Kelch-like ECH-associated Protein Research Articles

Modulation of NRF2: Biological Dualism in Cancer, Targets and Possible Therapeutic Applications.

Significance: The nuclear factor erythroid 2-related factor 2 (NRF2)-Kelch-like ECH-associated protein 1 (KEAP1) system is a master regulator of redox homeostasis and cell adaptation to a variety of exogenous and endogenous stressors. Accumulating evidence from the last decade indicates that the impairment of the redox balance leads to oxidative stress (OS), a common alteration occurring in many human acute and chronic inflammatory diseases, such as cancer, diabetes, neurodegeneration, and metabolic disorders, and aging. Recent Advances: Being located at the intersection of crucial signaling pathways, NRF2 can influence several cellular functions, which extend beyond the maintenance of the redox balance and include cellular metabolism, proteostasis, mitochondrial function and inflammation. For this reason, there is a growing interest in the pharmacologic manipulation of NRF2 for therapeutic purposes, which requires the accurate knowledge of the cell context and the specific time frame both of NRF2 activation and inhibition. This appears to be an important prerequisite and reflects the extreme complexity of the NRF2 signaling, characterized by an intrinsic dualism that mediates beneficial or detrimental effects even in the same biological process. Critical Issues: Of crucial importance will be to understand whether the NRF2 activity modulation might be exploited to exert beneficial outcomes in patients suffering from pathological conditions, in which the OS and the deregulation of inflammatory processes play a crucial role. Future Directions: In this review, we discuss the dual involvement of NRF2 in aging, neurodegeneration, metabolic diseases, long-COVID-19, and carcinogenesis and we present an overview of the most recent therapeutic modulators of NRF2, particularly emphasizing on those selected for clinical trials. Antioxid. Redox Signal. 40, 636-662.

Oxidative stress and inflammatory pathways in female eating disorders and borderline personality disorders with emotional dysregulation as linking factors with impulsivity and trauma

BackgroundBorderline personality disorder (BPD) and eating disorders (ED) are both disorders with emotional dysregulation that may share some similar biological underpinnings, leading to oxidative/inflammatory alterations. Unfortunately, to date, no studies have assessed the relationship between clinical features, inflammatory alterations and childhood trauma across these disorders. Our aim was to identify the potential common and disorder-specific inflammatory pathways and examine possible associationsbetween these dysregulated pathways and the clinical features. MethodsWe studied a sample of 108 women (m = 27.17 years; sd = 7.64), divided into four groups: 23 patients with a restrictive ED (ED-R), 23 patients with a bingeeating/ purging ED (ED-P) and 26 patients with BPD; whereas the control group included 23 healthy subjects. Several inflammatory/oxidative parameters: tumor necrosis factor alpha (TNFα), Thiobarbituric Acid Reactive Substances (TBARS), inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX2), p38 mitogenactivated protein kinases, ERK mitogen-activated protein kinases and c-Jun NH2- terminal kinase (JNK), and some antiinflammatory antioxidant elements: glutathione peroxidase (GPx), superoxide dismutase (SOD), catalase (CAT), Kelch-like ECHassociated protein (Keap1) were determined in plasma or peripheral blood mononuclear cells. Furthermore, clinical, impulsivity, trauma and eating behavior questionnaires were administered. ResultsThree main inflammatory/oxidative components were extracted using principal component analysis (59.19 % of biomarker variance explained). Disorder-specific dysfunction in the inflammatory and oxidative pathways in patients with BPD and ED were revealed by means of relationships with specific principal components (p < .01). BPD patients showed higher levels of a component featured by elevated levels of JNK and lower of GPx and SOD. ED-R and impulsivity were associated with a component featured by the activation of ERK and negative influence of Keap1. The component featured by the suppression of catalase and COX2 was associated with both ED subtypes and trauma exposure. ConclusionSeveral risk factors such as trauma, impulsivity and eating disorder symptoms were transdiagnostically associated with some inflammatory alterations regardless of diagnosis. These findings suggest that the clinical profile comprising trauma exposure and an emotional dysregulation disorder might constitute a specific endophenotype highly linked with inflammatory alterations.

