycle to delay the senescence of fresh-cut red pitaya (Selenicereus undatus) fruit

Mechanical damage during processing can accelerate the senescence and deterioration of fresh-cut red pitaya (Selenicereus undatus), thereby affecting its nutritional quality and commodity value. In this study, the effects of a low-concentration and long-term (10 μL L−1 for 60 min) gaseous ozone treatment on the quality, reactive oxygen species (ROS) metabolism, and enzyme and non-enzyme antioxidant activities of fresh-cut red pitaya fruit were investigated. The results showed that ozone treatment delayed the decline in fruit firmness, total soluble solids, and titratable acid content. Ozone treatment also maintained a higher free radical scavenging capacity and lower membrane lipid peroxidation than the control treatment. It prevented the rapid increase in ROS level by improving the activity of antioxidant enzymes, including superoxide dismutase, peroxidase, and catalase. Additionally, the treatment activated the ascorbate–glutathione cycle. Compared with the control group, ozone treatment maintained the activities of ascorbic acid peroxidase, dehydroascorbic acid reductase, monodehydroascorbic acid reductase, and glutathione reductase. Although the contents of ascorbic acid (AsA) and dehydroascorbic acid (DHA) decreased within 10 days of storage, the AsA/DHA and glutathione/oxidized glutathione ratios in the ozone-treated group were higher than those in the control group from day 0 to day 6. Thus, ozone treatment regulated ROS metabolism, enhanced the ROS-scavenging system, maintained the redox balance of fresh-cut red pitaya, and delayed senescence.

Nitrogen limitation and hydrogen peroxide act synergistically to enhance lipids accumulation via ROS/Ca2+ dependent mechanism in Chlorella sorokiniana

The main aim of the present study is to assess the effects of combined nitrogen limitation and hydrogen peroxide (H2O2) treatment on lipid accumulation in Chlorella sorokiniana. Furthermore, to study the regulative networks involved in the lipid accumulation of microalgae, the relationships between reactive oxygen species (ROS), Ca2+ and four lipogenic genes (mitogen-activated protein kinase: MAPK; heteromeric acetyl-CoA carboxylase beta subunit: accD; b-ketoacyl-ACP synthase III: KASIII; diacylglycerol acyltransferase: DGAT) were also analyzed. The results indicated that the contents of chlorophyll a, carbohydrate and protein were decreased, while the lipid content and levels of endogenous ROS, Ca2+ were increased under nitrogen limitation groups with H2O2 added. Further research indicated that the increased ROS level could affect lipid accumulation through lipogenic genes expression. In addition, the interplays of ROS and Ca2+ may be found. This study provides an effective strategy to improve the lipid accumulation in C. sorokiniana, and offers evidence of ROS, Ca2+ and lipogenic genes in the accommodation of lipid biosynthesis in microalgae under combined treatment of nitrogen limitation and H2O2.

Antioxidant Test, Total Alkaloid Content, Antimitosis Test and HPTLC Test of Chamomile Flower Extract (Matricaria Chamomilla L.)

Oxygen is one of the supports of multicellular life and is necessary for aerobic metabolism. However, the use of oxygen can produce by-products, one of which is reactive oxygen species (ROS) which can also increase due to external factors such as exposure to ionizing rays, and vehicle smoke. When reactive oxygen species levels are high, the body can experience a condition called oxidative stress, which is a condition that can cause tissue damage. To overcome this, antioxidants are needed, namely compounds that can donate electrons to free radicals to neutralize their effects. Antioxidants are divided into endogenous antioxidants and exogenous antioxidants. Endogenous antioxidants are antioxidants found in the body. Exogenous antioxidants are antioxidants that come from outside the body. Because there is an increase in ROS levels due to external influences, an increase in exogenous antioxidants is needed, one of which is chamomile flowers. This study evaluated qualitative phytochemical levels using the Harnborne method, total antioxidant capacity using the Blois method, total quantitative levels of alkaloids using the Trivedi et al method, BSLT toxicity using the Meyer method, and fingerprint analysis using HPTLC. The obtained chamomile flowers are dried and extracted by maceration using methanol solvent. The test results on chamomile flowers revealed phytochemical content in the form of alkaloids, betasianin, cardioglycosides, coumarins, flavonoids, phenols, quinones, saponins, steroids, tepenoids and tannins. Antioxidant ability 209.27 g/ml; Total alkaloid content of 12.62 g/ml; toxicity 174.39 g/mL; As well as fingerprint analysis chamomile flowers have active ingredients. Therefore, chamomile flowers can serve as a moderate antioxidant with antimitotic activity.

