Activation Of Nrf2 Pathway Research Articles

Photobiomodulation Controls Keratinocytes Inflammatory Response through Nrf2 and Reduces Langerhans Cells Activation.

Photobiomodulation (PBM) is rapidly gaining traction as a valuable tool in dermatology for treating many inflammatory skin conditions using low levels of visible light or near-infrared radiation. However, the physiological regulatory pathways responsible for the anti-inflammatory effect of PBM have not been well defined. Since previous studies showed that nuclear factor-erythroid 2 like 2 (Nrf2) is a master regulator of the skin inflammatory response, we have addressed its role in controlling inflammation by PBM. Primary human keratinocytes (KCs) stimulated with 2,4-dinitrochlorobenzene (DNCB) to mimic pro-inflammatory stress were illuminated with two wavelengths: 660 nm or 520 nm. Both lights significantly reduced the mRNA expression of the DNCB-triggered TNF-α, IL-6, and IL-8 cytokines in KCs, while they enhanced Nrf2 pathway activation. PBM-induced Nrf2 is a key regulator of the inflammatory response in KCs since its absence abolished the regulatory effect of light on cytokines production. Further investigations of the mechanisms contributing to the immunoregulatory effect of PBM in inflamed human skin explants showed that 660 nm light prevented Langerhans cells migration into the dermis, preserving their dendricity, and decreased pro-inflammatory cytokine production compared to the DNCB-treated group. This study is the first to report that the PBM-mediated anti-inflammatory response in KCs is Nrf2-dependent and further support the role of PBM in skin immunomodulation. Therefore, PBM should be considered a promising alternative or complementary therapeutic approach for treating skin-related inflammatory diseases.

Nuclear Factor Erythroid 2-Related Factor 2 and Its Targets in Skeletal Muscle Repair and Regeneration.

Significance: Skeletal muscles have a robust regenerative capacity in response to acute and chronic injuries. Muscle repair and redox homeostasis are intimately linked; increased generation of reactive oxygen species leads to cellular dysfunction and contributes to muscle wasting and progression of muscle diseases. In exemplary muscle disease, Duchenne muscular dystrophy (DMD), caused by mutations in the DMD gene that encodes the muscle structural protein dystrophin, the regeneration machinery is severely compromised, while oxidative stress contributes to the progression of the disease. The nuclear factor erythroid 2-related factor 2 (NRF2) and its target genes, including heme oxygenase-1 (HO-1), provide protective mechanisms against oxidative insults. Recent Advances: Relevant advances have been evolving in recent years in understanding the mechanisms by which NRF2 regulates processes that contribute to effective muscle regeneration. To this end, pathways related to muscle satellite cell differentiation, oxidative stress, mitochondrial metabolism, inflammation, fibrosis, and angiogenesis have been studied. The regulatory role of NRF2 in skeletal muscle ferroptosis has been also suggested. Animal studies have shown that NRF2 pathway activation can stop or reverse skeletal muscle pathology, especially when endogenous stress defence mechanisms are imbalanced. Critical Issues: Despite the growing recognition of NRF2 as a factor that regulates various aspects of muscle regeneration, the mechanistic impact on muscle pathology in various models of muscle injury remains imprecise. Future Directions: Further studies are necessary to fully uncover the role of NRF2 in muscle regeneration, both in physiological and pathological conditions, and to investigate the possibilities for development of new therapeutic modalities. Antioxid. Redox Signal. 38, 619-642.

Loureirin C inhibits ferroptosis after cerebral ischemia reperfusion through regulation of the Nrf2 pathway in mice