SUMOylation of the ubiquitin ligase component KEAP1 at K39 upregulates NRF2 and its target function in lung cancer cell proliferation

Nuclear Factor Erythroid 2-Related Factor 2 (NRF2) is important for the expression of genes associated with oxidative stress. The levels of NRF2 are controlled by Kelch-like ECH-associated protein 1 (KEAP1)-dependent degradation. Although oxidative stress is known to suppress KEAP1 activity to stabilize the levels of NRF2, the mechanism for this control is unclear. Here, we identify that KEAP1 is modified by SUMO1 at the lysine residue position 39 (K39). Arginine replacement of this lysine (K39R) in KEAP1 did not affect its stability, subcellular localization, or dimerization but promoted the formation of the Cullin 3 ubiquitin ligase and increased NRF2 ubiquitination. This was accompanied by decreased NRF2 expression. Gene reporter assays showed that the transcription of antioxidant response elements was heightened in KEAP1-WT cells compared to cells expressing the KEAP1-K39R SUMO1 substrate mutant. Consistent with this, chromatin immunoprecipitation assays revealed higher NRF2 binding to the promoter regions of antioxidant genes in cells expressing the KEAP1-WT compared to the KEAP1-K39R mutant protein in H1299 lung cancer cell. The significance of this suppression of KEAP1 activity by its SUMOylation was tested in a subcutaneous tumor model of H1299 lung cancer cell lines that differentially expressed the WT and K39R KEAP1 constructs. This model showed that mutating the SUMOylation site on KEAP1 altered the production of reactive oxygen species and suppressed tumor growth. Taken together, our study recognizes that NRF2-dependent redox control is regulated by the SUMOylation of KEAP1. These findings identify a potential new therapeutic option to counteract oxidative stress.

Alantolactone triggers oxeiptosis in human ovarian cancer cells via Nrf2 signaling pathway

IntroductionA growing body of evidence indicated that Alantolactone (ALT) promotes Reactive Oxygen Species (ROS) generation exclusively in cancer cells. Therefore, the aim of this study was to investigate the effect of ALT on the molecular mechanism of oxeiptosis, as a novel cell death pathway due to the high levels of intracellular ROS in ovarian cancer. MethodsMTT assay was used to evaluate the effect of ALT on SKOV3 cell viability. mRNA and protein expression levels of Nrf2 (nuclear factor erythroid 2–related factor 2), KEAP1 (Kelch-like ECH-associated protein 1), PGAM5 (phosphoglycerate mutase family member 5), AIFM1 (Mitochondrial Apoptosis-Inducing Factor), Glutathione synthetase (GSS) and glutathione peroxidase (GPX) were analyzed by real time PCR and western blotting methods respectively. ResultsOur findings showed that ALT inhibits the proliferation of skov3 cells in a time and dose dependent manner and IC50 was 32 μM at 24h.A significant down-regulation of Nrf2, GSH and GPX mRNA levels was seen in skov3 cells incubated with 32 and 64 μM of ALT in comparison with control group, while, mRNA expression levels of PGAM5 and KEAP1 were increased.Western blot analysis showed that ALT significantly decreases protein levels of Nrf2 and increases PGAM5 and KEAP1.ALT dephosphorylated PS116-AIFM1 and total AIFM1 protein level was elevated. ConclusionOur results provided evidence that ALT could be a potential option for ovarian cancer treatment by ROS-mediated oxeiptosis.

Total triterpenes of Euphorbium alleviates rheumatoid arthritis via Nrf2/HO-1/GPX4 pathway

This study aims to investigate the effect and mechanism of total triterpenes of Euphorbium in treating rheumatoid arthritis(RA). The rat model of RA was established with Freund's complete adjuvant(FCA). Male rats were randomly assigned into control, model, Tripterygium glycosides(7.5 mg·kg~(-1)), and low-, medium-, and high-dose total triterpenes of Euphorbium(32, 64, and 128 mg·kg~(-1), respectively) groups, with 10 rats in each group. In other groups except the control group, 0.2 mL FCA was injected into the right hind toe. Rats in the intervention groups were administrated with corresponding drugs by gavage, and the control group and the model group with the same volume of 0.5% CMC-Na solution once a day. During the treatment period, the swelling degree of the hind paw was measured and the arthritis was scored until day 30. At the end of drug administration, the pathological changes of the joint tissue were observed by hematoxylin-eosin staining. The content of malondialdehyde(MDA), glutathione(GSH), and Fe~(2+) and the activity of superoxide dismutase(SOD) in the joint tissue were measured by biochemical colorimetry. RT-PCR was performed to determine the mRNA levels of nuclear factor erythroid 2-related factor 2(Nrf2), glutathione peroxidase 4(GPX4), and acyl-CoA synthetase long chain family member 4(ACSL4) in the joint tissue. Western blot was employed to determine the protein levels of Nrf2, Kelch-like ECH-associated protein 1(Keap1), heme oxygenase-1(HO-1), NAD(P)H quinone oxidoreductase 1(NQO1), SOD2, GPX4, and ACSL4 in the joint tissue. The results showed that the treatment with Tripterygium glycosides(7.5 mg·kg~(-1)) and total triterpenes of Euphorbium(32, 64, and 128 mg·kg~(-1)) alleviated the swelling degree of bilateral hind limbs, decreased the arthritis score, reduced joint tissue lesions and the content of MDA and Fe~(2+) in the joint tissue, and increased GSH content and SOD activity. Furthermore, the interventions up-regulated the protein and mRNA levels of Nrf2 and GPX4, down-regulated the protein and mRNA levels of ACSL4, and up-regulated the protein levels of Keap1, NQO1, HO-1, and SOD2 in the Nrf2/HO-1/GPX4. In summary, the total triterpenes of Euphorbium can treat RA by inhibiting lipid peroxidation and abnormal ferroptosis, which may involve the Nrf2/HO-1/GPX4 signaling pathway.

