Nrf2-related Genes Research Articles

The comparative anti-oxidant and anti-inflammatory efficacy of postbiotics and probiotics through Nrf-2 and NF-kB pathways in DSS-induced colitis model

IBD is a disorder which could be caused by oxidative stress. This investigation aims to determine if probiotics and postbiotics can control oxidative stress and inflammation and compare the effectiveness of these two probiotic and postbiotic mixtures of substances. 88 strains of Lactobacillus and Bifidobacterium were tested for antioxidant activity. Male wild-type C57BL/6 mice were divided into four experimental groups, namely high fat diet (HFD) + PBS, HFD + DSS, HFD + DSS + 109 cfu/ml of probiotics, and HFD + DSS + 109 cfu/ml of postbiotics. The phenotypical indices and pathological scores were assessed. The expression of genes related to NF-kB and Nrf2 signaling pathways and enzymes associated with oxidant/anti-oxidant activities, and proinflammatory/inflammatory cytokines were assessed. In contrast to the groups exposed to DSS, mice treated with probiotics mixture and postbiotics mixture alongside DSS displayed alleviation of DSS-induced adverse effects on phenotypical characteristics, as well as molecular indices such as the Nrf2 and NF-kB related genes, with a greater emphasis on the postbiotics component. In accordance with the findings of the present investigation, it can be inferred that even in using a high-fat dietary regimen as an inducer of oxidative stress, the emergence of inflammation can be effectively addressed through the utilization of probiotics and, more specifically, postbiotics.

Upregulation of CIRP by its agonist prevents the development of heart failure in myocardial infarction rats

BackgroundDownregulated expression of cold-inducible RNA binding protein (CIRP), a stress-response protein, has been demonstrated in the hearts of patients with heart failure (HF). However, whether CIRP plays a critical role in the pathogenesis of HF remains unknown. Zr17-2 is a recently identified CIRP agonist, which can enhance the expression of CIRP in hearts. Herein, we evaluated the effects of zr17-2 on the development of HF in a rat model of myocardial infarction (MI).MethodsMale SD rats were pretreated with CIRP agonist zr17-2 or vehicle saline for 6 consecutive days, followed by MI induction. 1-week post-MI, cardiac function, and structural and molecular changes were determined by echocardiography and molecular biology methods.ResultsExcitingly, we found that pretreatment with zr17-2 significantly attenuated MI-induced cardiac dysfunction and dilation, coupled with reduced infarction size and cardiac remodeling. In addition, increased inflammatory response in the peri-infarcted heart including macrophage infiltration and the expression of inflammatory genes were all significantly decreased by zr17-2 pretreatment, suggesting an anti-inflammatory effect of zr17-2. Moreover, zr17-2 pretreatment also upregulated the antioxidant genes (e.g. NQO-1, Nrf2, and HO-1) level in the hearts. In isolated cultured cardiomyocytes, pretreatment with zr17-2 markedly attenuated cell injury and apoptosis induced by oxidative injury, along with elevation of Nrf2-related antioxidant genes and CIRP. However, silencing CIRP abolished zr17-2’s antioxidant effects against oxidative injury, confirming that zr17-2’s role is dependent on CIRP.ConclusionCollectively, our study suggests CIRP plays a crucial role in the development of HF and a beneficial effect of CIRP agonist in preventing MI-induced HF, possibly via anti-inflammatory and anti-oxidant pathways.

Lactational transfer of sulforaphane-N-acetylcysteine in vivo and in human breast milk

Sulforaphane (SFN) is a bioactive phytonutrient found in cruciferous vegetables. There is a lack of detailed information on the lactational transfer of SFN and SFN metabolites, and potential pharmacological effects on breastfeeding infants. We carried out two maternal supplementation studies in a mouse model, wherein lactating dams received either vehicle, 300 or 600 ppm SFN from postnatal day (PND) 1 to 5, or in a second experiment, vehicle or 600 ppm SFN from PND 1 to 14. The parent compound was only detectable in milk and plasma from dams receiving 600 ppm SFN for five days. The predominant metabolite SFN-N-acetylcysteine (SFN-NAC) was readily detected in milk from dams receiving 300 and 600 ppm SFN for five days or 600 ppm for 14 days. Maternal SFN-NAC plasma levels were elevated in both 600 ppm groups. Maternal hepatic and pulmonary expression of NRF2-related genes, Nqo1, Gsta2, Gstm1, and Gstp1, were significantly increased, generally following a dose-response; however, offspring induction varied. PND5 neonates in the 600-ppm group exhibited significantly elevated expression of Nqo1, Gsta2, and Gstp1 in liver, and Gstm1 and Gstp1 in lung. Findings support maternal dietary supplementation with SFN induces NRF2-related gene expression in neonates via lactational transfer of SFN-NAC. However, NQO1 enzyme activity was not significantly elevated, highlighting the need to optimize dosing strategy. Additionally, in a pilot investigation of lactating women consuming a typical diet, without any purified SFN supplementation, 7 out of 8 breast milk samples showed SFN-NAC above the limit of quantification (LOQ). Notably, the one sample below the LOQ was collected from the only participant who reported no consumption of cruciferous vegetables in the past 24 h. The parent compound was not detected in any of the human breast milk samples. Overall, these data indicate lactational transfer of SFN-NAC at dietary relevant levels. Future studies are needed to evaluate pharmacokinetics and pharmacodynamics of lactational transfer for potential preventive or therapeutic effects in breastfeeding children.