Amber Extract Suppressed Mast Cell-Mediated Allergic Inflammation via the Regulation of Allergic Mediators—An In Vitro Study

The various clinical approaches for treating allergy-related diseases have shown modest progress in low side effects and improved clinical outcomes. Therefore, finding alternative anti-allergic agents is crucial. The present study explored the anti-allergic effects of amber extract (fossilized tree resin) in RBL-2H3 mast cells stimulated with different allergens. In order to support the information on the inflammatory effect of the amber extract, NO production analysis on RAW 264.7 cells was conducted. β-Hexosaminidase release, an indicator of the efficacy of the amber extract in preventing mast cell activation and degranulation, reactive oxygen species (ROS) generation, and the effect of the amber extract on key cytokines production on RBL-2H3 cells, was evaluated. The results demonstrated that amber extract at concentrations up to 50 μg/mL had no cytotoxic effect on RAW 264.7 and RBL-2H3 cells. Amber extract inhibited NO production in RAW 264.7 cells. Treatment with amber extract significantly suppressed the release of β-hexosaminidase, especially at 50 μg/mL. Furthermore, amber extract suppressed the significantly increased ROS levels induced by allergen stimulation and allergy-associated cytokines. The results also suggested that amber extract exerts anti-allergic inflammatory effects by inhibiting the MAPK and NF-κB signaling pathways, resulting in decreased cytokines production. Thus, the amber extract is a promising anti-allergic agent.

Cyanidin-3-O-Glucoside Induces the Apoptosis of Human Gastric Cancer MKN-45 Cells through ROS-Mediated Signaling Pathways.

Cyanidin-3-O-glucoside (C3G), an active ingredient in anthocyanins, mainly exists in dark cereals. C3G was investigated for its effect on human gastric cancer (GC) cells, together with its molecular mechanism. The CCK-8 assay results showed that C3G had significant antiproliferative effects on GC cells, but it had little effect on normal cells. Western blot and flow cytometry results showed that C3G regulated the reduction of mitochondrial membrane potential and arrested the cell cycle in the G2/M phase through the AKT signaling pathway, causing the cells to undergo apoptosis. Additionally, in MKN-45 cells, C3G markedly raised intracellular reactive oxygen species (ROS) levels. The wound healing assay and Transwell assay results showed that MKN-45 cell migration was significantly inhibited. Western blot results showed that the expression of E-cadherin protein was upregulated and the expressions of β-catenin, N-cadherin, and Vimentin were downregulated. Additionally, following N-acetylcysteine treatment, the expression levels of these proteins were reduced. In conclusion, C3G caused MKN-45 cells to undergo apoptosis; arrested the cell cycle in the G2/M phase; hindered cell migration; and activated the MAPK, STAT3, and NF-κB signaling pathways, by inducing an increase in ROS levels. Thus, C3G may be a promising new medication for the treatment of GC.

Reduced coenzyme Q synthesis confers non-target site resistance to the herbicide thaxtomin A

Herbicide resistance in weeds is a growing threat to global crop production. Non-target site resistance is problematic because a single resistance allele can confer tolerance to many herbicides (cross resistance), and it is often a polygenic trait so it can be difficult to identify the molecular mechanisms involved. Most characterized molecular mechanisms of non-target site resistance are caused by gain-of-function mutations in genes from a few key gene families-the mechanisms of resistance caused by loss-of-function mutations remain unclear. In this study, we first show that the mechanism of non-target site resistance to the herbicide thaxtomin A conferred by loss-of-function of the gene PAM16 is conserved in Marchantia polymorpha, validating its use as a model species with which to study non-target site resistance. To identify mechanisms of non-target site resistance caused by loss-of-function mutations, we generated 107 UV-B mutagenized M. polymorpha spores and screened for resistance to the herbicide thaxtomin A. We isolated 13 thaxtomin A-resistant mutants and found that 3 mutants carried candidate resistance-conferring SNPs in the MpRTN4IP1L gene. Mprtn4ip1l mutants are defective in coenzyme Q biosynthesis and accumulate higher levels of reactive oxygen species (ROS) than wild-type plants. Mutants are weakly resistant to thaxtomin A and cross resistant to isoxaben, suggesting that loss of MpRTN4IP1L function confers non-target site resistance. Mutants are also defective in thaxtomin A metabolism. We conclude that loss of MpRTN4IP1L function is a novel mechanism of non-target site herbicide resistance and propose that other mutations that increase ROS levels or decrease thaxtomin A metabolism could contribute to thaxtomin A resistance in the field.