The Preventive Mechanism of Anserine on Tert-Butyl Hydroperoxide-Induced Liver Injury in L-02 Cells via Regulating the Keap1-Nrf2 and JNK-Caspase-3 Signaling Pathways.

Anserine is a naturally occurring histidine dipeptide with significant antioxidant activities. This study aimed to investigate the preventive mechanism of anserine on tert-butyl hydroperoxide (TBHP)-induced liver damage in a normal human liver cell line (L-02 cells). The L-02 cells were pretreated with anserine (10, 20, and 40 mmol/L) and then induced with 400 μmol/L of TBHP for 4 h. The results showed that the survival rates of L-02 cells and the contents of GSH were significantly increased with the pretreatment of anserine; the activities of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in the extracellular fluid were sharply decreased; and the formation of reactive oxygen species (ROS), nuclear fragmentation, and apoptosis were significantly inhibited. In addition, anserine could bind to the Kelch domain of Kelch-like ECH-associated protein 1 (Keap1) with a binding force of -7.2 kcal/mol; the protein expressions of nuclear factor-erythroid 2-related factor-2 (Nrf2), quinone oxidoreductase 1 (NQO1), heme oxygenase-1 (HO-1), and Bcl-2 were upregulated by anserine in TBHP-induced L-02 cells, with the downregulation of p-JNK and caspase-3. In conclusion, anserine might alleviated liver injury in L-02 cells via regulating related proteins in the Keap1-Nrf2 and JNK-Caspase-3 signaling pathways.

Role of the Nrf2 Signaling Pathway in Ovarian Aging: Potential Mechanism and Protective Strategies.

The ovary holds a significant role as a reproductive endocrine organ in women, and its aging process bears implications such as menopause, decreased fertility, and long-term health risks including osteoporosis, cardiovascular disorders, and cognitive decline. The phenomenon of oxidative stress is tightly linked to the aging metabolic processes. More and more studies have demonstrated that oxidative stress impacts both physiologic and pathologic ovarian aging, and the nuclear factor erythroid 2-related factor 2 (Nrf2) pathway plays a crucial role in regulating the antioxidant response. Furthermore, various therapeutic approaches have been identified to ameliorate ovarian aging by modulating the Nrf2 pathway. This review summarizes the important role of the Nrf2/ Kelch-like ECH-associated protein 1 (Keap1) signaling pathway in regulating oxidative stress and influencing ovarian aging. Additionally, it highlights the therapeutic strategies aimed at targeting the Nrf2/Keap1 pathway.

FSP1 confers ferroptosis resistance in KEAP1 mutant non-small cell lung carcinoma in NRF2-dependent and -independent manner

Ferroptosis, a type of cell death induced by lipid peroxidation, has emerged as a novel anti-cancer strategy. Cancer cells frequently acquire resistance to ferroptosis. However, the underlying mechanisms are poorly understood. To address this issue, we conducted a thorough investigation of the genomic and transcriptomic data derived from hundreds of human cancer cell lines and primary tissue samples, with a particular focus on non-small cell lung carcinoma (NSCLC). It was observed that mutations in Kelch-like ECH-associated protein 1 (KEAP1) and subsequent nuclear factor erythroid 2-related factor 2 (NRF2, also known as NFE2L2) activation are strongly associated with ferroptosis resistance in NSCLC. Additionally, AIFM2 gene, which encodes ferroptosis suppressor protein 1 (FSP1), was identified as the gene most significantly correlated with ferroptosis resistance, followed by multiple NRF2 targets. We found that inhibition of NRF2 alone was not sufficient to reduce FSP1 protein levels and promote ferroptosis, whereas FSP1 inhibition effectively sensitized KEAP1-mutant NSCLC cells to ferroptosis. Furthermore, we found that combined inhibition of FSP1 and NRF2 induced ferroptosis more intensely. Our findings imply that FSP1 is a crucial suppressor of ferroptosis whose expression is partially dependent on NRF2 and that synergistically targeting both FSP1 and NRF2 may be a promising strategy for overcoming ferroptosis resistance in cancer.