BackgroundIschemic stroke (IS) is considered as a serious cerebral vascular disease. Ferroptosis is a novel type of regulated cell death (RCD), that closely related to the occurrence and progress of IS. Loureirin C, a type of dihydrochalcone compound derived from the Chinese Dragon's blood (CDB). The effective components extracted from CDB have shown neuroprotective effects in ischemia reperfusion models. However, the role of Loureirin C in mice after IS is not well understood. Thus, it is worth to identify the effect and mechanism of Loureirin C on IS. PurposeThe present research aims to prove the existence of ferroptosis in IS and explore whether Loureirin C can inhibit ferroptosis by regulating nuclear factor E2 related factor 2 (Nrf2) pathway in mice and exert neuroprotective effects on IS models. MethodsMiddle cerebral artery occlusion and reperfusion (MCAO/R) model was established to evaluate the occurrence of ferroptosis and the potential Loureirin C brain-protective effect in vivo. The analysis of free iron, glutamate content, reactive oxygen species (ROS) and lipid peroxidation levels, along with transmission electron microscope (TEM) was applied to prove the existence of ferroptosis. The function of Loureirin C on Nrf2 nuclear translocation was verified by immunofluorescence staining. In vitro, primary neurons and SH-SY5Y cells were processed with Loureirin C after oxygen and glucose deprivation-reperfusion (OGD/R). ELISA kits, western blotting, co-immunoprecipitation (Co-IP) analysis, immunofluorescence, and quantitative real-time PCR were devoted to proving the neuroprotective effects of Loureirin C on IS via regulating ferroptosis and Nrf2 pathways. ResultsThe results showed that Loureirin C not only dramatically alleviated brain injury and inhibited neurons ferroptosis in mice after MCAO/R, but also dose-dependently reduce ROS accumulation in ferroptosis after OGD/R. Further, Loureirin C inhibits ferroptosis by activating Nrf2 pathway, and promoting nuclear translocation of Nrf2. Besides, Loureirin C increases heme oxygenase 1 (HO-1), quinone oxidoreductase 1 (NQO1) and glutathione peroxidase 4 (GPX4) content after IS. Intriguingly, the anti-ferroptosis effect of Loureirin C is weakened by Nrf2 knockdown. ConclusionOur discoveries first revealed that the inhibitory action of Loureirin C on ferroptosis may greatly depend on its adjusting effect on the Nrf2 pathway, suggesting that Loureirin C could act as a novel anti-ferroptosis candidate and play a therapeutic role in IS. These novel discoveries on the role of Loureirin C on IS models reveal an innovative method that may contribute to neuroprotection for the prevention of IS.

Polydatin Alleviates Chronic Stress-Induced Depressive and Anxiety-like Behaviors in a Mouse Model.

We aimed to investigate whether polydatin could suppress stress-induced depression- and anxiety-like behaviors in a mouse model. Mice were divided into the control group, chronic unpredictable mild stress (CUMS) exposure group, and CUMS mice treated with polydatin group. Following CUMS exposure and polydatin treatment, mice were subjected to behavioral assays to assess depressive-like and anxiety-like behaviors. Synaptic function was determined by the levels of brain-derived neurotrophic factor (BDNF), postsynaptic density protein 95 (PSD95), and synaptophysin (SYN) in the hippocampus and cultured hippocampal neurons. The number and length of dendrites were assessed in cultured hippocampal neurons. Finally, we investigated the effect of polydatin on CUMS-induced inflammation and oxidative stress in the hippocampus by measuring inflammatory cytokine levels, oxidative stress markers such as reactive oxygen species, glutathione peroxidase, catalase, and superoxide dismutase, as well as components of the Nrf2 signaling pathway. Polydatin alleviated CUMS-induced depressive-like behaviors in forced swimming, tail suspension and sucrose preference tests, and anxiety-like behaviors in marble-burying and elevated plus maze tests. Polydatin increased the number and length of dendrites of cultured hippocampal neurons from mice exposed to CUMS and alleviated CUMS-induced synaptic deficits by restoring BDNF, PSD95, and SYN levels in vivo and in vitro. Importantly, polydatin inhibited CUMS-induced hippocampal inflammation and oxidative stress and suppressed the activation of NFκB and Nrf2 pathways. Our study suggests that polydatin may be an effective drug for the treatment of affective disorders through inhibiting neuroinflammation and oxidative stress. Our current findings warrant further study to investigate the potential clinical application of polydatin.

Melatonin attenuates high glucose‑induced endothelial cell pyroptosis by activating the Nrf2 pathway to inhibit NLRP3 inflammasome activation.

Endothelial injury induced by hyperglycemia is the most critical initial step in the development of diabetic vasculopathy. The aim of this present study was to explore the prevention and treatment strategies and elucidate the specific mechanism of diabetes‑induced vascular endothelial injury. Melatonin, a hormone secreted by the pineal gland to regulate biological rhythm, serves an important role in maintaining human physiological function. Pyroptosis is a type of newly discovered inflammatory cell death. The current study first found by western blotting that melatonin could activate nuclear factor erythroid 2‑related factor 2 (Nrf2) pathway in human umbilical vein endothelial cells (HUVECs) under high glucose (HG) condition. Second, it found that pretreatment with Luzindole, a specific inhibitor of melatonin receptor (MT1/MT2), significantly reduced the activation of Nrf2 pathway by melatonin in HUVECs. It also found that pretreatment with melatonin or a specific NOD‑like receptor family, pyrin domain‑containing 3 (NLRP3) inhibitor (MCC950) pretreatment reduced HG‑induced endothelial cell pyroptosis. Finally, it was found that the protective effect of melatonin against reactive oxygen species/NLRP3 inflammasome pathway activation induced by HG in HUVECs was decreased after Nrf2 knockdown. In conclusion, the present study showed that melatonin may serve a protective role in HG‑induced vascular endothelial cell pyroptosis by activating the Nrf2 pathway to inhibit NLRP3 inflammasome activation. In addition, it was further found that melatonin attenuated HG‑induced vascular endothelial cell injury by interacting with its receptors (MT1/MT2) to promote activation of Nrf2 pathway.