Effects of exogenous spermidine on autophagy and antioxidant capacity in ovaries and granulosa cells of Sichuan white geese.

Autophagy can inhibit ovarian senescence induced by oxidative stress and regulate follicle development and atresia, but its mechanism is still unclear. Exogenous spermidine can induce autophagy and scavenge reactive oxygen species (ROS). In this experiment, oxidative stress in Sichuan white geese ovaries and follicular granulosa cells (GCs) was caused by 3-nitropropionic acid (3-NPA) and spermidine was added to explore the effect of exogenous spermidine inducing autophagy and inhibiting oxidative stress in vivo and in vitro. Research results showed that putrescine, spermidine and spermine contents in goose ovaries in the group treated with spermidine combined with 3-NPA were 2.70, 1.94, and 1.70 times higher than those in the group treated with 3-NPA, respectively (P < 0.05). The contents of spermidine and spermine in GCs were 1.37 and 0.89 times higher in the spermidine in combination with the 3-NPA group than in the 3-NPA group, respectively (P < 0.05). LC3 and p62 were mainly expressed in the follicular granulosa layer. The LC3-II/I ratio and p62 level in GCs in the spermidine combined with 3-NPA treatment group were 1.37 and 0.77 times higher than that of the 3-NPA treatment group, respectively (P < 0.05). 3-NPA treatment significantly increased ROS level and the apoptosis rate in GCs, while the combined treatment of spermidine and 3-NPA reversed this change (P < 0.05). In conclusion, spermidine alleviated the oxidative damage induced by 3-NPA by improving the antioxidant capacity of ovaries and follicular GCs of Sichuan white geese and may be alleviated by inducing autophagy in GCs.

Dexamethasone Induces Senescence-Associated Changes in Trabecular Meshwork Cells by Increasing ROS Levels Via the TGFβ/Smad3-NOX4 Axis

Glaucoma is a serious complication of glucocorticoid (GC) therapy arising through elevations in intraocular pressure (IOP). Dexamethasone (DEX) is reported to contribute to elevated IOP through different effects on the trabecular meshwork but whether DEX contributes to glaucoma development through the induction of cellular senescence is still unclear. We explored the actions of DEX on transformed human trabecular meshwork cells (HTMCs) using RNA-seq and conducted bioinformatic analyses to determine the affected pathways. Among the 4,103 differentially expressed genes identified in transformed HTMCs treated with 400 nM DEX (2,036 upregulated and 2,067 downregulated genes, respectively), bioinformatic analyses revealed significant enrichment and potential interplay between the transforming growth factor beta (TGFβ)41; signaling and cellular senescence pathways. DEX treatment induced senescence changes in primary and transformed HTMCs as indicated by increases in SA-β-gal positivity, interleukin (IL)-6 secretion, and senescence-associated heterochromatin foci (SAHF) along with selective accumulation of senescence marker p15 and elevations in reactive oxygen species (ROS) levels. Notably, the DEX-induced senescence changes were rescued by treatment with the TGFβ/Smad3 pathway inhibitor SIS3. Furthermore, we show that DEX increases cellular ROS levels via upregulation of NADPH oxidase 4 (NOX4) through activation of Smad3, and that SIS3 decreases ROS levels by downregulating NOX4. Instructively, inhibiting NOX4 with GLX351322 and scavenging ROS with NAC were both effective in preventing DEX-induced senescence changes. Similarly, we found in the mouse model that DEX-ac upregulated p15 and NOX4 expression in the trabecular meshwork, with cotreatment with GLX351322 alleviating elevations in IOP. We establish that DEX induces senescence changes in HTMCs by increasing ROS levels via the TGFβ/Smad3/NOX4 axis, increasing IOP and contributing to glaucoma development.

FABP4 Regulates Cell Proliferation, Stemness, Apoptosis, and Glycolysis in Colorectal Cancer via Modulating ROS/ERK/mTOR Pathway.

