Nrf2 Levels Research Articles

Polydatin protects against DSS-induced ulcerative colitis via Nrf2/Slc7a11/Gpx4-dependent inhibition of ferroptosis signalling activation

IntroductionUlcerative colitis (UC), a form of inflammatory irritable bowel disease, is characterized by a recurrent and persistent nonspecific inflammatory response. Polydatin (PD), a natural stilbenoid polyphenol with potent properties, exhibits unexpected beneficial effects beyond its well-documented anti-inflammatory and antioxidant activities. In this study, we presented evidence that PD confers protection against dextran sodium sulfate (DSS)-induced ulcerative colitis.MethodsThe protective effect of PD on colitis was examined in cultured caco-2 cells and DSS-induced colitis mouse model. Bulk RNA sequencing and differential gene expression analysis were used to investigate the protective mechanism of PD on DSS-induced colitis. Ferroptosis was determined by MDA levels, SOD levels, mitochondrial iron accumulation and ROS production. Ferroptosis-related proteins Slc7a11, Nrf2 and Gpx4 levels were measured by western blot, immunohistochemical and immunofluorescence staining.ResultsPD mitigated the DSS-induced increases in pro-inflammatory cytokines (IL-6, TNF-α, and IL-1β), alleviated colon length shortening, reduced morphological damage to the intestinal mucosa, and preserved tight junction proteins (TJ) occludin and Zonula occludens-1 (ZO-1) in both caco-2 cells and murine models of colitis. Mechanistically, PD reversed the reduction of Nrf2, Slc7a11 and Gpx4, the degree of nuclear translocation of Nrf2 induced by DSS in vitro and in vivo significantly. Moreover, the protective effect of PD is attenuated by erastin and resembled that of Fer-1 in caco-2 cells model.DiscussionOur study suggested that PD protects against DSS-induced ulcerative colitis via Nrf2/Slc7a11/Gpx4-dependent inhibition of ferroptosis signalling activation. Further investigation into the precise mechanisms underlying this phenomenon is warranted. The findings presented herein indicated that PD may serve as a potential therapeutic agent for patients with UC.

Physalis Calyx seu Fructus relieves chicken intestinal damage to heat via improving the antioxidant ability

Heat-stress-induced oxidative and inflammatory responses were important factors contributing to chicken intestinal damage. The purpose of this study was based on the antioxidant and anti-inflammatory activities of Physalis Calyx seu Fructus (Jin Deng Long, JDL) to investigate its efficacy and mechanism in relieving chicken heat stress damage. Primary chicken embryo duodenum cells and 90 30-day-old specific-pathogen-free chicken were randomly divided into control and JDL groups to establish heat stress models in vitro and in vivo. The mitigating effect was assessed through the oxidation-related enzymes and key genes, histopathology, and inflammatory factors. The results demonstrated that 100 µg/mL JDL extract could effectively alleviate heat stress damage to chicken embryo duodenum cells at 42°C. A strong antioxidant capacity of 100 µg/mL JDL extract was shown in the downregulation of LDH (at 5 h, P < 0.01) and MDA (at 5 h, P < 0.05), in the upregulation of SOD (at 5 and 10 h, P < 0.01), CAT (at 5 h, P < 0.01), and GSH-PX and T-AOC (at 0 h, P < 0.01) as well as in the high transcription level of NQO1 (at 5 and 10 h, P < 0.05) and HO-1 (at 5 and 10 h, P < 0.01). Supplements with 1 and 3 g/kg b.wt, respectively, in the drinking water both suppressed the rise of body temperature and had light pathological lesions of chicken duodenal tissues caused by heat stress at 40 ± 1°C. Accordingly, the chicken of JDL extract groups showed a lower inflammatory response as manifested by a lower level of IL-10 and higher levels of IL-6 and TNF-α and a strong antioxidant capacity characterized by lower level of MDA and higher levels of SOD and GSH-PX in the serum as well as also showed a higher transcription level of Nrf2, NQO1, and HO-1 in the duodenal tissues. In conclusion, JDL extract relieved chicken intestinal damage to heat via improving the antioxidant ability and reducing the inflammatory response.

Maternal exercise before and during pregnancy protects against genotoxicity and promotes offspring hippocampal health in mice prenatally exposed to high fructose.

The fetal brain is susceptible to programming effects during pregnancy, potentially leading to long-term consequences for offspring's cognitive health. Fructose intake is thought to adversely affect fetal brain development, whereas physical exercise before and during pregnancy may be protective. Therefore, this study aimed to assess biochemical and genotoxic changes in maternal hippocampi and behavioral, genotoxic, and biochemical alterations in offspring hippocampi. Seventy female mice were exposed to fructose (20%/L) and/or voluntary physical exercise (VPE) pre-pregnancy for eight weeks, and then mated and exposure was continued until weaning. Offspring were evaluated at 60 days old using behavioral test, genotoxic and biochemical markers. Fructose induced long-term memory impairment in male offspring, which was alleviated by VPE. VPE mitigated DNA damage from maternal fructose consumption in both maternal and offspring hippocampi in female offspring, VPE increased levels of APE-1, NRF2, and CREB proteins, whereas in males, OGG1 levels upregulate. Fructose consumption led to oxidative stress and antioxidant defense alterations in offspring, while VPE mitigated these effects. Telomere shortening was observed in male offspring from mothers who consumed fructose during pregnancy. Our findings suggest that exposure to fructose during (pre)pregnancy and lactation has adverse effects on offspring's hippocampi later in life, and VPE has a protective effect. Overall, the study underscores the significance of maternal dietary and physical habits on long term offspring health, with an emphasis on implications for adult cognitive function.