Baicalin ameliorates high fat diet-induced nonalcoholic fatty liver disease in mice via adenosine monophosphate-activated protein kinase-mediated regulation of SREBP1/Nrf2/NF-κB signaling pathways.

Nonalcoholic fatty liver disease (NAFLD) is a prevalent chronic liver disease around the world, imposing severe threats on human health. Unfortunately, no clinically approved drugs are available for use as yet. Baicalin (BA) is reported to have hepatoprotective effects, and it is not clear whether BA can treat NAFLD and how. Here, a high-fat diet (HFD)-induced NAFLD mouse model was established to explore the protective roles and mechanisms of BA against HFD-induced NAFLD. Physiochemical results showed that BA exhibited significantly protective effects against HFD-induced NAFLD in mice. Liver transcriptomic analysis revealed that BA attenuated HFD-induced NAFLD via activating AMPK pathway, which was confirmed by the AMPK inhibitor Compound C. Additionally, the expression changes of AMPK downstream genes demonstrated that BA exerted ameliorative effects against NAFLD through AMPK-mediated inhibition of SREBP1 and NF-κB pathways, and activation of Nrf2 pathway. Taken together, our study reveals the protective roles of BA against HFD-caused NAFLD through AMPK-mediated modulation of SREBP1/Nrf2/NF-κB pathways, suggesting that BA has potential drug development implications. Most importantly, our study creates a paradigm through the combination of molecular biology and bioinformatics for further studies of action mechanisms of biomolecules combating diseases.

Berberine Alleviates Doxorubicin-Induced Myocardial Injury and Fibrosis by Eliminating Oxidative Stress and Mitochondrial Damage via Promoting Nrf-2 Pathway Activation.

Doxorubicin (DOX)-related cardiotoxicity has been recognized as a serious complication of cancer chemotherapy. Effective targeted strategies for myocardial protection in addition to DOX treatment are urgently needed. The purpose of this paper was to determine the therapeutic effect of berberine (Ber) on DOX-triggered cardiomyopathy and explore the underlying mechanism. Our data showed that Ber markedly prevented cardiac diastolic dysfunction and fibrosis, reduced cardiac malondialdehyde (MDA) level and increased antioxidant superoxide dismutase (SOD) activity in DOX-treated rats. Moreover, Ber effectively rescued the DOX-induced production of reactive oxygen species (ROS) and MDA, mitochondrial morphological damage and membrane potential loss in neonatal rat cardiac myocytes and fibroblasts. This effect was mediated by increases in the nuclear accumulation of nuclear erythroid factor 2-related factor 2 (Nrf2) and levels of heme oxygenase-1 (HO-1) and mitochondrial transcription factor A (TFAM). We also found that Ber suppressed the differentiation of cardiac fibroblasts (CFs) into myofibroblasts, as indicated by decreased expression of α-smooth muscle actin (α-SMA), collagen I and collagen III in DOX-treated CFs. Pretreatment with Ber inhibited ROS and MDA production and increased SOD activity and the mitochondrial membrane potential in DOX-challenged CFs. Further investigation indicated that the Nrf2 inhibitor trigonelline reversed the protective effect of Ber on both cardiomyocytes and CFs after DOX stimulation. Taken together, these findings demonstrated that Ber effectively alleviated DOX-induced oxidative stress and mitochondrial damage by activating the Nrf2-mediated pathway, thereby leading to the prevention of myocardial injury and fibrosis. The current study suggests that Ber is a potential therapeutic agent for DOX-induced cardiotoxicity that exerts its effects by activating Nrf2.