Colorectal cancer is a common digestive tract malignancy. This study aimed to expound the functional role of fatty-acid-binding protein 4 (FABP4) and the potential underlying mechanisms in the development of colorectal cancer. Several techniques were utilized to investigate the role of FABP4 in colorectal cancer. FABP4 mRNA expression was quantified using Real time-quantitative PCR (RT-qPCR). Cell counting kit-8 (CCK-8), 5-ethynyl-2'-deoxyuridine (EdU), sphere formation assays and flow cytometry evaluated cell growth, stemness, and apoptosis in SW480 and HT29 cells. Glycolysis was assessed via extracellular acidification rate (ECAR) , lactate production, glucose uptake, adenosine triphosphate (ATP)/adenosine 5'-diphosphate (ADP) ratio, and Glut1 and Elevated lactate dehydrogenase A (LDHA) protein expression. Reactive oxygen species (ROS) levels were analyzed by flow cytometry. Western blot measured the protein expression of FABP4, Proliferating cell nuclear antigen (PCNA), Bax, Bcl-2, Glut1, LDHA, stemness makers (Sox2, Oct4, and ALDHA1), and extracellular regulated protein kinase (ERK)/mammalian target of rapamycin (mTOR) pathway proteins. In vivo experiments, BALB/c nude mice (n = 12) were inoculated with 200 μL HT29 cells (5 × 106 cells) transfected with sh-FABP4 or short hairpin (sh)-negative control (NC), forming two groups with 6 mice each. The in vivo mice tumor model allowed for evaluating FABP4's impact on tumor growth. FABP4 was significantly upregulated in colorectal cancer tissues and cells (p < 0.05). FABP4 knockdown markedly inhibited cell proliferation, stemness, and glycolysis, while promoting apoptosis in these cells (p < 0.05). Additionally, FABP4 depletion led to a significant increase in ROS level (p < 0.05). However, N-acetyl-L-cysteine (NAC) (p < 0.05), a ROS scavenger, mitigates these effects. Furthermore, the effects of FABP4 depletion on cell growth, stemness, glycolysis, and apoptosis in colorectal cancer cells were also retarded by NAC (p < 0.05). Notably, FABP4 knockdown also suppressed the ERK/mTOR pathway, suggesting its regulation via ROS (p < 0.05). In vivo study results showed, FABP4 depletion significantly curbed tumor growth in colorectal cancer (p < 0.05). These results suggest that FABP4 depletion inhibits colorectal cancer progression by modulating cell growth, stemness, glycolysis and apoptosis. This regulation occurs through the ROS/ERK/mTOR pathway.

Catalase promotes whitefly adaptation to high temperature by eliminating reactive oxygen species.

Thermal stress usually leads to excessive production of reactive oxygen species (ROS) in all aerobic organisms. Catalases (CAT) are the key antioxidant enzymes, which act as the first line of defense against ROS in the antioxidant pathway. The highly invasive and widely distributed whitefly Bemisia tabaci MED damages plants by feeding as well as by transmitting many plant viruses. Previous studies have shown that strong adaptability to high temperature helps explain the spread of MED around the world. However, the mechanism underlying high temperature adaptation of this pest is not well understood. In this study, 6 CAT genes were identified from the MED genome and transcriptome dataset, among which BtCAT1, BtCAT2, and BtCAT3 were found to be highly expressed in adults. The expression of BtCAT1, BtCAT2, or BtCAT3 increased with induction temperature and induction time. The MED was exposed with mean high temperature (30°C or 35°C) and a short-term extremely high temperature (39°C or 41°C) after the silencing of BtCAT1, BtCAT2, or BtCAT3 to significantly increased ROS levels by at least 0.5 times and significantly decreased survival rate and fecundity of MED adults. The ROS level in the treated specimens gradually returned to a normal level after 24h at 25°C, but the survival rate still declined significantly. Taken together, our results demonstrate that CAT could help B. tabaci adapt to long-term mean high temperatures and short-term extremely high temperatures by eliminating excessive ROS.

HslO ameliorates arrested ΔrecA polA cell growth and reduces DNA damage and oxidative stress responses

Chromosome damage combined with defective recombinase activity has been widely considered to render cells inviable, owing to deficient double-strand break repair. However, temperature-sensitive recAts polA cells grow well upon induction of DNA damage and supplementation with catalase at restrictive temperatures. These treatments reduce intracellular reactive oxygen species (ROS) levels, which suggests that recAts polA cells are susceptible to ROS, but not chronic chromosome damage. Therefore, we investigated whether polA cells can tolerate a complete lack of recombinase function. We introduced a ΔrecA allele in polA cells in the presence or absence of the hslO-encoding redox molecular chaperon Hsp33 expression plasmid. Induction of the hslO gene with IPTG resulted in increased cell viability in ΔrecA polA cells with the hslO expression plasmid. ΔrecA polA cells in the absence of the hslO expression plasmid showed rich medium sensitivity with increasing ROS levels. Adding catalase to the culture medium considerably rescued growth arrest and decreased ROS. These results suggest that hslO expression manages oxidative stress to an acceptable level in cells with oxidative damage and rescues cell growth. Overall, ROS may regulate several processes, from damage response to cell division, via ROS-sensitive cell metabolism.