Biogenic Selenium Nanoparticles Synthesized with Alginate Oligosaccharides Alleviate Heat Stress-Induced Oxidative Damage to Organs in Broilers through Activating Nrf2-Mediated Anti-Oxidation and Anti-Ferroptosis Pathways.

Selenium (Se) is an essential trace element for maintaining health due to its ideal antioxidant properties. We previously prepared a new type of biogenic selenium nanoparticles based on alginate oligosaccharides (SeNPs-AOS), and this study aimed to investigate the protective effects of SeNPs-AOS (Se particle size = 80 nm, Se content = 8%) on organ health in broilers challenged with HS. A total of 192 21-day-old Arbor Acres broilers were randomly divided into four groups according to a 2 × 2 experimental design, including a thermoneutral zone group (TN, raised under 23 ± 1.5 °C); TN + SeNPs-AOS group (TN group supplemented 5 mg/kg SeNPS-AOS); HS group (HS, raised under 33 ± 2 °C for 10 h/day); and HS + SeNPs-AOS group (HS group supplemented 5 mg/kg SeNPS-AOS). There were six replicates in each group (eight broilers per replicate). The results showed that SeNPs-AOS improved the splenic histomorphology, enhanced the activity of catalase (CAT) and glutathione peroxidase (GSH-Px) of the spleen, as well as upregulating the splenic mRNA expression of antioxidant-related genes in broilers under HS. In addition, SeNPs-AOS reversed the pathological changes in bursa caused by HS increased the activity of GST, GSH-Px, and CAT and upregulated the mRNA expression of Nrf2 and antioxidant-related genes in the bursa of heat-stressed broilers. In addition, dietary SeNPs-AOS improved the hepatic damage, increased the activity of GSH-Px in the liver, and upregulated the mRNA expression of antioxidant-related genes while downregulating the Keap1 gene expression of the liver in broilers during HS. Moreover, dietary SeNPs-AOS upregulated the anti-ferroptosis-related genes expression of liver in broilers under HS. In conclusion, dietary SeNPs-AOS could relieve HS-induced oxidative damage to the spleen, bursa of Fabricius and liver in broilers by upregulating the Nrf2-mediated antioxidant gene expression and SeNPs-AOS could also upregulate the expression of hepatic Nrf2-related anti-ferroptosis genes in heat-stressed broilers. These findings are beneficial for the development of new nano-antioxidants in broilers.

Penthorum chinense Prush extract alleviates aflatoxin B1-induced toxicity, oxidative stress and apoptosis via mediating Nrf2 signaling pathway in the Bursa of Fabricius of broilers

Aflatoxin B1 (AFB1) is the primary mycotoxin that is responsible for the severe issues plaguing poultry farming. The study was aimed to explore the relevant pathways connected with immunity (inflammation, oxidative stress and apoptosis) in an AFB1-challenged chicken by using Penthorum chinense Prush extract (PCPE) in Bursa of Fabricius (BF) of broilers. A total one hundred and eighty day-old broilers were divided into six groups: Control, AFB1 (3 mg/kg feed), Yin-Chen-Hao Tang extract (YCHT) (10 ml/kg feed), and PCPE groups (low 1 g/kg, medium 2 g/kg, and high 3 g/kg PCPE/kg feed) respectively. The results showed that AFB1-challenged birds showed significant decrease in growth, BF weight index, serum antioxidant biomarkers and histopathological changes in BF tissues. The mRNA analysis showed that AFB1 upregulated the apoptosis associated genes (caspase-3, caspase-9, Bak, Bax and p53) and downregulated BCL-2. Additionally, AFB1 downregulated expression level of Nuclear Factor EF-2 (Nrf2) related genes (Nrf2, HO-1, NQO1 and GCLC) in the BF of broilers. The PCPE treatment showed positive impact on final weight gain, bursal index, and reversing of pathological changes in the BF of AFB1-challanged broilers. PCPE ameliorated oxidative stress generated by AFB1, as an increase in antioxidant enzyme activities, alleviated histopathological changes in BF, enhanced the Nrf2 expression levels and lowered the apoptosis gene expressions as compared to AFB1. The findings revealed that PCPE activated the Nrf2 pathway, antioxidant defense system and modulated the apoptosis in the BF of broiler chicken.