Mefenamic Acid-Upregulated Nrf2/SQSTM1 Protects Hepatocytes against Oxidative Stress-Induced Cell Damage.

Mefenamic acid (MFA) is a commonly prescribed non-steroidal anti-inflammatory drug (NSAID) with anti-inflammatory and analgesic properties. MFA is known to have potent antioxidant properties and a neuroprotective effect against oxidative stress. However, its impact on the liver is unclear. This study aimed to elucidate the antioxidative effects of MFA and their underlying mechanisms. We observed that MFA treatment upregulated the nuclear factor erythroid 2-related factor 2 (Nrf2) pathway. Treatment with various anthranilic acid derivative-class NSAIDs, including MFA, increased the expression of sequestosome 1 (SQSTM1) in HepG2 cells. MFA disrupted the interaction between Kelch-like ECH-associated protein 1 (Keap1) and Nrf2, activating the Nrf2 signaling pathway. SQTM1 knockdown experiments revealed that the effect of MFA on the Nrf2 pathway was masked in the absence of SQSTM1. To assess the cytoprotective effect of MFA, we employed tert-Butyl hydroperoxide (tBHP) as a ROS inducer. Notably, MFA exhibited a protective effect against tBHP-induced cytotoxicity in HepG2 cells. This cytoprotective effect was abolished when SQSTM1 was knocked down, suggesting the involvement of SQSTM1 in mediating the protective effect of MFA against tBHP-induced toxicity. In conclusion, this study demonstrated that MFA exhibits cytoprotective effects by upregulating SQSTM1 and activating the Nrf2 pathway. These findings improve our understanding of the pharmacological actions of MFA and highlight its potential as a therapeutic agent for oxidative stress-related conditions.

Muscle p62 stimulates the expression of antioxidant proteins alleviating cancer cachexia.

Oxidative stress plays an important role in skeletal muscle atrophy during cancer cachexia, and more glycolytic muscles are preferentially affected. Sequestosome1/SQSTM1 (i.e., p62), particularly when phosphorylated at Ser 349 (Ser 351 in mice), competitively binds to the Kelch-like ECH-associated protein 1 (Keap1) activating Nuclear factor erythroid 2-related factor 2 (Nrf2). Nrf2 then stimulates the transcription of antioxidant/electrophile-responsive elements in target genes. However, a potential role for p62 in the protection of muscle wasting in cachexia remains to be determined. Here, using the well-established cachexia-inducing model of Lewis Lung Carcinoma (LLC) in mice we demonstrate higher expression of antioxidant proteins (i.e., NQO1, HO-1, GSTM1, CuZnSOD, MnSOD, and EcSOD) in the more oxidative and cachexia resistant soleus muscle than in the more glycolytic and cachexia prone extensor digitorum longus muscle. This was accompanied by higher p62 (total and phosphorylated) and nuclear Nrf2 levels in the soleus, which were paralleled by higher expression of proteins known to either phosphorylate or promote p62 phosphorylation (i.e., NBR1, CK1, PKCδ, and TAK1). Muscle-specific p62 gain-of-function (i.e., in p62 mTg mice) activated Nrf2 nuclear translocation and increased the expression of multiple antioxidant proteins (i.e., CuZnSOD, MnSOD, EcSOD, NQO1, and GSTM1) in glycolytic muscles. Interestingly, skeletal muscle Nrf2 haplodeficiency blunted the increases of most of these proteins (i.e., CuZnSOD, EcSOD, and NQO1) suggesting that muscle p62 stimulates antioxidant protein expression also via additional, yet to be determined mechanisms. Of note, p62 gain-of-function mitigated glycolytic muscle wasting in LLC-affected mice. Collectively, our findings identify skeletal muscle p62 as a potential therapeutic target for cancer cachexia.

TMED2 Induces Cisplatin Resistance in Breast Cancer via Targeting the KEAP1-Nrf2 Pathway.