Synthesis and Antiproliferative Effect of Halogenated Coumarin Derivatives.

A series of 6- and 6,8-halocoumarin derivatives have been investigated as potential antiproliferative compounds against a panel of tumor and normal cell lines. Cytotoxic effects were determined by the MTT method. To investigate the potential molecular mechanism involved in the cytotoxic effect, apoptosis assay, cell cycle analysis, reactive oxygen species (ROS), and reduced glutathione analysis were performed. Among the screened compounds, coumarins 6,8-dibromo-2-oxo-2H-chromene-3-carbonitrile 2h and 6,8-diiodo-2-oxo-2H-chromene-3-carbonitrile 2k exhibited the most antiproliferative effect in thyroid cancer-derived cells TPC-1. The apoptosis assay showed that both 2h and 2k induced apoptosis in TPC-1 thyroid cancer cells. According to these experiments, both coumarins induced a slight increase in TPC-1 cells in the G2/M phase and a decrease in the S phase. A significant increase in ROS levels was observed in TPC-1 treated with diiodocoumarin 2k, while the dibromocoumarin 2h induced a decrease in ROS in a dose and time-dependent manner.

The nuclear factor erythroid 2-related factor 2 agonist tert-butylhydroquinone improves bone marrow mesenchymal stromal cell function in prolonged isolated thrombocytopenia after allogeneic haematopoietic stem cell transplantation.

Prolonged isolated thrombocytopenia (PT) is a life-threatening comorbidity associated with allogeneic haematopoietic stem cell transplantation (allo-HSCT). Our previous study indicated that dysfunctional bone marrow mesenchymal stromal cells (BM MSCs) played a role in PT pathogenesis and that reactive oxygen species (ROS) accumulation was related to BM MSC senescence and apoptosis. However, the mechanism of the increase in ROS levels in the BM MSCs of PT patients is unknown. In the current case-control study, we investigated whether nuclear factor erythroid 2-related factor 2 (NRF2), which is a central regulator of the cellular anti-oxidant response that can clear ROS in human BM MSCs, was associated with PT after allo-HSCT. We evaluated whether an NRF2 agonist (tert-butylhydroquinone, TBHQ) could enhance BM MSCs from PT patients in vitro. We found that BM MSCs from PT patients exhibited increased ROS levels and reduced NRF2 expression. Multivariate analysis showed that low NRF2 expression was an independent risk factor for primary PT [p=0.032, Odds ratio (OR) 0.868, 95% confidence interval (CI) 0.764-0.988]. In-vitro treatment with TBHQ improved the quantity and function of BM MSCs from PT patients by downregulating ROS levels and rescued the impaired BM MSC support of megakaryocytopoiesis. In conclusion, these results suggested that NRF2 downregulation in human BM MSCs might be involved in the pathogenesis of PT after allo-HSCT and that BM MSC impairment could be improved by NRF2 agonist in vitro. Although further validation is needed, our data indicate that NRF2 agonists might be a potential therapeutic approach for PT patients after allo-HSCT.

Inhibitory effects of Ipomoea cairica extracts on the harmful algae Phaeocystis globosa.

Ipomoea cairica (L.) Sweet is an invasive plant that cause serious invasion and damage in South China. Phaeocystis globosa is a common harmful algal bloom species on the southeast coast of China. Both species cause great environmental disturbances and serious economic damage to the localregion. This study explored the potential inhibitory effects of I. cairica leaf extracts on P. globosa. The results showed that solitary cells growth was inhibited at extract concentrations higher than 0.25% (v/v). Although the colony diameter did not change, and the colony number increased rapidly in the first 36h, we found that cells in the colonies had been damaged using scanning electron microscope and SYTOX-Green staining at 48h. In addition, the rapid light-response curve of cells treated with extracts decreased, along with down-regulation of photosynthesis-related genes (psbA, psbD, and rbcL), suggesting damage to the photosynthetic system. Finally, the activities of antioxidant enzymes including superoxide dismutase, peroxidase, and catalase increased with increasing treatment time, indicating that cells activate antioxidant enzyme defense systems to alleviate the production of reactive oxygen species (ROS). Increased ROS levels disrupt cell membranes, alter cellular ultrastructures, and ultimately lead to cell death. This study not only achieved the reuse of invasive plant resources, but also demonstrated that I. cairica leaf extract has potential value as an algaecide.