Irisin drives macrophage anti-inflammatory differentiation via JAK2-STAT6-dependent activation of PPARγ and Nrf2 signaling.

Irisin is an exercise-induced myokine that alleviates inflammation and obesity. The induction of anti-inflammatory (M2) macrophage is facilitated for treatment of sepsis and associated lung damage. However, whether irisin drives macrophage M2 polarization remains unclear. Here, we found that irisin induced-macrophage anti-inflammatory differentiation in vivo using an LPS-induced septic mice model and in vitro using RAW64.7cells and bone marrow-derived macrophages (BMDMs). Irisin also promoted the expression, phosphorylation, and nuclear translocation of peroxisome proliferator-activated receptor gamma (PPAR-γ) and nuclear factor-erythroid 2-related factor 2 (Nrf2). Inhibition or knockdown of PPAR-γ and Nrf2 abolished irisin-induced accumulation of M2 macrophage markers, such as interleukin (IL)-10 and Arginase 1. Furthermore, dual-luciferase reporter and chromatin immunoprecipitation-quantitative PCR (ChIP-qPCR) assays confirmed that STAT6 boosts PPAR-γ and Nrf2 transcription by binding to their DNA promoters in irisin-stimulated macrophages. In contrast, STAT6 shRNA blocked the irisin-induced activation of Pparγ, Nrf2, and related downstream genes. Moreover, the interaction of irisin with its ligand integrin αVβ5 remarkably promoted Janus kinase 2 (JAK2) phosphorylation, while inhibition or knockdown of integrin αVβ5 and JAK2 attenuated the activation of STAT6, PPAR-γ, and Nrf2 signaling. Interestingly, co-immunoprecipitation (Co-IP) assay also revealed that the binding between JAK2 and integrin αVβ5 is critical for irisin-induced macrophage anti-inflammatory differentiation by enhancing the activation of the JAK2-STAT6 pathway. In conclusion, irisin boosted M2 macrophage differentiation by inducing JAK2-STAT6-dependent transcriptional activation of the PPAR-γ-related anti-inflammatory system and Nrf2-related antioxidant genes. The findings of this study suggest that the administration of irisin is a novel and promising therapeutic strategy for infectious and inflammatory diseases.

Nrf2 Overexpression Increases Risk of Venetoclax Resistance in Acute Myeloid Leukemia By Promoting Glycolysis

Backgroud and objective: Recurrence and drug resistance is still to be the primary barrier to acute myeloid leukemia (AML). Glycolysis is elevated in the tumor environment, which contributes to reduce the sensitivity of leukemia cells to chemotherapeuticsy. Venetoclax is a BCL-2 inhibitor that effectively improves clinical outcomes in the poor-prognosis category of AML patients unfit for intensive chemotherapy. Though the majority of patients respond to venetoclax-based treatment, duration of response remain inadequate. Nuclear factor-erythroid 2-related factor 2 (Nrf2) is a pleiotropic transcription factor, which plays an important role in regulating the metabolic homeostasis. The mechanism by which NRF2 mediated venetoclax-resistance of AML through promoting the metabolic homeostasis are poorly understood. Methods: The possible glycolysis related genes regulating by NRF2 were confirmed with Gene Transcription Regulation Database (GTRD) and Knock TF. Venetoclax-resistant MV4-11 cells(MV4-11/VEN cells) were newly established through continuous exposure of MV4-11 cells to venetoclax. Drug sensitivity were compared between MV4-11 and MV4-11/VEN cells by Cell Counting Kit-8 (CCK8). In addition, flow cytometry was used to observe the apoptosis process. Glucose and lactate kits were utilized to detect glucose consumption and lactate levels. The expression of NRF2 and glycolysis related genes were examined by quantitative reverse transcription-PCR (RT-PCR) and Western blot. Finally, we transfected AML cell lines by constructing LV-NRF2 lentiviral vector, and integrating siRNA of glycolysis related gene separately to detect the above index changes in leukemia cells. Results: MV4-11/VEN cells displayed more than 10-fold resistance to venetoclax compared with MV4-11 cells (P<0.05). MV4-11/VEN cells also exhibited a higher glycolysis rate and increased expression level of NRF2. These effectsIt could be reversed by knock down of NRF2 in MV4-11/VEN cells. Further, NRF2 overexpression could protect the AML cells from apoptosis induced by venetoclax in vitro and increased the rate of drug resistance associated with glycolysis. Bioinformatics analysis suggested that glycolytic genes that may be transcriptionally regulated by NRF2 include G6PD, PKFP, SOD1, GCLC, and GOT1. Among these candidates, the expression of SOD1 was significantly decreased after knockdown of NRF2. Furthermore, Nrf2 overexpression induced the expression of SOD1, which enhanced the rate of glycolysis. Inhibition of SOD1 could reverse the changes in leukemia cells induced by NRF2. Conclusion: As an important component of the metabolic homeostasis, glycolysis are closely related to the venetoclax resistance of AML. NRF2 could be a key component mediating the interaction between glycolysis and leukemia cells of AML This results might provide a new entry point for deepening the mechanism of AML progression and provides a new strategy for clinical practice.