OP30 Human umbilical cord mesenchymal stem cells derived exosomes alleviate Ulcerative Colitis by inhibiting macrophage ferroptosis via miR-23b-3p/Nrf2 pathway

Abstract Background Human umbilical cord mesenchymal stem cell derived exosomes (HucMSC-Exo) have broad application prospects in treatment of various disease, including ulcerative colitis (UC). Macrophages can trigger intestinal mucosal inflammation thereby participating in the onset and development of UC. As a newly discovered iron-dependent cell death pathway, ferroptosis has also been found accompanied by inflammatory response in recent studies. We speculated that there could be relationship between macrophage ferroptosis and inflammatory response in UC. Furthermore, the role of macrophage ferroptosis in the process of HucMSC-Exo alleviating UC remains undefined. This study aims to explore the therapeutic effect and mechanism of HucMSC-Exo on UC by regulating macrophage ferroptosis. Methods The gene expression of ferroptosis-related proteins was assessed in colon samples from healthy controls and UC patients to explore the relationship between ferroptosis and UC. In vivo, colitis was induced by dextran sulfate sodium in mice. HucMSC-Exo was administered intravenously to estimate its curative effect, and its gut homing was evaluated by animal live imaging analysis. In vitro, HucMSC-Exo were cocultured with ferroptosis inducer RSL3-treated bone marrow-derived macrophages (BMDMs), RAW264.7 cell line, followed by examining the effect of HucMSC-Exo. Transfection of microRNA mimics and gene knockdown were used to confirm the role of miR-23b-3p in HucMSC-Exo regulating of macrophage ferroptosis. Results Compared with healthy individuals, ferroptosis may be involved in the mucosal inflammation of UC patients. HucMSC-Exo were found to successfully target damaged colon tissues in vivo. HucMSC-Exo treatment significantly improved the pathological damage of colon tissue, increased the expression of glutathione peroxidase 4 (GPX4), while markedly decreasing the levels of iron and lipid ROS, and reducing the secretion of interleukin (IL)-1β and IL-6, and then relieved macrophage ferroptosis to alleviate murine colitis. In vitro experiments showed that HucMSC-Exo could be effectively absorbed by macrophages and rescued RSL3-induced ferroptosis, which was consistent with the effect of ferrostatin-1, a ferroptosis-specifific inhibitor. Mechanismly, HucMSC-exo could inhibit macrophage ferroptosis by delivering its highly expressed miR-23b-3p to activate Nrf2 pathway. Additionally, the protection of HucMSC-exo for UC was attenuated by decreasing the expression of miR-23b-3p or inhibiting Nrf2 pathway. Conclusion HucMSC-Exo could activate Nrf2 antioxidant pathway and abrogate macrophage ferroptosis by delivering miR-23b-3p and ultimately ameliorate colitis.

Longitudinal Effects of 1-Year Smoking Cessation on Human Bronchial Epithelial Transcriptome

Longitudinal Effects of 1-Year Smoking Cessation on Human Bronchial Epithelial Transcriptome

Withaferin A, a steroidal lactone, selectively protects normal lymphocytes against ionizing radiation induced apoptosis and genotoxicity via activation of ERK/Nrf-2/HO-1 axis

Increasing use of ionizing radiation (IR) in medicine, industry, agriculture and research ensues potential health hazards if not used properly or contained effectively. However, radioprotectors which are effective in clinical and/or accidental radiation exposures are still elusive. In this direction, we have explored the radioprotective potential of Withaferin A, a plant withanolide, which was recently shown to be safe and well tolerated in cancer patients in a clinical trial and is also known to be a radio-sensitizer in cancer cells. Our results show that, Withaferin A (WA) protected only normal lymphocytes, but not cancer cells, against IR-induced apoptosis and offered radioprotection even when added post-radiation exposure. WA treatment led to significant inhibition of IR-induced caspase-3 activation and decreased IR-induced DNA damage to lymphocytes and bone-marrow cells. WA reduced intracellular ROS and GSH levels and only thiol based anti-oxidants could abrogate the radio-protective effects of WA, indicating a crucial role of cellular/protein thiols in its biological activity. The inability of WA-glutathione adduct to offer radioprotection further underscored the role of cellular thiols. WA induced pro-survival transcription factor, Nrf-2, and expression of cytoprotective genes HO-1, catalase, SOD, peroxiredoxin-2 via ERK. Further, WA administration could rescue mice against radiation induced mortality, DNA damage, increase in micro-nucleated polychromatic erythrocytes (mn-PCEs) and increased ratio of polychromatic erythrocytes (PCEs) to Normochromatic Erythrocytes (NCEs) in bone-marrow, demonstrating its potent in vivo the radio-protective efficacy. In conclusion, WA selectively protects normal cells against IR-induced apoptosis via activation of cytoprotective Nrf-2 pathway.