Ethyl Acetate Extract of Cichorium glandulosum Activates the P21/Nrf2/HO-1 Pathway to Alleviate Oxidative Stress in a Mouse Model of Alcoholic Liver Disease

Background: Alcoholic liver disease (ALD) is a significant global health concern, primarily resulting from chronic alcohol consumption, with oxidative stress as a key driver. The ethyl acetate extract of Cichorium glandulosum (CGE) exhibits antioxidant and hepatoprotective properties, but its detailed mechanism of action against ALD remains unclear. This study investigates the effects and mechanisms of CGE in alleviating alcohol-induced oxidative stress and liver injury. Methods: Ultra-Performance Liquid Chromatography coupled with Quadrupole-Orbitrap Mass Spectrometry (UPLC-Q-Orbitrap-MS) was used to identify CGE components. A C57BL/6J mouse model of ALD was established via daily oral ethanol (56%) for six weeks, with CGE treatment at low (100 mg/kg) and high doses (200 mg/kg). Silibinin (100 mg/kg) served as a positive control. Liver function markers, oxidative stress indicators, and inflammatory markers were assessed. Transcriptomic and network pharmacology analyses identified key genes and pathways, validated by reverse transcription quantitative polymerase chain reaction (RT-qPCR) and Western blotting. Results: UPLC-Q-Orbitrap-MS identified 81 CGE compounds, mainly including terpenoids, flavonoids, and phenylpropanoids. CGE significantly ameliorated liver injury by reducing alanine aminotransferase (ALT), aspartate aminotransferase (AST), and alkaline phosphatase (ALP) levels and enhancing antioxidative markers such as total antioxidant capacity (T-AOC) and total superoxide dismutase (T-SOD) while lowering hepatic malondialdehyde (MDA) levels. Inflammation was mitigated through reduced levels of Tumor Necrosis Factor Alpha (TNF-α), Interleukin-1 Beta (IL-1β), and C-X-C Motif Chemokine Ligand 10 (CXCL-10). Transcriptomic and network pharmacology analysis revealed seven key antioxidant-related genes, including HMOX1, RSAD2, BCL6, CDKN1A, THBD, SLC2A4, and TGFβ3, validated by RT-qPCR. CGE activated the P21/Nuclear Factor Erythroid 2-Related Factor 2 (Nrf2)/Heme Oxygenase-1 (HO-1) signaling axis, increasing P21, Nrf2, and HO-1 protein levels while suppressing Kelch-like ECH-associated Protein 1 (Keap1) expression. Conclusions: CGE mitigates oxidative stress and liver injury by activating the P21/Nrf2/HO-1 pathway and regulating antioxidant genes. Its hepatoprotective effects and multi-target mechanisms highlight CGE’s potential as a promising therapeutic candidate for ALD treatment.

Nrf2 mediates mitochondrial and NADPH oxidase-derived ROS during mild heat stress at 40 °C.

Hyperthermia is an adjuvant to chemotherapy and radiotherapy and sensitizes tumors to these treatments. However, repeated heat treatments result in acquisition of heat resistance (thermotolerance) in tumors. Thermotolerance is an adaptive survival response that appears to be mediated by upregulated cellular defenses. However, the mechanisms of activation remain unclear. When HeLa cells were exposed to mild heat shock at 40 °C for 3 h, levels of superoxide and peroxides increased. Cells were treated with mitochondrial antioxidant MitoQ and NADPH oxidase (NOX) inhibitor apocynin to characterize the contribution of these two sources to the total reactive oxygen species (ROS) pool. We found that both mitochondria and NOX are sources of ROS during mild heat shock at 40 °C. Heat-derived ROS are thought to activate the adaptive survival response at 40 °C. Nrf2, the master regulator of the cellular antioxidant response, is thought to play a pivotal role in establishing the adaptive survival response. Nrf2 was overexpressed or knocked down to assess its role. Moreover, Nrf2 levels correlate with the cellular redox state, and do so via scavenging of mitochondria- and NOX-derived ROS. Knockdown of Nrf2 markedly increased levels of ROS that were scavenged by either apocynin or MitoQ. Finally, critical defense proteins such as DJ-1 and PGAM5 seemed to require a two-key activation system mediated by Nrf2 and mitochondrial ROS. Our study characterized mitochondrial and NOX-derived ROS as being essential in activating cellular defenses alongside Nrf2 and underlines potential therapeutic targets that may contribute to the acquisition of thermotolerance.