Cisplatin is the first-line treatment for breast cancer, but it faces challenges of drug resistance. This study investigated new molecular mechanisms underlying cisplatin resistance in breast cancer. We analyzed sequencing data from the TCGA database to identify potential associations between transmembrane emp24 protein transport domain containing 2 (TMED2) and breast cancer. Western blotting, real-time PCR, CCK-8, and TUNEL assays were used to measure the effects and molecular mechanism of TMED2 on cisplatin resistance in MCF-7 and MDA-MB-231 cell lines. TMED2 was overexpressed in breast cancer and associated with poor prognosis. TMED2 increased cisplatin resistance in breast cancer cells in vitro via promoting ubiquitination of Kelch-like ECH-associated protein 1 (KEAP1), relieving inhibition of KEAP1 on nuclear factor erythroid 2-related factor 2 (Nrf2), and increasing expression of downstream drug resistance related genes, such as heme oxygenase 1 (HO-1) and NAD (P) H quinone oxidoreductase 1 (NQO1). We identified a new molecular mechanism by which TMED2 affects cisplatin resistance in breast cancer. Our results provide theoretical guidance for future clinical applications.

Effects of dietary eicosapentaenoic acid on the growth performance, fatty acid profile, immunity and heat tolerance of juvenile abalone Haliotis discus hannai Ino

A 93-day growth trial was performed to evaluate the influences of graded dietary EPA levels (0.03% (control group), 0.24%, 0.63%, 1.07%, 1.72% and 2.25%) on the growth performance, immunity and heat tolerance of juvenile abalone Haliotis discus hannai with a mean initial weight of 4.15 ± 0.18 g. After the feeding trial, the findings found that the highest weight gain rate (WGR) belonged to the groups fed with 0.63% ∼ 1.72% of dietary EPA (P < 0.05). The lowest feed conversion ratio was distinguished in the groups with 0.63% ∼ 1.07% of dietary EPA (P < 0.05). Also, EPA supplementation in abalone feed increased the content of crude lipid in the soft body and the activity of lipase in the intestine (P < 0.05). The total antioxidative capacity increased when dietary EPA levels rose from 0.03% to 1.07%, and then decreased (P < 0.05). Furthermore, compared to the control group, the 0.24% and 0.24% ∼ 0.63% of dietary EPA significantly down-regulated the mRNA levels of IκB kinase and kelch-like ECH-associated protein 1, respectively (P < 0.05). However, nuclear factor-κB and interleukin 1 receptor-associated kinase 4 genes expression in the digestive gland were significantly up-regulated by the use of 1.72% and 2.25% of EPA in diet (P < 0.05). The mRNA levels of tumor necrosis factor α and interleukin 16 were significantly up-regulated by the 2.25% of dietary EPA (P < 0.05). The 1.72% of dietary EPA significantly increased the falling rate of abalone after the heat stress compared with the control (P < 0.05). In conclusion, the optimal dietary EPA requirement of juvenile abalone was determined as 1.36% based on quadratic regression analysis against WGR. Moderate dietary EPA content (0.63% ∼ 1.07%) improved the immunity and heat resistance of abalone. Excessive dietary EPA content (1.72% ∼ 2.25%) decreased the growth, meanwhile, increasing the oxidative stress, inflammation and susceptibility of abalone to heat stress.

Metabolic barriers in non-small cell lung cancer with LKB1 and/or KEAP1 mutations for immunotherapeutic strategies.

Currently, immune checkpoint inhibitors (ICIs) are widely considered the standard initial treatment for advanced non-small cell lung cancer (NSCLC) when there are no targetable driver oncogenic alternations. NSCLC tumors that have two alterations in tumor suppressor genes, such as liver kinase B1 (LKB1) and/or Kelch-like ECH-associated protein 1 (KEAP1), have been found to exhibit reduced responsiveness to these therapeutic strategies, as revealed by multiomics analyses identifying immunosuppressed phenotypes. Recent advancements in various biological approaches have gradually unveiled the molecular mechanisms underlying intrinsic reprogrammed metabolism in tumor cells, which contribute to the evasion of immune responses by the tumor. Notably, metabolic alterations in glycolysis and glutaminolysis have a significant impact on tumor aggressiveness and the remodeling of the tumor microenvironment. Since glucose and glutamine are essential for the proliferation and activation of effector T cells, heightened consumption of these nutrients by tumor cells results in immunosuppression and resistance to ICI therapies. This review provides a comprehensive summary of the clinical efficacies of current therapeutic strategies against NSCLC harboring LKB1 and/or KEAP1 mutations, along with the metabolic alterations in glycolysis and glutaminolysis observed in these cancer cells. Furthermore, ongoing trials targeting these metabolic alterations are discussed as potential approaches to overcome the extremely poor prognosis associated with this type of cancer.

Echinochrome Ameliorates Physiological, Immunological, and Histopathological Alterations Induced by Ovalbumin in Asthmatic Mice by Modulating the Keap1/Nrf2 Signaling Pathway.