Garlic-Derived Metabolites Exert Antioxidant Activity, Modulate Gut Microbiota Composition and Limit Citrobacter rodentium Infection in Mice.

The garlic-derived compounds propyl propane thiosulfinate (PTS) and propyl propane thiosulfonate (PTSO) are metabolites with putative health benefits against intestinal inflammation that may be related to their antioxidant activity. However, the underlying mechanisms remain unclear, and whether PTS-PTSO can promote gut health by altering the microbiota and exert protection against enteric pathogens needs further investigation. Here, we explored the antioxidant activity of PTS-PTSO in murine macrophages in vitro, and in an in vivo model of bacterial infection with the bacterial pathogen Citrobacter rodentium. PTS-PTSO attenuated reactive oxygen species in lipopolysaccharide-stimulated macrophages in a nuclear factor erythroid factor 2-related factor 2 (Nrf2)-dependent manner, decreased nitric oxide levels both in macrophages in vitro and in the sera of mice fed PTS-PTSO, and had putatively beneficial effects on the commensal gut microbiota. Importantly, PTS-PTSO decreased faecal C. rodentium counts, concomitant with upregulation of Nrf2-related genes in colon tissue. Thus, PTS-PTSO mediates Nrf2-mediated antioxidant activity and modulates gut microbiota, which may protect the host against C. rodentium colonization. Our results provide further insight into how PTS-PTSO and related bioactive dietary compounds may reduce enteric infections.

The Gut Microbial Metabolite Pyrogallol Is a More Potent Inducer of Nrf2-Associated Gene Expression Than Its Parent Compound Green Tea (-)-Epigallocatechin Gallate.

(-)-Epigallocatechin gallate (EGCG) has been associated with multiple beneficial effects. However, EGCG is known to be degraded by the gut microbiota. The present study investigated the hypothesis that microbial metabolism would create major catechol-moiety-containing microbial metabolites with different ability from EGCG to induce nuclear factor-erythroid 2-related factor 2 (Nrf2)-mediated gene expression. A reporter gene bioassay, label-free quantitative proteomics and reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) were combined to investigate the regulation of Nrf2-related gene expression after exposure of U2OS reporter gene or Hepa1c1c7 cells in vitro to EGCG or to its major microbial catechol-moiety-containing metabolites: (-)-epigallocatechin (EGC), gallic acid (GA) and pyrogallol (PG). Results show that PG was a more potent inducer of Nrf2-mediated gene expression than EGCG, with a 5% benchmark dose (BMD5) of 0.35 µM as compared to 2.45 µM for EGCG in the reporter gene assay. EGC and GA were unable to induce Nrf2-mediated gene expression up to the highest concentration tested (75 µM). Bioinformatical analysis of the proteomics data indicated that Nrf2 induction by PG relates to glutathione metabolism, drug and/or xenobiotics metabolism and the pentose phosphate pathway. Taken together, our findings demonstrate that the microbial metabolite PG is a more potent inducer of Nrf2-associated gene expression than its parent compound EGCG.

Heat stress in broilers of liver injury effects of heat stress on oxidative stress and autophagy in liver of broilers

This study aimed to investigate the effect of chronic heat stress on oxidative stress in liver of broilers. In our study, chickens were randomly allocated to control 1 group (control 7 d), heat stress 1 group (HS1, 7 d), control 2 group (control 14 d) and heat stress 2 group (HS2, 14 d), with 30 replicates in each group. Broilers in heat stress groups exposed 8 h/day heat stress (35 ± 2°C) for 7 or 14 consecutive days, and the rest of time per day were kept at 23 ± 2℃ the same as control group broilers. Growth performance and the liver tissues were collected for histological observation and detection of organ index and liver redox status. The serum indicators (alanine aminotransferase [ALT] and aspartate aminotransferase [AST]) related to liver injury were determined. Moreover, Nrf2-related genes and protein expression levels in liver were measured. The results showed that in heat stress group broilers the body weight gain, feed conversion ratio, liver weight, and liver index were decreased, inflammatory cells infiltration in liver, and serum AST level was enhanced, compared with control group broilers. Moreover, the hepatic malondialdehyde (MDA) and superoxide dismutase (SOD) level were increased after 1 wk of heat stress. Nrf2, Sqstm1/p62, HO-1, and NQO1 mRNA expressions in the liver of broilers were decreased by heat stress. P62 and p-p62 protein expressions were significantly up-regulated, but Nrf2 and keap1 protein level was decreased in heat stress group broilers as compared to control group. The mRNA expression levels of Beclin1, LC3-I, LC3-II were down-regulated significantly with heat stress for 1 wk. The mRNA expression level of mTOR up-regulated after 2 wk of heat stress. In conclusion, heat stress induced liver injury of broilers by down-regulating Nrf2-keap1 signaling pathway and autophagy.