Discovery of 15-deoxynaphthomycins activating the antioxidant NRF2-ARE pathway from Streptomyces sp. N50 via genome mining, global regulator introduction, and molecular networking

Genome mining is a promising avenue for expanding the repertoire of microbial natural products, which are important for drug development. This approach involves predicting genetically encoded small molecules by examining bacterial genomes via accumulated knowledge of microbial biosynthesis. However, it is also important that the microbes produce the predicted molecule in practice. Here, we introduce an endophytic Streptomyces sp. N50, which was isolated from the medicinal plant Selaginella tamariscina. Upon sequencing its entire genome, 33 biosynthetic gene clusters (BGCs) were identified in a chromosome and a megaplasmid. Subsequent genome mining revealed that the new 15-deoxynaphthomycin could be produced due to the presence of an enoyl reductase domain, which is absent in the known BGC of naphthomycin, a type of ansamycin antibiotics. In addition, the engineered strain with the introduction of the global regulatory gene afsR2 into N50 successfully produced 15-deoxynaphthomycins. Furthermore, molecular network analysis via MS/MS selectively confirmed the presence of additional sulfur-containing 15-deoxynaphthomycin congeners. Eventually, six new 15-deoxynaphthomycins were isolated and elucidated from the engineered strain N50. This family of compounds is known to exhibit various biological activities. Also, the presence of quinone moieties in these compounds, which are known to activate NRF2, they were tested for their ability to activate NRF2. Among the new compounds, three (1, 5, and 6) activated the antioxidant NRF2-ARE signaling pathway. Treatment with these compounds significantly elevated NRF2 levels in HepG2 cells and further induced the expression of NRF2 target genes associated with the antioxidant response. This study suggests that the combination of genome mining, gene engineering and molecular networking is helpful for generating new small molecules as pharmaceutical candidates from microorganisms.

SENP1 inhibits aerobic glycolysis in Aβ1-42-incubated astrocytes by promoting PUM2 deSUMOylation

Alzheimer’s disease (AD), the most prevalent form of dementia in the elderly, involves critical changes such as reduced aerobic glycolysis in astrocytes and increased neuronal apoptosis, both of which are significant in the disease’s pathology. In our study, astrocytes treated with amyloid β1-42 (Aβ1-42) to simulate AD conditions exhibited upregulated expressions of small ubiquitin-like modifier (SUMO)-specific protease 1 (SENP1) and Pumilio RNA Binding Family Member 2 (PUM2), alongside decreased levels of Nuclear factor erythroid 2-related factor 2 (NRF2). SENP1 is notably the most upregulated SUMOylation enzyme in Aβ1-42-exposed astrocytes. Functional assays including Ni2+-Nitrilotriacetic acid (NTA) agarose bead pull-down and co-immunoprecipitation (Co-IP) confirmed SENP1’s role in actively deSUMOylating PUM2, thereby enhancing its stability and expression. The interaction between PUM2 and the 3’ untranslated region (3’UTR) of NRF2 mRNA reduces NRF2 levels, subsequently diminishing the transcriptional activation of critical glycolytic enzymes, Hexokinase 1 (HK1) and Glucose Transporter 1 (GLUT1). These changes contribute to the observed reduction in glycolytic function in astrocytes, exacerbating neuronal apoptosis. Targeted interventions, such as knockdown of Senp1 or Pum2 or overexpression of NRF2 in APPswe/PSEN1dE9 (APP/PS1) transgenic mice, effectively increased HK1 and GLUT1 levels, decreased apoptosis, and alleviated cognitive impairment. These findings highlight the important roles of the SENP1/PUM2/NRF2 pathway in influencing glucose metabolism in astrocytes, presenting new potential therapeutic targets for AD.Graphical 1. SENP1 deSUMOylates PUM2 enhancing its expression in Aβ1-42-microenvironment.2. PUM2 binds to NRF2 mRNA reducing its stability and expression in Aβ1-42-microenvironment.3. Reduced NRF2 reduces its transcriptional promoting effects on HK1 and GLUT1 in Aβ1-42-microenvironment.4. Reduced HK1 and GLUT1 decreases aerobic glycolysis in astrocytes, leading to an increase in co-culture neuronal apoptosis in Aβ1-42-microenvironment.

Angiotensin type 1 and type 2 receptors-induced mitochondrial dysfunction promotes ferroptosis in cardiomyocytes.

Previous studies suggest that ferroptosis is involved in cardiovascular diseases. The aim of the present study is to investigate the causal relationship between angiotensin II type 1 and type 2 receptors (AT1/2R) activities and mitochondrial dysfunction in induction of cardiomyocyte ferroptosis. Human AC16 cardiomyocytes were first pre-treated with an AT1/2R blockers, before stimulated with angiotensin II (Ang II) for 24 h. The redox status of the cardiomyocytes were assessed by measuring the cellular malondialdehyde (MDA), superoxide dismutase (SOD), and Nicotinamide-adenine dinucleotide phosphate, (NADPH) levels using biochemical methods. Mitochondrial reactive oxygen specifics (mitROS), mitochondrial memebrane potential, and Fe2+ levels were determined using flow cytometry. The signaling pathways, including the glutathione peroxidase 4 (GPX4), heme oxygenase-1 (HO-1), sirtuin1, and ferroptosis suppressor protein 1 (FSP1)-coenzyme Q10 (CoQ10) pathways, were evaluated using western blotting. Our results demonstrated that Ang II significantly elevated the levels of MDA, Fe2+, mitoROS, and FtMt and markedly reduced SOD, NADPH, mitochondrial membrane potential, GPX4, HO-1, Sirt1, SFXN1, Nrf2, and FSP1 levels in cardiomyocyte, which were reversed by blockade of AT1/2R. Our results suggest that AT1/2R signaling can induce myocardial ferroptosis by impairing mitochondrial function via multiple signaling pathways, including the cyst (e)ine /GSH/GPX4 axis and FSP1/coenzyme Q10 (CoQ10) axis.