Triptolide promotes ferroptosis by suppressing Nrf2 to overcome leukemia cell resistance to doxorubicin.

Doxorubicin (DOX) is an extensively used chemotherapeutic drug to treat leukemia. However, there remains a pivotal clinical problem of resistance to DOX in patients with leukemia. Erythroid 2‑related factor2 (Nrf2) is a master regulator of antioxidation response which serves a critical role in maintaining cellular oxidative homeostasis. However, whether Nrf2 is involved in DOX resistance is not totally clear. It is well‑documented that triptolide, a widely used drug to treat autoimmune disorders, possesses anti‑cancer activities, yet whether triptolide affects leukemia cell sensitivity to DOX remains to be elucidated. The present study aimed to determine the role of triptolide‑mediated downregulation of Nrf2 in regulating leukemia cell ferroptosis and resistance to DOX. For this purpose, low‑dose DOX was used to establish DOX‑resistant K562 cells and HL‑60 cells. Nrf2 mRNA and protein expression were examined by quantitative PCR and western blotting assays. The effects of triptolide on leukemia cell viability, reactive oxygen species (ROS) levels, or lipid oxidation were determined by CCK8 assay, DCFH‑DA assay, or BODIPY 581/591 C11 assay, respectively. The results show that Nrf2 expression was significantly upregulated in DOX‑resistant leukemia cells and clinical leukemia samples. Silencing of Nrf2 significantly sensitized leukemia cells to DOX. Furthermore, it was demonstrated that triptolide inhibited Nrf2 expression and induced leukemia cell ferroptosis, as evidenced by increased ROS levels and lipid oxidation as well as decreased glutathione peroxidase4 expression. Ectopic expression of Nrf2 significantly rescued triptolide‑induced leukemia cell ferroptosis. Notably, the present study showed that triptolide re‑sensitized DOX‑resistant leukemia cells to DOX. In conclusion, the present study indicated that Nrf2 served a critical role in leukemia cell resistance to DOX and triptolide‑induced ferroptosis and suggested a potential strategy of combination therapy using triptolide and DOX in leukemia treatment.

Selenium deficiency increased duodenal permeability and decreased expression of antimicrobial peptides by activating ROS/NF-κB signal pathway in chickens.

Selenium (Se) is an essential trace element for the body. Various organs of the body, including the intestine, are affected by its deficiency. Se deficiency can induce oxidative stress and inflammatory responses in the intestine. It can also increase intestinal permeability and decrease intestinal immune function in mammals. However, the detailed studies, conducted on the intestinal molecular mechanisms of Se deficiency-induced injury in poultry, are limited. This study explored the adverse effects of Se deficiency on intestinal permeability and its mechanism. A Se-deficient chicken model was established, and the morphological changes in the chicken duodenum tissues were observed using a light microscope and transmission electron microscope (TEM). Western blotting, qRT-PCR, and other methods were used to detect the expression levels of selenoproteins, oxidative stress indicators, inflammatory factors, tight junction (TJ) proteins, antimicrobial peptides, and other related indicators in intestinal tissues. The results showed that Se deficiency could decrease the expression levels of selenoproteins and antioxidant capacity, activate the nuclear factor kappa-B (NF-κB) pathway, cause inflammation, and decrease the expression levels of TJ proteins and antimicrobial peptides in the duodenum tissues. The study also demonstrated that Se deficiency could increase intestinal permeability and decrease antimicrobial peptides via reactive oxygen species (ROS)/NF-κB. This study provided a theoretical basis for the scientific prevention and control of Se deficiency in poultry. Se deficiency decreased the expression levels of selenoproteins and increased ROS levels to activate the NF-κB pathway, resulting in the production of pro-inflammatory cytokines, reducing the expression levels of TJ protein, and weakening the expression of antimicrobial peptides, which contributed to the higher intestinal permeability. Oxidative stress weakened the expression of antimicrobial peptides.

Effects of Antioxidant Gene Overexpression on Stress Resistance and Malignization In Vitro and In Vivo: A Review.