(±)-5-bromo-2-(5-fluoro-1-hydroxyamyl) Benzoate Protects Against Oxidative Stress Injury in PC12 Cells Exposed to H2O2 Through Activation of Nrf2 Pathway.

Background: Oxidative stress is associated with the pathogenesis of ischemic stroke (±)-5-bromo-2-(5-fluoro-1-hydroxyamyl) benzoate (BFB) is a novel compound modified by dl-3-n-butylphthalide (NBP). Here, we hypothesized that BFB may protect the PC12 cells against H2O2-induced oxidative stress injury through activation of the Nrf2 pathway. Methods: We measured the cell viability and levels of lactate dehydrogenase (LDH), malondialdehyde (MDA), glutathione (GSH), and reactive oxygen species (ROS) to determine the construction of the H2O2-induced models of oxidative stress in PC12 cells. Additionally, apoptotic cell death, mitochondrial membrane potential, and cellular morphology were examined to determine the effect of BFB on oxidative stress injury in H2O2-treated PC12 cells. The expression levels of Nrf2-related and autophagy-related genes and proteins were detected using real time quantative PCR (RT-qPCR), Western Blot, and immunofluorescence analyses. Results: Our study showed that BFB treatment reduced the elevated levels of MDA, LDH, and ROS, and decreased cell viability and GSH in H2O2-treated PC12 cells. We also observed the elevated expression of Nrf2 pathway-related factors and intranuclear transitions and found that Nrf2 inhibitors (ML385) could block the protective effect of BFB. The inhibitory effect of BFB on oxidative stress may be partially regulated by Nrf2 activation, and the initiation and induction of autophagy. Conclusion: BFB inhibited H2O2-induced oxidative stress injury in PC12 cells by activating the Nrf2 pathway, initiating and inducing autophagy, suggesting that BFB may be a promising therapeutic agent in treating neurological disorders like cerebral ischemia.

Anti‐depressant lithium compounds reliefs cardiorespiratory disturbances and improves cardiac function in experimental non‐ischemic heart failure

Non‐ischemic heart failure (HF) is a complex medical problem with a high incidence among the older population. While several efforts have been made to improve HF outcomes there is an urgent need for new strategies to reduce mortality rates. Sympathoexcitation and disordered breathing are both considered hallmarks of HF and are closely related to disease progression. Lithium salts has been largely used as anti‐depressants; however, lithium compounds also showed several pleiotropic effects some of them linked to the origin of cardiorespiratory disorders in HF (i.e. oxidative stress). Accordingly, using drug repurposing strategy we aimed to define whether lithium treatment improve cardiorespiratory function in experimental HF. Male Sprague‐Dawley rats underwent volume overload to induce non‐ischemic HF. Four weeks after HF induction, an osmotic minipump containing lithium chloride (Li+, 2 meq/Kg/day) was subcutaneously implanted for drug delivery. Cardiac autonomic and baroreflex function, ventilatory chemoreflex drive and disordered breathing were evaluated after 4 weeks of Li+ treatment in freely moving rats. In addition, cardiac function was determine using PV‐loops. Compared to Sham rats, HF animals displayed autonomic imbalance, reduced baroreflex sensitivity, higher incidence of cardiac arrhythmias, enhanced central chemoreflex and breathing disorders. Cardiac diastolic dysfunction was evident in HF animals compared to Sham animals. HF rats that underwent Li+ treatment showed significant (p&lt;0.05) improvements in cardiac autonomic function as evidenced by reduced low‐to‐high frequency ratio of heart rate variability (LF/HF: 2.8 ± 0.3 vs. 0.9 ± 0.4; HF+Veh vs. HF+Li+), baroreflex sensitivity (BRS: 2.0 ± 0.5 vs. 5.8 ± 1.0 ms/mmHg; HF+Veh vs. HF+Li+) and reduced cardiac arrhythmogenesis (PVC: 220 ± 50 vs. 30 ± 10 events/h; HF+Veh vs. HF+Li+). Furthermore, Li+ treatment in HF completely normalized central chemoreflex drive. Indeed, hypercapnic ventilatory responses obtained in HF+Li+ rats were undistinguishable from the ones obtained in Sham rats (HCVR: 1.7 ± 0.4 vs. 2.0 ± 0.4 DVe/FiCO2; HF+Veh vs. HF+Li+). Also, breathing disorders (i.e. apneas/hypopneas) were markedly reduced by Li+ treatment in HF (AHI: 8.5 ± 1.0 vs. 3.7 ± 0.8; HF+Veh vs. HF+Li+). Lastly, HF+Li+ treated rats displayed marked improvements in cardiac diastolic function (~2.5‐fold reduction in EDPVR) compared to HF+Veh treated rats. We found that the beneficial effects of Li+were partially associated with upregulation of the Nrf2 pathway and related antioxidant genes (i.e. SOD2), reductions in tissue oxidative stress and decreases in the expression of pro‐inflammatory cytokines (i.e. TNF‐a) in HF rats. Our results support the notion that lithium compounds may offer new avenues for the treatment of cardiorespiratory disorders and cardiac diastolic dysfunction in non‐ischemic HF.