Euglena gracilis-derived β-glucan ameliorates particulate matter (PM2.5)-induced airway inflammation by modulating nuclear factor kappa B, mitogen-activated protein kinase, and nuclear factor erythroid 2-related factor 2 signaling pathways in A549 cells and BALB/c mice.

This study aimed to investigate the effects of β-glucan derived from Euglena gracilis (EGB), an edible microalga, on particulate matter (PM2.5)-induced airway inflammation in A549 cells and BALB/c mice. EGB effectively suppressed the mRNA and protein levels of inflammatory cytokines (IL-6, IL-1β, TNF-α, IL-8) and mediators (iNOS, COX-2), while inhibiting the NF-κB and MAPK signaling pathways triggered by PM2.5 exposure and reducing nuclear NF-κB levels. Additionally, EGB decreased PM2.5-induced ROS production and increased the protein levels of NRF2 and HO-1, along with genes encoding antioxidant enzymes (catalase, GPx, SOD1), associated with elevated nuclear NRF2 levels. EGB reduced immune cell infiltration and inflammatory cytokine levels in BALF and serum, both of which increased by PM2.5 exposure. EGB also significantly increased alveolar numbers while decreasing the gene expression of MMP1/9/13. Furthermore, EGB suppressed PM2.5-induced bronchial thickening and collagen-1 deposition by downregulating TGF-β1 expression, and alleviated goblet cell hyperplasia and mucin production in lung tissues. These results suggest that EGB effectively reduces PM2.5-induced airway inflammation by suppressing NF-κB and MAPK signaling pathways, lowering pro-inflammatory cytokines, and activating the NRF2-HO-1 signaling pathway to enhance antioxidant enzyme expression. This study highlights the potential of EGB as an edible functional agent for controlling PM-related airway inflammation.

Innovative rheumatoid arthritis management using injection replacement approach via dual therapeutic effects of hyalurosomes-encapsulated luteolin and dexamethasone.

Rheumatoid arthritis is a highly prevalent debilitating condition linked to inflammation. The effectiveness of the present therapeutic techniques is constrained; so, there is an urgent requirement for a novel nanoplatform entailing drugs with proven efficacy. The current work highlighted the development of dexamethasone and luteolin co-encapsulated hyalurosomes (LUT-DEX hyalurosomes). High entrapment efficiency of 92.79 % and 81.21 % for DEX and LUT, respectively in addition to sustained release of both drugs were attained, where only 45 % DEX and 75.87 % LUT were released after 24 h indicating the possibility of a persistent therapeutic impact. A spherical nano-system with smooth edges and a characteristic layer of hyaluronic acid surrounding the core of the particles was evidenced by a transmission electron microscope. The efficacy of LUT-DEX hyalurosomes was evaluated in-vision vivo using a rat model of rheumatoid arthritis initiated by Complete Freund's Adjuvant (CFA). Histological examination and serum concentrations of malondialdehyde (MDA), interleukin 1ß (IL1ß), tumour necrosis factor-alpha (TNF-α), interleukin 3 (MMP-3), and nuclear factor (erythroid-derived) Like 2 NRF2) were also evaluated. The dual drug-loaded hyalurosomes demonstrated 2.9-, 3.2-, 2.5- and 2.7-fold decreases in MMP3, TNF-α, MDA and IL1, respectively, compared with the positive control group. Conversely, the negative control group demonstrated the highest NRF2 level followed by LUT-DEX hyalurosomes, comparison compared to the positive control group which demonstrated the lowest NRF2 level. The histological examination of the joints confirmed the superior effect of the dual drug encapsulated nano delivery system in reducing joint swelling and inflammation achieving similar results as the negative control group. Ultimately, the developed hyalurosomes co-encapsulating dexamethasone and luteolin, possess the potential to serve as a highly auspicious innovative strategy for managing rheumatoid arthritis.

Inhibition of ASIC1a reduces ferroptosis in rheumatoid arthritis articular chondrocytes via the p53/NRF2/SLC7A11 pathway.