Unraveling Hepatic Metabolomic Profiles and Morphological Outcomes in a Hybrid Model of NASH in Different Mouse Strains.

Nonalcoholic fatty liver disease (NAFLD) encompasses nonalcoholic steatohepatitis (NASH) and affects 25% of the global population. Although a plethora of experimental models for studying NASH have been proposed, still scarce findings regarding the hepatic metabolomic/molecular profile. In the present study, we sought to unravel the hepatic metabolomic profile of mice subjected to a hybrid model of NASH, by combining a Western diet and carbon tetrachloride administration, for 8 weeks, in male C57BL/6J and BALB/c mice. In both mouse strains, the main traits of NASH-metabolic (glucose intolerance profile), morphologic (extensive microvesicular steatosis and fibrosis, lobular inflammation, and adipose tissue-related inflammation/hypertrophy), and molecular (impaired Nrf2/NF-κB pathway dynamics and altered metabolomic profile)-were observed. The hepatic metabolomic profile revealed that the hybrid protocol impaired, in both strains, the abundance of branched chain-aromatic amino acids, carboxylic acids, and glycosyl compounds, that might be linked to the Nrf2 pathway activation. Moreover, we observed a strain-dependent hepatic metabolomic signature, in which the tricarboxylic acid metabolites and pyruvate metabolism were dissimilarly modulated in C57BL/6J and BALB/c mice. Thus, we provide evidence that the strain-dependent hepatic metabolomic profile might be linked to the distinct underlying mechanisms of NASH, also prospecting potential mechanistic insights into the corresponding disease.

Duality of Nrf2 in iron-overload cardiomyopathy.

Cardiomyopathy deeply affects quality of life and mortality of patients with b-thalassemia or with transfusion-dependent myelodysplastic syndromes. Recently, a link between Nrf2 activity and iron metabolism has been reported in liver ironoverload murine models. Here, we studied C57B6 mice as healthy control and nuclear erythroid factor-2 knockout (Nrf2-/-) male mice aged 4 and 12 months. Eleven-month-old wild-type and Nrf2-/- mice were fed with either standard diet or a diet containing 2.5% carbonyl-iron (iron overload [IO]) for 4 weeks. We show that Nrf2-/- mice develop an age-dependent cardiomyopathy, characterized by severe oxidation, degradation of SERCA2A and iron accumulation. This was associated with local hepcidin expression and increased serum non-transferrin-bound iron, which promotes maladaptive cardiac remodeling and interstitial fibrosis related to overactivation of the TGF-b pathway. When mice were exposed to IO diet, the absence of Nrf2 was paradoxically protective against further heart iron accumulation. Indeed, the combination of prolonged oxidation and the burst induced by IO diet resulted in activation of the unfolded protein response (UPR) system, which in turn promotes hepcidin expression independently from heart iron accumulation. In the heart of Hbbth3/+ mice, a model of b-thalassemia intermedia, despite the activation of Nrf2 pathway, we found severe protein oxidation, activation of UPR system and cardiac fibrosis independently from heart iron content. We describe the dual role of Nrf2 when aging is combined with IO and its novel interrelation with UPR system to ensure cell survival. We open a new perspective for early and intense treatment of cardiomyopathy in patients with b-thalassemia before the appearance of heart iron accumulation.

Dysregulation and Epigenetic Reprogramming of NRF2 Signaling Axis Promote Acquisition of Cisplatin Resistance and Metastasis in Head and Neck Squamous Cell Carcinoma.

Cisplatin (CDDP)-based chemotherapy is a first-line treatment for patients with advanced head and neck squamous cell carcinomas (HNSCC), despite a high rate of treatment failures, acquired resistance, and subsequent aggressive behavior. The purpose of this study was to study the mechanism of CDDP resistance and metastasis in HNSCC. We investigated the role of NRF2 pathway activation as a driven event for tumor progression and metastasis of HNSCC. Human HNSCC cell lines that are highly resistant to CDDP were generated. Clonogenic survival assays and a mouse model of oral cancer were used to examine the impact of NRF2 activation in vitro and in vivo on CDDP sensitivity and development of metastasis. Western blotting, immunostaining, whole-exome sequencing, single-cell transcriptomic and epigenomic profiling platforms were performed to dissect clonal evolution and molecular mechanisms. Implantation of CDDP-resistant HNSCC cells into the tongues of nude mice resulted in a very high rate of distant metastases. The CDDP-resistant cells had significantly higher expression of NRF2 pathway genes in the presence of newly acquired KEAP1 mutations, or via epigenomic activation of target genes. Knockdown of NRF2 or restoration of the wild-type KEAP1 genes resensitized resistant cells to CDDP and decreased distant metastasis (DM). Finally, treatment with inhibitor of glutaminase-1, a NRF2 target gene, alleviated CDDP resistance. CDDP resistance and development of DM are associated with dysregulated and epigenetically reprogrammed KEAP1-NRF2 signaling pathway. A strategy targeting KEAP1/NRF2 pathway or glutamine metabolism deserves further clinical investigation in patients with CDDP-resistant head and neck tumors.