Antagonistic effects of selenium on lead-induced oxidative stress and apoptosis of Leydig cells in sheep

The objective of this study was to investigate the antagonistic effects of selenium (Se) on lead (Pb)-induced oxidative stress and apoptosis of sheep Leydig cells and its underlying mechanism. Leydig cells collected from 8-month-old sheep were treated with Pb (40 μmol/L) and/or Se (2 μmol/L), respectively. CCK-8 assay was used to detect cell proliferation and apoptosis after cultured for 48 h. The abundances of pro-apoptosis (BAX, CASPASE 3 and CASPASE 8) and NRF2-related (NRF2, HO-1, NQO1 and γ-GCS) genes were detected by real-time PCR and western blot analysis, respectively.The results showed that the highest cell viability was observed in the Se group. Compared with the control group, Pb treatment led to the higher ROS level and greater abundances of BAX, CASPASE 3 and CASPASE 8 mRNA transcripts. Treatment with Pb + Se resulted in an increased (P < 0.05) abundances of NRF2, HO-1, NQO1 and γ-GCS mRNA transcripts and proteins. Compared with the Pb group, the Se + Pb treatment dramatically decreased (P < 0.05) the ROS level and relative abundances of pro-apoptosis genes. The greater (P < 0.05) abundances of pro-apoptosis and NRF2-related mRNA transcripts and proteins were also obtained in the Se + Pb group. The abundances of BAX, CASPASE 3 and CASPASE 8 genes in the SeML385 group were greater (P < 0.05) than in the Se group. Compared with the corresponding groups without ML385, treatment with ML385 decreased (P < 0.05) cell viability and the relative abundances of pro-apoptosis and NRF2-related genes.These results indicate that Pb-induced oxidative stress can inhibit the viability of Leydig cells by modulating pro-apoptosis gene expression. NRF2 pathway could be involved in the antagonistic effect of Se on Pb-induced apoptosis of Leydig cells in sheep. This study is expected to provide some experimental evidences for Se treatment of Pb-induced reproductive disorders.

Activation of the Nrf2 Pathway Prevents Mitochondrial Dysfunction Induced by Caspase-3 Cleaved Tau: Implications for Alzheimer's Disease.

Alzheimer’s disease (AD) is characterized by memory and cognitive impairment, accompanied by the accumulation of extracellular deposits of amyloid β-peptide (Aβ) and the presence of neurofibrillary tangles (NFTs) composed of pathological forms of tau protein. Mitochondrial dysfunction and oxidative stress are also critical elements for AD development. We previously showed that the presence of caspase-3 cleaved tau, a relevant pathological form of tau in AD, induced mitochondrial dysfunction and oxidative damage in different neuronal models. Recent studies demonstrated that the nuclear factor (erythroid-derived 2)-like 2 (Nrf2) plays a significant role in the antioxidant response promoting neuroprotection. Here, we studied the effects of Nrf2 activation using sulforaphane (SFN) against mitochondrial injury induced by caspase-3 cleaved tau. We used immortalized cortical neurons to evaluate mitochondrial bioenergetics and ROS levels in control and SFN-treated cells. Expression of caspase-3 cleaved tau induced mitochondrial fragmentation, depolarization, ATP loss, and increased ROS levels. Treatment with SFN for 24 h significantly prevented these mitochondrial abnormalities, and reduced ROS levels. Analysis of Western blots and rt-PCR studies showed that SFN treatment increased the expression of several Nrf2-related antioxidants genes in caspase-3 cleaved tau cells. These results indicate a potential role of the Nrf2 pathway in preventing mitochondrial dysfunction induced by pathological forms of tau in AD.

Genetic Variations on Redox Control in Cardiometabolic Diseases: The Role of Nrf2.

The transcription factor Nrf2 is a master regulator of multiple cytoprotective genes that maintain redox homeostasis and exert anti-inflammatory functions. The Nrf2-Keap1 signaling pathway is a paramount target of many cardioprotective strategies, because redox homeostasis is essential in cardiovascular health. Nrf2 gene variations, including single nucleotide polymorphisms (SNPs), are correlated with cardiometabolic diseases and drug responses. SNPs of Nrf2, KEAP1, and other related genes can impair the transcriptional activation or the activity of the resulting protein, exerting differential susceptibility to cardiometabolic disease progression and prevalence. Further understanding of the implications of Nrf2 polymorphisms on basic cellular processes involved in cardiometabolic diseases progression and prevalence will be helpful to establish more accurate protective strategies. This review provides insight into the association between the polymorphisms of Nrf2-related genes with cardiometabolic diseases. We also briefly describe that SNPs of Nrf2-related genes are potential modifiers of the pharmacokinetics that contribute to the inter-individual variability.