The activation of acid-sensing ion channel 1a (ASIC1a) in response to extracellular acidification leads to an increase in extracellular calcium influx, thereby exacerbating the degeneration of articular chondrocytes in rheumatoid arthritis (RA). It has been suggested that the inhibition of extracellular calcium influx could potentially impede chondrocyte ferroptosis. The cystine transporter, solute carrier family 7 member 11 (SLC7A11), is recognized as a key regulator of ferroptosis. Recent studies suggest that the tumor suppressor gene p53 facilitates the induction of ferroptosis by suppressing the upregulation of SLC7A11. This process is mediated by the nuclear factor erythroid 2-related factor 2 (NRF2), a key transcription factor integral to the maintenance of cellular redox homeostasis and the regulation of inflammatory responses. This study aims to investigate the role of ASIC1a in the ferroptosis of RA chondrocytes and to determine the involvement of the p53/NRF2/SLC7A11 pathway in its underlying mechanism. Invitro experiments revealed that acidosis induces ferroptosis and reduces the expression of NRF2 and SLC7A11 in chondrocytes. Moreover, acidification significantly increased p53 protein levels in chondrocytes. Pifithrin-α (PFN-α), a p53 inhibitor, mitigated acidosis-induced ferroptosis and restored the diminished expression of NRF2 and SLC7A11. Furthermore, PcTx-1, an ASIC1a inhibitor, inhibited acidification-induced ferroptosis, enhanced the protein levels of SLC7A11 and NRF2, and reduced p53 expression. Invivo experiments demonstrated that the ASIC1a-specific inhibitor PcTx-1 ameliorated histopathological characteristics of ankle joints in collagen-induced arthritis (CIA) mice, decreased p53 expression, and enhanced NRF2 and SLC7A11 expression in chondrocytes. These findings suggest that ASIC1a inhibition may mitigate acidification-induced ferroptosis in articular chondrocytes in RA, potentially via the p53/NRF2/SLC7A11 pathway.

Physalins and neophysalins from the calyx of Physalis alkekengi: Structures and anti-inflammatory efficacy.

To explore potential anti-inflammatory lead compounds, ten new physalin steroids, including three neophysalins (1, 4, and 9) and seven physalins (2, 3, 5-8, and 10), along with eleven known analogs, were isolated from an ethanol extract of the calyx of Physalis alkekengi. The new structures were rigorously determined through comprehensive HRESIMS, 1D/2D-NMR, and X-ray diffraction analysis. Among these compounds, 1 was identified as a new 1,10-seco-neophysalin, and 2 was identified as a new 11,15-cyclo-9,10-seco-physalin characterized by an aromatic A-ring. Evaluation of the anti-inflammatory activities of the isolated compounds revealed that compound 15 displayed remarkable potency, inhibiting NO production in RAW 264.7 macrophages with an IC50 value of 2.26±0.12μM, while exhibiting low cytotoxicity (IC50=90.45±6.10μM). It suppressed the expression levels of IL-6, IL-1β, iNOS and COX-2, while upregulating the expression levels of HO-1 and Nrf2, indicating the involvement of multiple mechanisms. Further studies indicated that compound 15 significantly attenuated the LPS-induced increase in the phosphorylation of p38, ERK, and JNK in a dose-dependent manner, indicating its potential to exert anti-inflammatory effects through modulation of the MAPK signaling pathway. In vivo studies further demonstrated the efficacy of compound 15, as it showed inhibitory activity against mouse ear edema, achieving an inhibition rate of 37.30% at a dosage of 25mg/kg. Importantly, compound 15 (25mg/kg) significantly ameliorated colitis-related symptoms in a dextran sodium sulfate (DSS)-induced ulcerative colitis (UC) model, highlighting its potential as a therapeutic candidate for inflammatory disorders.

Comparing the antioxidant effects of single and binary combinations of Lactiplantibacillus plantarum in vitro and in vivo and their application in yogurt.

Lactiplantibacillus plantarum (L. plantarum) have been studied for their antioxidant properties, which can mitigate oxidative stress and improve health outcomes. The study aimed to compare the antioxidant properties of single and binary L. plantarum and their impact on yogurt. L. plantarum 847 (Lp-C), L. plantarum 8014 (Lp-G), and their combination were chosen for their in vitro antioxidant potential. In vivo experiments were performed in Drosophila melanogaster (D. melanogaster) and results showed that binary L. plantarum significantly improved the survival time, weight, catalase activity and intestinal integrity in H2O2-induced flies. As compared with single L. plantarum treated flies, binary strains improved the survival curve, superoxide dismutase and catalase activities in females, prolonged the average survival time in males, and increased the expression level of keap1, Nrf2 and SOD genes in all genders. To explore the effect of single and binary L. plantarum on milk fermentation, the physicochemical properties and antioxidant activity of yogurt were detected, and results presented that yogurt fermented with L. plantarum exhibited the improved antioxidant capacity, with the binary strain combination demonstrating superior effects in rheological properties and the later period of yogurt storage. This research offers a foundation for choosing the combinations of lactic acid bacteria (LAB) with antioxidant properties.

Ashwagandha (Withania somnifera (L.) dunal) root extract containing withanolide a alleviates depression-like behavior in mice by enhancing the brain-derived neurotrophic factor pathway under unexpected chronic mild stress.