Asthma is a persistent inflammatory disease of the bronchi characterized by oxidative stress, airway remodeling, and inflammation. Echinochrome (Ech) is a dark-red pigment with antioxidant and anti-inflammatory activities. In this research, we aimed to investigate the effects of Ech against asthma-induced inflammation, oxidative stress, and histopathological alterations in the spleen, liver, and kidney in mice. Mice were divided into four groups (n = 8 for each): control, asthmatic, and asthmatic mice treated intraperitoneally with 0.1 and 1 mg/kg of Ech. In vitro, findings confirmed the antioxidant and anti-inflammatory activities of Ech. Ech showed antiasthmatic effects by lowering the serum levels of immunoglobulin E (IgE), interleukin 4 (IL-4), and interleukin 1β (IL-1β). It attenuated oxidative stress by lowering malondialdehyde (MDA) and nitric oxide (NO) contents and increasing reduced glutathione (GSH), superoxide dismutase (SOD), glutathione-s-transferase (GST), and catalase (CAT) in the liver, spleen, and kidney. Moreover, it protected asthma-induced kidney and liver functions by increasing total protein and albumin and decreasing aspartate aminotransferase (AST), alanine aminotransferase (ALT), creatinine, urea, and uric acid levels. Additionally, it ameliorated histopathological abnormalities in the lung, liver, spleen, and kidney. Additionally, molecular docking studies were used to examine the interactions between Ech and Kelch-like ECH-associated protein 1 (Keap1). PCR and Western blot analyses confirmed the association of Ech with Keap1 and, consequently, the regulatory role of Ech in the Keap1-(nuclear factor erythroid 2-related factor 2) Nrf2 signaling pathway in the liver, spleen, and kidney. According to our findings, Ech prevented asthma and its complications in the spleen, liver, and kidney. Inhibition of inflammation and oxidative stress are two of echinochrome's therapeutic actions in managing asthma by modulating the Keap1/Nrf2 signaling pathway.

MiR-200a attenuated oxidative stress, inflammation, and apoptosis in dextran sulfate sodium-induced colitis through activation of Nrf2.

Oxidative stress and inflammatory responses are critical factors in ulcerative colitis disease pathogenesis. Nuclear factor erythroid 2-related factor 2 (Nrf2) modulates oxidative stress and suppresses inflammatory responses, and the protective benefits of Nrf2 activation have been associated with the therapy of ulcerative colitis. MicroRNA-200a (miR-200a) could target Kelch-like ECH-associated protein 1 (Keap1) and activate the Nrf2-regulated antioxidant pathway. Nevertheless, whether miR-200a modulates the Keap1/Nrf2 pathway in dextran sulfate sodium (DSS)-induced colonic damage is unknown. Here, our research intends to examine the impact of miR-200a in the model of DSS-induced colitis. Prior to DSS intervention, we overexpressed miR-200a in mice for four weeks using an adeno-associated viral (AAV) vector to address this problem. ELISA detected the concentration of inflammation-related cytokines. The genes involved in inflammatory reactions and oxidative stress were identified using quantitative reverse transcription-polymerase chain reaction (qRT-PCR), western blot, and immunofluorescence. Moreover, we applied siRNAs to weakened Nrf2 expression to confirm the hypothesis that miR-200a provided protection via Nrf2. The present study discovered miR-200a down-regulation, excessive inflammatory activation, enterocyte apoptosis, colonic dysfunction, and Keap1/Nrf2 antioxidant pathway inactivation in mouse colitis and NCM460 cells under DSS induction. However, our data demonstrated that miR-200a overexpression represses Keap1 and activates the Nrf2 antioxidant pathway, thereby alleviating these adverse alterations in animal and cellular models. Significantly, following Nrf2 deficiency, we failed to observe the protective benefits of miR-200a against colonic damage. Taken together, through activating the Keap1/Nrf2 signaling pathway, miR-200a protected against DSS-induced colonic damage. These studies offer an innovative therapeutic approach for ulcerative colitis.

Intermittent cyclic mechanical compression promotes endplate chondrocytes degeneration by disturbing Nrf2/PINK1 signaling pathway-dependent mitophagy.