Esculetin alleviates murine lupus nephritis by inhibiting complement activation and enhancing Nrf2 signaling pathway

Ethnopharmacological relevanceEsculetin is a bioactive compound of medicinal herb Hydrangea paniculata, and has showed anti-oxidation and anti-inflammation bioactivities. Renal local oxidative stress and inflammation are import contributors for progression of lupus nephritis (LN). Aim of the studyIn the present study, the renal protective effect of esculetin against LN was evaluated using MRL/lpr mice. Materials and methodsMRL/lpr mice were orally administrated with esculetin (20 mg/kg and 40 mg/kg) from 10 to 20 weeks and then renal function and kidney pathology were analyzed. ResultsEsculetin significantly attenuated renal impairment in MRL/lpr mice by reducing blood urea nitrogen (BUN), serum creatinine (Scr) and albuminuria, and ameliorated the glomerular hypertrophy, tubular interstitial fibrosis and mononuclear cell infiltration into interstitium. mRNA microarray suggested that esculetin could significantly down-regulate complement cascade, inflammation and fibrosis pathway, and up-regulate Nrf2-related anti-oxidation genes. Most surprising finding in the current study was that esculetin could inhibit the complement activation both in classical and alternative pathway using in vitro hemolysis assay, further enzyme assay suggested that esculetin blocked the C3 convertase (C4b2a) to exert this inhibitory capability. Molecular docking predicted that esculetin had four conventional hydrogen bonds interacting with C4b2a, and CDOCKER energy is relatively lower. Luciferase reporter gene demonstrated that esculetin could activate Nrf2 signaling pathway, and further flow cytometry confirmed that anti-oxidation bioactivity of esculetin was dependent on Nrf2 activation. On the other hand, esculetin could inhibit NFκB nuclear translocation and TGFβ-smad3 profibrosis pathway. ConclusionEsculetin shows beneficial effect on LN progression, and it may be a good natural leading compound for design of chemical compounds to treat LN.

An Investigation into the Impact of a Glutaminase Inhibitor, Compound 968, on Nrf2 Signaling

Glutaminase is a critical enzyme that catalyzes the process of glutaminolysis for energy synthesis. Meanwhile, glutaminase also contributes to the pathological process of various diseases, such as cancer, neurodegenerative diseases, and inflammation. This leads to the discovery of glutaminase inhibitors for therapeutical uses. However, the mechanisms of the beneficial therapeutical effect of glutaminase inhibitors are still unclear. This pilot study aimed to determine the impact of a well-characterized glutaminase inhibitor, compound 968 (C968), on Nrf2 signaling. We performed molecular docking, luciferase assay, and quantitative PCR to determine the activation of Nrf2 and the expression of several Nrf2-related genes. These experiments found that C968 induced the Nrf2 activation and promoted the expression of Nrf2, heme oxygenase-1 (HO-1), and NAD(P)H Quinone Dehydrogenase-1 (NQO-1). All findings provide evidence that Nrf2 activation could be one of the mechanisms contributing to the therapeutical activity of C968, but more studies are warranted to further confirm this mechanism.

Keap1/Nrf2 Pathway Modifications In Muscle To Exercise Trained Compared To Sedentary Mice

Nrf2 is a protein/ transcription factor in the cytosol and bound to KEAP1. When released NRf2 can be degraded or translocated to the nucleus and bind to the antioxidant response element (ARE). Nrf2 binding to the ARE induces antioxidants to protect against oxidative stress. Chronic exercise training effects on KEAP1/Nrf2 proteins are not well understood. Studies reported Nrf2 increases in whole cell homogenates and/or nuclear concentration but have not included KEAP1 concentration. Therefore, the purpose of this study was to examine the changes that may occur in the KEAP1/Nrf2 proteins in response to exercise training in both the cytosol and the nucleus in muscle. PURPOSE: To examine the changes in KEAP1/Nrf2 proteins in response to exercise training in cytosol and nucleus in muscle. METHODS: Sedentary C57BL/6 mice (N = 9) were compared to endurance exercise trained mice (N = 14), (8 wks, 5 times/wk for 45-60 min). Trained mice were sacrificed 72 hours after the last exercise session and sedentary mice were sacrificed at the same time. Muscles were flash frozen in liquid nitrogen and later thawed, homogenized, and then underwent nuclear and cytosolic fractioning. Fractions were multiplex immunoblotted and labelled using fluorescent-conjugated primary antibodies (anti-: KEAP1, Nrf2, HO1, SOD1, GAPDH) (Santa Cruz Biotech). Membranes were analyzed using Typhoon scanner and normalized to GAPDH. Repeated measures ANOVAS were performed between groups, followed up with Tukey post-hoc pairwise comparisons. RESULTS: Nuclear Nrf2 showed a significant in the trained group compared to sedentary group across all muscles (69.5%, p = 0.04), with no difference in cytosol Nrf2. SOD1 increased in the cytosol fraction in trained (18%, p = 0.04) compared to sedentary group. KEAP1 content showed no change in the cytosol. HO1 showed no difference in concentration in the muscles. CONCLUSIONS: The results from this study showed that exercise training induced nuclear translocation of Nrf2 in the muscles tested with no change in KEAP1 concentration. SOD1 in the cytosol increased but HO1 did not suggesting selective increases in the downstream antioxidants. Further research is needed to understand Nrf2 nuclear translocation with endurance training. Future studies should examine Nrf2-DNA binding and determine mRNA of Nrf2-related genes.