Ashwagandha (Withania somnifera (L.) Dunal) root or whole-plant extracts are used to treat anxiety, insomnia, and other nervous system disturbances. We evaluated the neuroprotective and antidepressant effects of ashwagandha root extract (ARE) on corticosterone-exposed HT-22cells and unpredictable chronic mild stress (UCMS)-challenged mice. The neuroprotective properties of ARE containing withanolide A were assessed in HT-22cells subjected to corticosterone-induced oxidative stress. Additionally, the effects of ARE on depression-like behavior, stress-related hormones, and inflammatory cytokine levels were evaluated in a mouse model of UCMS. In HT-22cells, ARE (100 and 200μg/mL) and its constituent, withanolide A (1.56 and 3.12μg/mL), mitigated corticosterone-induced increases in MAO activity, ROS, and MDA levels. Treatment also reversed corticosterone-induced reductions in BDNF, TrkB, p-AKT, p-ERK, and p-CREB and normalized Nrf2 and Keap1 levels, thereby elevating HO-1 expression. In UCMS mice, ARE improved behavioral outcomes, increased sucrose preference, and reduced immobility in the forced swimming test while enhancing activity in the open field test and elevated plus maze. ARE decreased the levels of stress hormones (corticotropin-releasing hormone, adrenocorticotropic hormone, and corticosterone) and increased the levels of neurotransmitters (L-DOPA, 5-HTP, and serotonin). Histological analysis revealed that ARE reduced hippocampal cell loss. Additionally, ARE (60 and 100mg/kg) restored decreased levels of p-AKT, p-ERK, and p-CREB and lowered inflammation-related proteins (Cox2, iNOS, IL-6, IL-1β, TNF-α). These results indicate that ARE containing withanolide A exhibits notable neuroprotective and antidepressant properties.

Iristectorin A Ameliorates Cisplatin-Induced Hepatorenal Injury in Mice Through Modulation of the Nrf2/HO-1 Signaling Pathway.

Cisplatin (CIS) is a chemotherapeutic agent frequently used in cancer treatment. However, depending on the dosage and duration of use, CIS can lead to hepatotoxicity and nephrotoxicity. Iristectorin A (IRIS), a natural flavonoid, has been found to exhibit antioxidant and protective effects. In this paper, we scrutinized the effects and molecular mechanisms of the IRIS on CIS-induced liver and kidney damage in mice. IRIS administration alleviated CIS-induced elevations in AST, ALT, ALP, BUN, and creatinine levels by approximately 12%, 15%, 11%, 21%, and 15%, respectively. It also inhibited liver and kidney MDA levels by approximately 29% and 28%, while enhancing liver and kidney GSH, SOD, and CAT levels by 47%-60%, 85%-70%, and 90%-55%, respectively. IRIS enhanced liver and kidney mRNA expression levels of Nrf2 (by approximately 1.6- and 1.5-fold, respectively), HO-1 (by 1.5- and 1.5-fold, respectively), and Bcl-2 (by 1.5- and 1.4-fold, respectively). In addition, IRIS suppressed the mRNA expression levels of NF-κB (by 0.7- and 0.7-fold), TNF-α (by 0.7- and 0.7-fold), Bax (by 0.8- and 0.7-fold), and Cas-3 (by 0.9- and 0.7-fold). Protein expression analysis revealed that IRIS increased Nrf2 (by 1.5- to 1.2-fold) and Bcl-2 levels (by 1.3- to 1.7-fold), and reduced Bax (by 0.7- to 0.8-fold) and Cas-3 (by 0.8- and 0.8-fold) levels altered by CIS treatment. Moreover, IRIS administration prevented histopathological changes in the liver and kidney caused by CIS. Ultimately, IRIS was found to substantially mitigate CIS-induced hepatorenal injury by targeting oxidative stress, inflammation, and apoptosis through regulation of the Nrf2/HO-1 signaling pathway. Therefore, IRIS holds potential as a therapeutic adjuvant in the use of CIS.

Cold stimulated bronchial epithelial cells derived exosomes HMGB1 aggravates bronchial epithelial cells injury.

The aim of this study was to reveal the mechanism of cold stimulation (CS)-bronchial epithelial cells (BECs) derived exosomes (CS-BECs-exo) aggravated sepsis induced acute lung injury (SALI). CS-BECs-exo were separated by differential centrifugation and were characterized. Proteomics, immunoprecipitation, and RAGE knockout (RAGE) mice were used to investigate the mechanism of CS-BECs-exo aggravated SALI. The results of transmission electron microscope (TEM) showed that CS-BECs-exo showed a double-layer membrane structure like a saucer. Nanoparticle tracking analysis (NTA) particle size analysis showed that the average particle size of CS-BECs-exo was 123.6 nm. The results of proteomics showed that the expression level HMGB1 was significantly increased in CS-BECs-exo compared with BECs-exo. CS-BECs-exo significantly increased oxidative stress and inflammatory reaction of SALI. In addition, CS-BECs-exo significantly increased RAGE and decreased the levels of Nrf-2 and OH-1. RAGE knockout (RAGE KO) and silence of RAGE (RAGE siRNA) significantly canceled the effects of CS-BECs-exo on SALI. HMGB1 knockout (HMGB1) and silence of HMGB1 also significantly (HMGB1 siRNA) canceled the effects of CS-BECs-exo on SALI. In conclusion, CS-BECs-exo aggravated ALI in sepsis via HMGB1/RAGE/Nrf-2/OH-1 signal pathway.