Reactive oxygen species (ROS) are normal products of a number of biochemical reactions and are important signaling molecules. However, at the same time, they are toxic to cells and have to be strictly regulated by their antioxidant systems. The etiology and pathogenesis of many diseases are associated with increased ROS levels, and many external stress factors directly or indirectly cause oxidative stress in cells. Within this context, the overexpression of genes encoding the proteins in antioxidant systems seems to have become a viable approach to decrease the oxidative stress caused by pathological conditions and to increase cellular stress resistance. However, such manipulations unavoidably lead to side effects, the most dangerous of which is an increased probability of healthy tissue malignization or increased tumor aggression. The aims of the present review were to collect and systematize the results of studies devoted to the effects resulting from the overexpression of antioxidant system genes on stress resistance and carcinogenesis in vitro and in vivo. In most cases, the overexpression of these genes was shown to increase cell and organism resistances to factors that induce oxidative and genotoxic stress but to also have different effects on cancer initiation and promotion. The last fact greatly limits perspectives of such manipulations in practice. The overexpression of GPX3 and SOD3 encoding secreted proteins seems to be the "safest" among the genes that can increase cell resistance to oxidative stress. High efficiency and safety potential can also be found for SOD2 overexpression in combinations with GPX1 or CAT and for similar combinations that lead to no significant changes in H2O2 levels. Accumulation, systematization, and the integral analysis of data on antioxidant gene overexpression effects can help to develop approaches for practical uses in biomedical and agricultural areas. Additionally, a number of factors such as genetic and functional context, cell and tissue type, differences in the function of transcripts of one and the same gene, regulatory interactions, and additional functions should be taken into account.

Out of the Shadow: Blue Light Exposure Induces Apoptosis in Müller Cells

Awareness toward the risks of blue light (BL) exposure is rising due to increased use of BL-enriched LEDs in displays. Short-wave BL (400-500 nm) has a high photochemical energy, leading to the enhanced production of reactive oxygen species (ROS). BL potentially plays a role in causing dry eye, cataracts, and age-related macular degeneration (AMD). The effect of BL on retinal pigment epithelium cells (RPEs) or photoreceptors has been extensively investigated. In contrast, only a few studies have investigated the effects of BL exposure on Müller cells (MCs). This is mainly due to their lack of photosensitive elements and the common assumption that their reaction to stress is only secondary in disease development. However, MCs perform important supportive, secretory, and immune functions in the retina, making them essential for retinal survival. Increased oxidative stress is a key player in many retinal diseases such as AMD or glaucoma. We hypothesize that increased oxidative stress can also affect MCs. Thus, we simulated oxidative stress levels by exposing primary porcine MCs and human MIO-M1 cells to BL. To confirm the wavelength-specificity, the cells were further exposed to red (RL), purple (PL), and white light (WL). BL and WL exposure increased ROS levels, but only BL exposure led to apoptosis in primary MCs. Thus, BL accounted for the harmful part of WL exposure. When cells were simultaneously exposed to BL and RL (i.e., PL), cell damage due to BL could be partly prevented, as could the inhibition of p53, demonstrating the protective effect of RL and p53 dependency. In contrast, BL hardly induced apoptosis in MIO-M1 cells, which is likely due to the immortalization of the cells. Therefore, enhanced oxidative stress levels can significantly harm MC function, probably leading to decreased retinal survival and, thus, further enhancing the progression of retinal diseases. Preventing the cell death of these essential retinal cells represents a promising therapy option to enhance retinal survival.

The Genotoxicity of Acrylfentanyl, Ocfentanyl and Furanylfentanyl Raises the Concern of Long-Term Consequences.

Three fentanyl analogues Acrylfentanyl, Ocfentanyl and Furanylfentanyl are potent, rapid-acting synthetic analgesics that recently appeared on the illicit market of new psychoactive substances (NPS) under the class of new synthetic opioids (NSO). Pharmacotoxicological data on these three non-pharmaceutical fentanyl analogues are limited and studies on their genotoxicity are not yet available. Therefore, the aim of the present study was to investigate this property. The ability to induce structural and numerical chromosomal aberrations in human lymphoblastoid TK6 cells was evaluated by employing the flow cytometric protocol of the in vitro mammalian cell micronucleus test. Our study demonstrated the non-genotoxicity of Fentanyl, i.e., the pharmaceutical progenitor of the class, while its illicit non-pharmaceutical analogues were found to be genotoxic. In particular, Acrylfentanyl led to a statistically significant increase in the MNi frequency at the highest concentration tested (75 μM), while Ocfentanyl and Furanylfentnyl each did so at both concentrations tested (150, 200 μM and 25, 50 μM, respectively). The study ended by investigating reactive oxygen species (ROS) induction as a possible mechanism linked to the proved genotoxic effect. The results showed a non-statistically significant increase in ROS levels in the cultures treated with all molecules under study. Overall, the proved genotoxicity raises concern about the possibility of serious long-term consequences.