Deficiency of Nrf2 exacerbates white matter damage and microglia/macrophage levels in a mouse model of vascular cognitive impairment

BackgroundChronic cerebral hypoperfusion causes damage to the brain’s white matter underpinning vascular cognitive impairment. Inflammation and oxidative stress have been proposed as key pathophysiological mechanisms of which the transcription factor Nrf2 is a master regulator. We hypothesised that white matter pathology, microgliosis, blood-brain barrier breakdown and behavioural deficits induced by chronic hypoperfusion would be exacerbated in mice deficient in the transcription factor Nrf2.MethodsMice deficient in Nrf2 (male heterozygote or homozygous for Nrf2 knockout) or wild-type littermates on a C57Bl6/J background underwent bilateral carotid artery stenosis (BCAS) to induce chronic cerebral hypoperfusion or sham surgery and survived for a further 6 weeks. White matter pathology was assessed with MAG immunohistochemistry as a marker of altered axon-glial integrity; alterations to astrocytes and microglia/macrophages were assessed with GFAP and Iba1 immunohistochemistry, and blood-brain barrier breakdown was assessed with IgG immunohistochemistry. Behavioural alterations were assessed using 8-arm radial arm maze, and alterations to Nrf2-related and inflammatory-related genes were assessed with qRT-PCR.ResultsChronic cerebral hypoperfusion induced white matter pathology, elevated microglial/macrophage levels and blood-brain barrier breakdown in white matter tracts that were increased in Nrf2+/− mice and further exacerbated by the complete absence of Nrf2. Chronic hypoperfusion induced white matter astrogliosis and induced an impairment in behaviour assessed with radial arm maze; however, these measures were not affected by Nrf2 deficiency. Although Nrf2-related antioxidant gene expression was not altered by chronic cerebral hypoperfusion, there was evidence for elevated pro-inflammatory related gene expression following chronic hypoperfusion that was not affected by Nrf2 deficiency.ConclusionsThe results demonstrate that the absence of Nrf2 exacerbates white matter pathology and microgliosis following cerebral hypoperfusion but does not affect behavioural impairment.

Monitoring oxidative stress, immune response, Nrf2/NF-κB signaling molecules of Rhynchocypris lagowski living in BFT system and exposed to waterborne ammonia

The present study was implemented to evaluate oxidative stress, immune response, Nrf2 and NF-κB signaling molecules related genes expression of Rhynchocypris lagowski living in biofloc technology (BFT) system and exposed to waterborne ammonia. According to the differences of C:N ratios, the experiment was divided into four groups: C:N 10.8:1 (control group), C:N 15:1, C:N 20: 1 and C:N 25:1. The results demonstrated that BFT can effectively regulate water quality and promote growth, and the C:N 20:1 group has the most significant effect (P < 0.05). Besides, significant increases in immune enzymes (lysozyme, complement C3, C4, immunoglobulin M and nitric oxide synthase) and anti-inflammatory factor (IL-2) activity of R. lagowski were emerged in the treatment C:N 20:1 after the 56-d growth experiment and the challenging trial (P < 0.05). Comparing the antioxidant status of R. lagowski in liver and gut before and after ammonia stress: superoxide dismutase, total antioxidant capacity and catalase activity in treatments C:N 20:1 were significant increased (P < 0.05), while the level of malondialdehyde was marked lower than that in control. Meanwhile, treatment C:N 20:1 considerably upregulated Nrf2 signaling molecules related genes and significantly down-regulated the expression of pro-inflammatory factor gene in NF-κB signaling pathway compared with the control (P < 0.05). These results indicated that BFT could enhance growth, antioxidant and immune response and regulate Nrf2 and NF-κB related genes expression in R. lagowski, with most excellent effects in fish given C:N 20:1 group.