Silencing PADI-2 induces antitumor effects by downregulating NF-κB, Nrf2/HO-1 and AKT1 in A549 lung cancer cells.

This study aimed to investigate the tumorigenic role and regulatory pathways of peptidyl arginine deiminase 2 (PAD-2) in A549 lung cancer cells following treatment with small interfering RNA (PADI-2 siRNA) or the pharmacological pan-PAD inhibitor BB-Cl amidine. A549 lung cancer cells were treated with PADI-2 siRNA to knock down PADI-2 expression or with BB-Cl amidine to inhibit PAD2 activity. The effects on cell proliferation, migration, invasion, and cell cycle phases were assessed. Additionally, the expression levels of nuclear factor erythroid 2 p45-related factor 2 (Nrf2), heme oxygenase-1 (HO-1), AKT serine/threonine kinase 1 (AKT), nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), interleukin 6 (IL6), and p53 were analyzed to elucidate the underlying mechanisms involved. The manipulation of PAD-2 expression or activity significantly influenced tumor cell behavior. Knockdown of PADI-2 in A549 cells reduced cell proliferation by inhibiting the S and G2 phases and decreasing cell migration and invasion. Inhibition of PADI-2 expression also suppressed the protein levels of Nrf2 and HO-1 via suppression of the AKT/NF-κB pathway. Furthermore, this inhibition enhanced the senescence-associated secretory phenotype (SASP) through the regulation of IL6 and p53, resulted in significant upregulation of SASP factors mainly, p21, Lamin B1 and HMGB1. Downregulation of PADI-2 attenuated the proliferation, migration, and invasion of A549 lung cancer cells by modulating the Nrf2/HO-1/AKT signaling pathway. It also increased senescence in A549 lung cancer cells via IL6 and p53 key regulators. These findings highlight the potential of PADI-2 as a therapeutic target in lung cancer treatment.

A Mechanism Study on the Antioxidant Pathway of Keap1-Nrf2-ARE Inhibiting Ferroptosis in Dopaminergic Neurons.

The pathology of Parkinson's disease (PD) indicates that iron deposition exists in dopaminergic neurons, which may be related to the death of cellular lipid iron peroxide. The extracellular autophagy adaptor SQSTM1(p62) of dopamine (DA) neurons can activate the intracellular Keap1-Nrf2-ARE signaling pathway to inhibit ferroptosis, which has a protective effect on DA neurons. The objective of this study was to investigate the protective mechanism of the Keap1- Nrf2-ARE antioxidant pathway against iron death in dopaminergic neurons. The experiment was divided into a control group (Control group), 1-methyl-4- phenylpyridiniumion control group (MPP+ Control group), p62 overexpression group (MPP+OVp62), and p62 overexpression no-load group (MPP+ OV-P62-NC). The inhibitors brusatol and ZnPP inhibited the activation of NF-E2-related factor 2(Nrf2) and Heme oxygenase-1(HO-1), respectively, and were divided into brusatol group (MPP+OV-p62+brusatol) and ZnPP group (MPP+OV-p62+ZnPP). RT-qPCR was used to detect transfection efficiency, and Cell Counting Kit-8 (CCK8) was used to detect cell activity. FerroOrange, 2,7-Dichlorodihydrofluorescein diacetate (DCFH-DA), and Liperfluo probes were used to detect intracellular iron, reactive oxygen species (ROS), and lipid peroxidation (LPO) levels. Western Blotting detected the levels of Nrf2, HO-1, Kelch-like ECH-associated protein1 (Keap1), and their downstream Glutathione peroxidase 4(GPX4) and Acyl-CoA synthetase long-chain family member 4(ACSL4). The levels of LGlutathione (GSH) and Malondialdehyde (MDA) were detected by GSH and MDA kits, and the activation of Keap1-Nrf2-ARE pathway was verified at the cellular level to have an antioxidant protective effect on iron death in dopaminergic neurons. (1) The results of RT-qPCR showed that compared with the control group, the expression of the p62 gene was significantly increased in the MPP+OV-p62 groups (P = 0.039), and the p62 gene was significantly increased in the brusatol and ZnPP groups, indicating successful transfection (P =0.002; P=0.008). (2) The immunofluorescence probe flow results showed that compared to the normal control group, the contents of three kinds of probes in MPP+ model group were significantly increased (P =0.001; P ＜0.001; P＜0.001), and the contents of three kinds of probes in MPP+OV-p62 group were decreased compared to the MPP+ model group (P =0.004). The results indicated that the levels of iron, ROS, and LPO were increased in the MPP+ group and decreased in the MPP+OV-p62 group. (3) Compared with the control group, the expressions of Nrf2, HO-1, and GPX4 in the MPP+OV-p62 group were increased (P =0.007; P =0.004; P=0.010), and the expressions of Keap1 and ACSL4 in MPP+p62 overexpression group were decreased (P =0.017; P =0.005). Compared with the MPP+ control group, Nrf2 and GPX4 were increased in the MPP+OV-p62 group, and ACSL4 was decreased in the MPP+OVp62 group (P =0.041; P ＜0.001; P ＜0.001). The results of the GSH and MDA kit showed that compared with the normal control group, the content of GSH in the MPP+ control group was decreased (P < 0.01), and the content of MDA was increased (P < 0.01). Compared with the MPP+ model group, GSH content was increased (P = 0.003), and MDA content was decreased in the MPP+OV-p62 group (P < 0.001). Nrf2, HO-1, and GPX4 increased in the MPP+p62 overexpression group but decreased in the brusatol group and ZnPP group (P < 0.001). Based on the transfection of P62 plasmid, it was found that P62 plasmid can inhibit the lipid peroxidation of iron death in dopaminergic nerve cells by activating the Nrf2 signaling pathway, thus playing a protective role in dopaminergic nerve cells.