PNaktide mitigates inflammation-induced neuronal damage and behavioral deficits through the oxidative stress pathway

Neuroinflammation is closely related to the etiology and progression of neurodegenerative diseases such as Parkinson disease and Alzheimer disease. pNaktide, an Src inhibitor, exerts antioxidant effects by mimicking Na/K-ATPase. It has been verified that its anti-inflammation and anti-oxidation ability could be embodied in obesity, steatohepatitis, uremic cardiomyopathy, aging, and prostate cancer. This study aimed to investigate the effects and mechanisms of pNaktide in lipopolysaccharide (LPS)-induced behavioral damage, neuroinflammation, and neuronal damage. We found that pNaktide improved anxiety, memory, and motor deficits. pNaktide inhibited MAPK and NF-κB pathways induced by TLR4 activation, inhibited the NLRP3 inflammasome complex, and reduced the expression of inflammatory factors, complement factors, and chemokines. pNaktide inhibited the activation of Nrf2 and HO-1 antioxidant stress pathways by LPS and reduced the level of oxidative stress. Inhibition of autophagy and enhancement of apoptosis induced by LPS were also alleviated by pNaktide, which restored LPS-induced injury to newborn neurons in the hippocampus region. In summary, pNaktide attenuates neuroinflammation, reduces the level of oxidative stress, has neuroprotective effects, and may be used for the treatment of neuroinflammation-related diseases.

Inhibitors of Keap1-Nrf2 protein-protein interaction reduce estrogen responsive gene expression and oxidative stress in estrogen receptor-positive breast cancer

Estrogen contributes to the development of breast cancer through estrogen receptor (ER) signaling and by generating genotoxic metabolites that cause oxidative DNA damage. To protect against oxidative stress, cells activate nuclear factor erythroid 2-related factor 2 (Nrf2) and its downstream cytoprotective genes that initiate antioxidant responses and detoxify xenobiotics. Nrf2 activation occurs by inhibiting the protein-protein interaction (PPI) between Nrf2 and its inhibitor Keap1, which otherwise targets Nrf2 for ubiquitination and destruction. In this study, we examined a series of novel direct inhibitors of Keap1-Nrf2 PPI in their role in promoting the availability of Nrf2 for antioxidant activity and attenuating estrogen-mediated responses in breast cancer. ER-positive human breast cancer cells MCF-7 were treated with 17β-estradiol (E2) in the presence or absence of selected Keap1-Nrf2 PPI inhibitors. Keap1-Nrf2 PPI inhibitors suppressed the mRNA and protein levels of estrogen responsive genes induced by E2 exposure, such as PGR. Keap1-Nrf2 PPI inhibitors caused significant activation of Nrf2 target genes. E2 decreased the mRNA and protein level of the Nrf2 target gene NQO1, and the Keap1-Nrf2 PPI inhibitors reversed this effect. The reversal of E2 action by these compounds was not due to binding to ER as ER antagonists. Further, a selected compound attenuated oxidative stress induced by E2, determined by the level of a biomarker 8-oxo-deoxyguanosine. These findings suggest that the Keap1-Nrf2 PPI inhibitors have potent antioxidant activity by activating Nrf2 pathways and inhibit E2-induced gene and protein expression. These compounds may serve as potential chemopreventive agents in estrogen-stimulated breast cancer.

Kaempferol prevents acetaminophen-induced liver injury by suppressing hepatocyte ferroptosis via Nrf2 pathway activation.