Hypoxia-Induced Oxidative Stress Promotes Proteasome Inhibitor Resistance Via Upregulation of Heme Oxygenase-1 in Multiple Myeloma

Background Multiple myeloma (MM) is a hematopoietic malignancy for which proteasome inhibitors have become available in recent years. However, many patients develop resistance to these drugs during treatment. Therefore, it is important to elucidate the mechanisms underlying resistance acquisition by proteasome inhibitors. Side population (SP) cells, which have a high drug efflux capacity, as well as hypoxic responses in the microenvironment, have both provided important insights into drug resistance in MM; however, little is known about the characteristics of SP cells in hypoxic microenvironments. Aim and Methods To find novel therapeutic target for drug resistant MM cells, the SP cells and non-SP cells of MM cell lines (RPMI-8226 and KMS-11) were sorted for normoxic and hypoxic cultures (1% O2) for 48 h, respectively, and cDNA microarray assays were performed on these samples. Then, we conducted functional analysis of hypoxia-SP specific genes. Results Our comprehensive gene expression analysis identified four coding genes (HMOX1, BACH2, DUX4, and RASGEF1B) and seven non-coding genes (TUSC8, LOC101928738, lnc-WRAP73-1, LINC-PINT, LINC01004, MIR146A, and lnc-FAM133B-1) that are specifically highly expressed in SP cells under hypoxic conditions. Among protein-coding genes, RT-qPCR showed very low expression of BACH2 and no expression of DUX4; we decided to investigate the function of HMOX1/heme oxygenase-1 (HO-1) in hypoxic environments. One of the functions of HO-1 is to protect cells from apoptosis induced by reactive oxygen species (ROS). We found a significant increase in ROS under hypoxic conditions compared with normoxic conditions in MM cells, including MM patient samples. HMOX1 knockdown significantly increased ROS levels under both normoxic and hypoxic conditions. These results suggest that hypoxic stress induced ROS and that HO-1 can neutralize hypoxia-induced ROS. The addition of PMA, a ROS inducer, significantly increased HMOX1/HO-1 in MM cells. Conversely, the addition of NAC, a ROS neutralizer, suppressed HMOX1/HO-1 expression under hypoxic conditions. These results suggest that HO-1 is induced in hypoxic environments by a ROS-mediated pathway. Next, we investigated the involvement of HO-1 in the effects of proteasome inhibitors in hypoxic environments. MTX assays showed that bortezomib against HMOX1 knockdown KMS-11 reduced cell viability under hypoxic rather than normoxic conditions. We transplanted HMOX1 knockdown or control KMS-11 (1×106 cells) into immunodeficient mice, injected bortezomib (1.0 mg/kg) intraperitoneally on days 14 and 18, and sacrificed the mice on day 22. There was no significant difference in subcutaneous tumor volume and weight between the control and HMOX1 knockdown cell lines, whereas HMOX1 knockdown resulted in a significant bortezomib-induced reduction in subcutaneous tumor volume and weight (p = 0.0009 and 0.0096, respectively). Lastly, we examined whether excessive levels of ROS enhance the effect of bortezomib. PMA and a low concentration of bortezomib (5 nM, which is lower than that for IC50) were added to KMS-11 and RPMI-8226, and an apoptosis assay was performed. As a result, PMA or a low concentration of bortezomib increased apoptotic cells by only a small percentage, while simultaneous exposure to both significantly increased apoptotic cells synergistically. This result indicates that excessive ROS levels enhance the anti-myeloma effect of proteasome inhibitors. Additionally, in clinical datasets, HMOX1 had a strong and significantly positive correlation with MAFB, but not MAF. We investigated whether MAFB actually regulates HMOX1 under hypoxic conditions. MAFB/MafB was induced by hypoxia even in a cell line without t(14;20) (q32;q12), such as KMS-11. Knockdown of MAFB significantly decreased HMOX1/HO-1 expression only under hypoxic condition, suggesting that MafB induction is involved in the elevated expression of HO-1 under hypoxic conditions. Conclusion We clarified the contribution of the hypoxia-ROS-HO-1 axis to proteasome inhibitor resistance in hypoxic environments. This axis might also be involved in hypoxia-induced MafB expression. Because mechanisms that reduce excessive ROS in hypoxic microenvironments are thought to be closely related to cell survival and drug resistance, targeting this mechanism may be a new strategy to overcome drug resistance in refractory MM.