An abnormal mechanical load is a pivotal inducer of endplate cartilage degeneration, which subsequently promotes intervertebral disc degeneration. Our previous study indicated that intermittent cyclic mechanical compression (ICMC) promotes endplate chondrocyte degeneration, but the mechanism underlying this effect is unclear. In this study, we investigated PTEN-induced kinase 1(PINK1) dependent mitophagy during ICMC-induced endplate chondrocyte degeneration. Furthermore, we determined whether NF-E2-related factor 2 (Nrf2) activation correlated with PINK1-dependent mitophagy regulation and increased oxidation resistance of endplate chondrocytes under ICMC application. First, we generated a mechanical compression-induced endplate chondrocyte degeneration model in vitro and in vivo. ICMC was found to promote endplate chondrocyte extracellular matrix degradation. PINK1-mediated mitophagy was suppressed in the ICMC-stimulated endplate chondrocytes, while increased mitochondrial reactive oxygen species generation suggested that mitophagy is involved in the protective effect of mechanical strain on endplate chondrocytes. Moreover, Nrf2 expression, interaction with Kelch-like ECH-associated protein (Keap1), and nuclear translocation were inhibited by ICMC. Nrf2 overexpression inhibited reactive oxygen species production and reversed ICMC-induced endplate chondrocyte degeneration. Transfection with PINK1 shRNA abolished this effect and partially blocked Nrf2-induced mitophagy. Our findings suggested that ICMC could inhibit the Nrf2/PINK1 signaling pathway to reduce the mitophagy levels which significantly promote oxidative stress and thereby endplate chondrocyte degeneration. Therapeutic regulation of the Nrf2/PINK1 signaling pathway may be an efficient anabolic strategy for inhibiting this process.

Protective Effects and Mechanisms of Pectolinarin against H2O2-Induced Oxidative Stress in SH-SY5Y Neuronal Cells.

This study aims to investigate the protective effects and mechanisms of pectolinarin against oxidative stress-induced cell damage in SH-SY5Y cells. Neurodegenerative diseases-such as Alzheimer's disease-are potentially associated with oxidative stress, which causes excessive production of reactive oxygen species (ROS) that damage DNA and proteins in neuronal cells. The results of this study demonstrate that pectolinarin can scavenge hydroxyl and nitric oxide radicals in a concentration-dependent manner. Moreover, pectolinarin significantly increased cell viability while reducing ROS production and LDH release in the hydrogen peroxide (H2O2)-induced control group. Additionally, Pectolinarin recovered protein expression from H2O2-altered levels back to close-to-normal SH-SY5Y cell levels for components of the oxidative stress, inflammation, and apoptosis pathways-such as nuclear factor erythroid 2-related factor 2 (Nrf2), kelch-like ECH-associated protein (Keap1), anti-heme oxygenase 1 (HO-1), inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), interleukin-1β (IL-1β), B-cell lympho-ma-2 (Bcl-2) protein, and Bcl-2-associated X protein (Bax). These findings suggest that pectolinarin has the potential to be used as a plant material for functional foods to be applied in the treatment of neurodegenerative diseases, such as Alzheimer's disease, by mitigating oxidative stress-induced damage to neuronal cells.

TNFAIP2 confers cisplatin resistance in head and neck squamous cell carcinoma via KEAP1/NRF2 signaling

BackgroundDrug resistance limits the treatment effect of cisplatin-based chemotherapy in head and neck squamous cell carcinoma (HNSCC), and the underlying mechanism is not fully understood. The aim of this study was to explore the cause of cisplatin resistance in HNSCC.MethodsWe performed survival and gene set variation analyses based on HNSCC cohorts and identified the critical role of tumor necrosis factor alpha-induced protein 2 (TNFAIP2) in cisplatin-based chemotherapy resistance. Half-maximal inhibitory concentration (IC50) examination, colony formation assays and flow cytometry assays were conducted to examine the role of TNFAIP2 in vitro, while xenograft models in nude mice and 4-nitroquinoline N-oxide (4NQO)-induced HNSCC models in C57BL/6 mice were adopted to verify the effect of TNFAIP2 in vivo. Gene set enrichment analysis (GSEA) and coimmunoprecipitation coupled with mass spectrometry (Co-IP/MS) were performed to determine the mechanism by which TNFAIP2 promotes cisplatin resistance.ResultsHigh expression of TNFAIP2 is associated with a poor prognosis, cisplatin resistance, and low reactive oxygen species (ROS) levels in HNSCC. Specifically, it protects cancer cells from cisplatin-induced apoptosis by inhibiting ROS-mediated c-JUN N-terminal kinase (JNK) phosphorylation. Mechanistically, the DLG motif contained in TNFAIP2 competes with nuclear factor-erythroid 2-related factor 2 (NRF2) by directly binding to the Kelch domain of Kelch-like ECH-associated protein 1 (KEAP1), which prevents NRF2 from undergoing ubiquitin proteasome-mediated degradation. This results in the accumulation of NRF2 and confers cisplatin resistance. Positive correlations between TNFAIP2 protein levels and NRF2 as well as its downstream target genes were validated in HNSCC specimens. Moreover, the small interfering RNA (siRNA) targeting TNFAIP2 significantly enhanced the cisplatin treatment effect in a 4NQO-induced HNSCC mouse model.ConclusionsOur results reveal the antioxidant and cisplatin resistance-regulating roles of the TNFAIP2/KEAP1/NRF2/JNK axis in HNSCC, suggesting that TNFAIP2 might be a potential target in improving the cisplatin treatment effect, particularly for patients with cisplatin resistance.