In vivo, in vitro, and in silico approaches in the detailed study of di-butyl phthalate (DBP), a plasticizer-induced lung fibrosis via Nrf-2/Keap-1/HO-1 pathway and its regulation.

The alveolar epithelium is a crucial barrier against external threats, yet it becomes a key player in initiating pulmonary fibrosis when compromised. Despite its importance, the intricate relationship between, DBP exposure and alveolar epithelial cell injury ensuing pro-fibrotic effects remains poorly understood. Phthalates, ubiquitous in nature, pose a significant risk to lung health upon inhalation, acting as immune triggers that cause airway inflammation and epithelial damage. We aimed to investigate the impact of intranasal administration ofDi-butyl Phthalate (DBP) inhalation, and its probable effects on normal and asthmatic lungs. DBP was administered via intranasal route in normal and OVA-induced asthmatic mice. DBP exposure enhanced oxidative stress and inflammatory parameters, leading to exacerbated asthmatic response and oxidative lung damage. Enhanced accumulation of immune cells, bronchial thickening, and collagen deposition was noted in histopathological investigations of DBP-exposed lung sections. Curcumin, a plant-derived molecule, significantly mitigated DBP-exposed asthma exacerbations by suppressing NF-κB expression and enhancing NRF2 levels via the Nrf-2/Keap-1/HO-1 signaling pathway. FACS analysis revealed increased CD11b+ cells (32%) in asthmatic mice which were reduced in the curcumin pre-treatment group (10.5%). Enhanced epithelial to mesenchymal transition (EMT) was noted in mice lungs and A549 cells where E-cadherin expression was reduced as compared to Vimentin, and α-SMA. Apart from aggravated airway inflammation, DBP exposure damages healthy lungs also. MMP-9/TIMP-1 ratios and collagen-1 levels were restored which were enhanced after DBP exposure. Moreover, antioxidant enzyme levels such as NQO-1, HO-1, and Catalase were significantly enhanced (p<0.01) and comparable to dexamethasone, a conventional corticosteroid. Notably, both dexamethasone and curcumin treatments effectively regulated the stimulation and accumulation of Nrf-2 in the nucleus, promoting antioxidant production and offering potential therapeutic benefits in mitigating pulmonary fibrosis. OVA and DBP alone caused DNA damage in the lung cells where increasedpercentage of damaged DNA movement in thetail, tail length, tail moment, and olive tail moment indicated severe damage in theDBP and OVA combined exposure strategies. Dexamethasone and Curcumin treatments reduced theextent of the DNA damage indicating anti-inflammatory and ant-oxidative potentials. Moreover, in silico studies are supportive of therapeutic potential of Curcumin and Dexamethasone in DBP-induced lung inflammation and fibrosis.

Improving anti-oxidant stress treatment of subarachnoid hemorrhage through self-assembled nanoparticles of oleanolic acid

Subarachnoid hemorrhage (SAH) is a life-threatening acute hemorrhagic cerebrovascular disease, with early brain injury (EBI) being the main cause of high mortality and severe neurological dysfunction. Oxidative stress plays a crucial role in the pathogenesis of EBI. In this study, we synthesized antioxidant stress nanoparticles based on self-assembled oleanolic acid (OA) using the solvent volatilization method. X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR), and transmission electron microscopy (TEM) techniques were employed to analyze and understand the self-assembly mechanism of oleic acid nanoparticles (OA NPs). The TUNEL assay, Nissl staining, and brain water content measurements were conducted to investigate the impact of OA NPs on cortical neuronal injury. Additionally, Western blot analysis was performed to investigate the antioxidant stress mechanism of OA NPs. The result showed that OA NPs exhibited a spherical structure with an average diameter of 168 nm. The application of OA NPs in SAH has been found to contribute to the reduction of keap1 protein levels and an increase in the nuclear level of Nrf2. As a result, the transcription of antioxidant stress proteins, including HO1 and NQO1, is triggered. The activation of the antioxidant stress pathway by OA NPs ultimately leads to a decrease in neuron damage and an improvement in neurological dysfunction. In conclusion, we successfully designed and synthesized OA NPs that can efficiently target the site of SAH. These nanoparticles have demonstrated their potential as antioxidants for the treatment of SAH, offering significant clinical applications.