Acetaminophen (APAP)-induced liver injury (AILI) has become a growing public health problem. Ferroptosis, an iron-dependent form of cell death associated with lipid peroxide accumulation, has been recently implicated in AILI. The activation of the Nrf2 signaling pathway is a potential therapy for AILI. Kaempferol (KA), a flavonoid widely existing in edible plants, has been reported to exert profound anti-inflammatory and antioxidant activities. This study aimed to investigate whether KA exerts anti-AILI effects via the Nrf2 signaling pathway. Mice were fasted for 22 h and injected intraperitoneally with APAP (250 mg kg-1) to induce AILI. Mice were pre-injected intragastrically with KA for 2 h followed by APAP injection. The hepatic injury was observed by H&E staining. Biochemical parameters of the serum and liver were measured using kits. KA alleviated hepatic injury and inflammatory response in AILI mice and ameliorated APAP-induced hepatic iron overload and oxidative stress in mice. In addition, the protective effects of KA against APAP-induced hepatotoxicity were examined in L02 cells in vitro. Cell viability was assayed by the CCK8 assay. Mitochondrial reactive oxygen species (ROS) in L02 cells were detected by MitoSox fluorescence. KA reversed the APAP-induced decrease in cell viability and GSH levels and inhibited the accumulation of intracellular ROS. Furthermore, KA activated the Nrf2 pathway and upregulated Gpx4 in mouse livers and L02 cells to inhibit ferroptosis induced by APAP. Finally, molecular docking indicated the potential interaction of KA with Keap1. Taken together, KA ameliorated oxidative stress and ferroptosis-mediated AILI by activating Nrf2 signaling.

The ethanolic extract of Curcuma longa grown in Korea exhibits anti-neuroinflammatory effects by activating of nuclear transcription factor erythroid-2-related factor 2/heme oxygenase-1 signaling pathway

BackgroundCurcuma longa has been used as spices, food preservative, coloring material, and traditional medicine. This plant also has long been used for a variety of diseases including dyslipidemia, stomach disorders, arthritis, and hepatic diseases. The aim of the present investigation was to examine the anti-neuroinflammatory effects of the 50% ethanolic extract of C. longa in lipopolysaccharide (LPS)-induced BV2 microglial cells.MethodsGriess reaction was employed to measure the production of nitric oxide (NO), and the levels of prostaglandin E2 (PGE2) and pro-inflammatory cytokines such as interleukin 1-beta (IL-1β), IL-6 and tumor necrosis factor-α (TNF-α) were determined by using profit ELISA kits. Western blotting was used to determine the expression of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), nuclear factor kappa B (NF-κB), mitogen activated protein kinases (MAPKs), heme oxygenase-1 (HO-1) and nuclear factor erythroid-2-related factor 2 (Nrf2).ResultsPre-treatment with CLE inhibited the overproduction and overexpression of pro-inflammatory mediators including NO, PGE2, iNOS, COX-2, and pro-inflammatory cytokines such as IL-1β, IL-6 and TNF-α in LPS-induced BV2 cells. In addition, CLE suppressed the activation of the NF-κB and three MAPK signaling pathways. Treatment with CLE induced HO-1 protein expression by activating Nrf2 pathway, and inhibiting the HO-1 expression reversed the anti-inflammatory effect of CLE.ConclusionCLE showed anti-neuroinflammatory effects against LPS-induced microglial cells activation through the inhibition of production and expression of pro-inflammatory mediators by negative regulation of the NF-κB and MAPK signaling pathways. These anti-neuroinflammatory effects of CLE were mediated by HO-1/Nrf2 signaling pathway. Taken together, the present study suggests a potent effect of CLE to prevent neuroinflammatory diseases. It is necessary to perform additional efficacy evaluation through in vivo experiments.

Hydroxytyrosol attenuates diquat-induced oxidative stress by activating Nrf2 pathway and modulating colonic microbiota in mice

This study was conducted to investigate the antioxidant effects of hydroxytyrosol (HT) administration in diquat (DQ)-challenged mice. The results showed that HT treatment markedly alleviated DQ-induced oxidative stress, which was indicated by the enhanced total antioxidant capacity (T-AOC), increased activities of superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and catalase and decreased malondialdehyde (MDA) concentration in serum. Additionally, HT increased the mRNA expression levels of NF-E2-related factor 2 (Nrf2) and its downstream genes, including NADPH quinone oxidoreductase 1 (NQO1) and catalase (CAT) in the small intestine of DQ-challenged mice. 16S rRNA gene sequencing results showed that HT treatment increased the relative abundance of Firmicutes and Lactobacillus and decreased the relative abundance of Bacteroidetes. Interestingly, Pearson correlation analysis showed that there were strong association between colonic Firmicutes, Lactobacillus, and Bacteroidetes and the activities of serum antioxidant enzymes. Meanwhile, HT significantly enhanced the colonic butyrate concentration in DQ-challenged mice. Additionally, HT treatment decreased the serum metabolites involving in glycerophospholipid metabolism, pentose, and glucuronate interconversions, which were associated with alleviated oxidative stress. These results indicate that oral administration of 100 mg/kg body weight HT alleviates oxidative stress in DQ-challenged mice, which may involve Nrf2 signaling pathways via modulation of colonic microbiota.