β-Endorphin (an endogenous opioid) inhibits inflammation, oxidative stress and apoptosis via Nrf-2 in asthmatic murine model

Asthma, a chronic respiratory disease is characterized by airway inflammation, remodelling, airflow limitation and hyperresponsiveness. At present, it is considered as an umbrella diagnosis consisting several variable clinical presentations (phenotypes) and distinct pathophysiological mechanisms (endotypes). Recent evidence suggests that oxidative stress participates in airway inflammation and remodelling in chronic asthma. Opioids resembled by group of regulatory peptides have proven to act as an immunomodulator. β-Endorphin a natural and potent endogenous morphine produced in the anterior pituitary gland play role in pain modulation. Therapeutic strategy of many opioids including β-Endorphin as an anti‑inflammatory and antioxidative agent has not been yet explored despite its promising analgesic effects. This is the first study to reveal the role of β-Endorphin in regulating airway inflammation, cellular apoptosis, and oxidative stress via Nrf-2 in an experimental asthmatic model. Asthma was generated in balb/c mice by sensitizing with 1% Toulene Diisocyanate on day 0, 7, 14 and 21 and challenging with 2.5% Toulene Diisocyanate from day 22 to 51 (on every alternate day) through intranasal route. β-Endorphin (5 µg/kg) was administered through the nasal route 1 h prior to sensitization and challenge. The effect of β-Endorphin on pulmonary inflammation and redox status along with parameters of oxidative stress were evaluated. We found that pre-treatment of β-Endorphin significantly reduced inflammatory infiltration in lung tissue and cell counts in bronchoalveolar lavage fluid. Also, pre-treatment of β-Endorphin reduced reactive oxygen species, Myeloperoxidase, Nitric Oxide, Protein and protein carbonylation, Glutathione Reductase, Malondialdehyde, IFN-γ, and TNF-α. Reversely, β-Endorphin significantly increased Superoxide dismutase, Catalase, glutathione, Glutathione-S-Transferase, and activation of NF-E2-related factor 2 (Nrf-2) via Kelch-like ECH-associated protein 1 (Keap1), independent pathway in the lung restoring architectural alveolar and bronchial changes. The present findings reveal the therapeutic potency of β-END in regulating asthma by Keap-1 independent regulation of Nrf-2 activity. The present findings reveal the therapeutic potency of β-Endorphin in regulating asthma.

Follicular fluid derived exosomal miR-4449 regulates cell proliferation and oxidative stress by targeting KEAP1 in human granulosa cell lines KGN and COV434

Polycystic ovary syndrome (PCOS) is characterized by ovulatory dysfunction, hyperandrogenism, and polycystic ovary morphology, affecting more and more women of reproductive age. Our study aimed to explore the molecular mechanism and effect of exosomal miR-4449 on granulosa cells (GCs). Two immortalized human ovarian granulosa cells (KGN and COV434 cells) were used for in vitro functional studies. Our study found that follicular fluid (FF) derived exosomal miR-4449 was significantly decreased in women with PCOS compared with the control patients. And exosomal miR-4449 could alleviate GCs oxidative stress (OS) and promote GCs proliferation, while the opposite trend was observed after inhibiting the expression of miR-4449. In addition, we demonstrated that Kelch-like ECH-associated protein 1(KEAP1) was a direct target of miR-4449 through dual-luciferase reporter assay, and the expression patterns of KEAP1 and miR-4449 in PCOS FF-derived exosomes were exactly opposite. In addition, KEAP1/NRF2 signaling pathway may play an important role in GCs proliferation and OS. Our results demonstrated that the decreased FF-derived exosomal miR-4449 expression in PCOS might aggravate the OS of GCs and inhibit GCs proliferation via KEAP1/NRF2 signaling pathway. Exosomal miR-4449 might be a potential biomarker for the diagnosis of PCOS. Our study contributes to a new understanding of the pathogenesis of PCOS.