Blockage of Nrf2 and autophagy by L-selenocystine induces selective death in Nrf2-addicted colorectal cancer cells through p62-Keap-1-Nrf2 axis

Persistent Nrf2 activation is typically noted in many cancers, including colorectal cancer (CRC), aiding cancer cells in overcoming growth stress and promoting cancer progression. Sustained Nrf2 activation, which is beneficial for cancer cells, is called “Nrf2 addiction”; it is closely associated with malignancy and poor prognosis in patients with cancer. However, Nrf2 inhibitors may have adverse effects on normal cells. Here, we found that the selenocompound l-selenocystine (SeC) is selectively cytotoxic in the Nrf2-addicted CRC cell line WiDr cells, but not in non–Nrf2-addicted mesenchymal stem cells (MSCs) and normal human colon cells. Another CRC cell line, C2BBe1, which harbored lower levels of Nrf2 and its downstream proteins were less sensitive to SeC, compared with the WiDr cells. We further demonstrated that SeC inhibited Nrf2 and autophagy activation in the CRC cells. Antioxidant GSH pretreatment partially rescued the CRC cells from SeC-induced cytotoxicity and Nrf2 and autophagy pathway inhibition. By contrast, SeC activated Nrf2 and autophagy pathway in non–Nrf2-addicted MSCs. Transfecting WiDr cells with Nrf2-targeting siRNA decreased persistent Nrf2 activation and alleviated SeC cytotoxicity. In KEAP1-knockdown C2BBe1 cells, Nrf2 pathway activation increased SeC sensitivity and cytotoxicity. In conclusion, SeC selectively attacks cancer cells with constitutively activated Nrf2 by reducing Nrf2 and autophagy pathway protein expression through the P62–Nrf2–antioxidant response element axis and eventually trigger cell death.

SULFORAPHANE ADMINISTRATION AFTER HEMORRHAGIC SHOCK/RESUSCITATION IN MICE REDUCES THE SECRETION OF INFLAMMATORY CYTOKINES AND INCREASES THE IMMUNOCOMPETENCE OF SPLENIC MACROPHAGES.

Objective : The purpose of this study was to investigate the immunomodulatory effects of sulforaphane (SFN), a nuclear factor erythroid 2-related factor (Nrf2) pathway activator, on splenic macrophages' immunocompetence after hemorrhagic shock/resuscitation (HS/R). Methods : Male C57/BL6 wild-type mice (n = 6 per group) were subjected to either pressure-controlled HS (MAP, 35-45 mm Hg) or a sham procedure surgery (without HS). After 90 minutes of HS, fluid resuscitation with withdrawn blood and 0.9% NaCl was performed. Sulforaphane (50 mg/kg of body weight) was applied intraperitoneally immediately after the resuscitation phase as well as 24 and 48 h thereafter, depending on group allocation. The mice were killed at 6, 24, and 72 h after resuscitation. After killing, spleens were harvested to perform Nrf2 immunofluorescence histology. Splenic macrophages were isolated and cultured to measure cytokine secretion in the cell culture supernatant. Furthermore, macrophages isolated after 24-hour resuscitation were treated with 100 ng/mL of bacterial LPS to measure immunocompetence. Matrix-assisted laser desorption/ionization mass spectrometry imaging was performed to verify the distribution of SFN in the spleen after intraperitoneal injection. Results : We showed that administered SFN reached the spleen within the first hour after administration. Furthermore, we identified that SFN increased splenic Nrf2 activation and decreased cytokine expression in splenic macrophages after HS/R. In addition, we showed that SFN exhibited splenic anti-inflammatory properties of macrophages in vitro (IL-6/IL-10-ratio of the HS/R group: 51.79 ± 9.99 [at 6 h] and 15.70 ± 3.35 [at 24 h] vs. HS/R + SFN group: 20.54 ± 5.35 [at 6 h] and 8.60 ± 2.37 [at 24 h], P < 0.05). Furthermore, SFN improved in vitro splenic macrophage immunocompetence after HS/R, as evidenced by the increased secretion of inflammatory cytokines in response to LPS stimulation in vitro . Conclusions : Our study shows that SFN can reduce inflammatory cytokines secreted by splenic macrophages after HS/R and increase their immunocompetence toward a more anti-inflammatory profile.