[Essential oil of Cinnamomum camphora mitigates depression-like behavior in mice by regulating Nrf2/HO-1 pathway and inhibiting inflammatory cytokines].

This study aims to investigate the essential oil(EOL) of Cinnamomum camphora regarding its anti-depression effect and mechanism in regulating inflammatory cytokines and the nuclear factor erythroid 2-related factor 2(Nrf2)/heme oxygenase-1(HO-1) pathway. A mouse model of depression was established by intraperitoneal injection of lipopolysaccharide(LPS). Open field, elevated plus maze, and forced swimming tests were carried out to examine mouse behaviors. Western blot and qRT-PCR were employed to determine the expression of proteins and genes in the Nrf2/HO-1 pathway in the hippocampus. The levels of tumor necrosis factor(TNF)-α, interleukin(IL)-6, and IL-1ß in the serum were measured by enzyme-linked immunosorbent assay(ELISA). The changes of apoptosis in mouse brain were detected by Tunel staining. Compared with the blank control group, the model group showed shortened distance travelled and time spent in the central zone and reduced number of entries in the central zone in the open field test. In the elevated plus maze test, the model group showed reduced open arm time(OT%) and open arm entries(OE%). In the force swimming test, the model group showed extended duration of immobility compared with the blank control group. Compared with the model group, the treatment with EOL significantly increased the distance travelled and time spent in the central zone and increased the number of entries in the central zone in the open field test. In addition, EOL significantly increased the OT% and OE% in the elevated plus maze and shor-tened the immobility duration in the forced swimming test. The model group showed lower expression levels of Nrf2 and HO-1 and hig-her levels of TNF-α, IL-6, and IL-1ß than the blank control group. Compared with the model group, the treatment with EOL up-regulated the expression levels of Nrf2 and HO-1 and lowered the levels of TNF-α, IL-6, and IL-1ß. The Tunel staining results showed that the apoptosis rate in the brain tissue of mice decreased significantly after the treatment with EOL. To sum up, EOL can mitigate the depression-like behaviors of mice by up-regulating the expression of Nrf2 and HO-1 and preventing hippocampal inflammatory damage. The findings provide empirical support for the application of EOL and aromatherapy in the treatment of depression.

[Fuyu Decoction ameliorates myocardial fibrosis in rat model of heart failure by regulating Nrf2/GPX4-mediated ferroptosis].

This study aims to investigate the effect and mechanism of Fuyu Decoction(FYD) in the treatment of myocardial fibrosis in the rat model of heart failure(HF). Sixty Wistar rats were randomized into a modeling group(n=50) and a sham group(n=10). A post-myocardial infarction HF model was established by ligating the left anterior descending coronary artery in rats. The successfully modeled rats were assigned into model, low-dose(2.5 g·kg~(-1)) FYD(FYD-L), high-dose(5.0 g·kg~(-1)) FYD(FYD-H), and FYD+Nrf2 inhibitor(ML385, 30 mg·kg~(-1)) groups(n=10). FYD was administrated by gavage and ML385 by intraperitoneal injection. The rats in the sham and model groups were administrated with equal amounts of normal saline by gavage. After 8 weeks of intervention, the cardiac function indicators were measured, and the myocardial tissue morphology and collagen deposition were observed. The positive expression of collagens Ⅰ and Ⅲ, apoptosis, and oxidative stress were examined, and the levels of Fe~(2+) and reactive oxygen species(ROS) were determined. The protein levels of nuclear factor erythroid 2-related factor 2(Nrf2), solute carrier family 7 member 11(SLC7A11), glutathione peroxidase 4(GPX4), and acyl-coenzyme A synthase long chain family member 4(ACSL4) in the myocardial tissue were determined. Compared with sham group, the model group showed decreased left ventricular ejection fraction(LVEF) and left ventricular fractional shortening(LVFS), increased left ventricular end internal dimension in systole(LVIDs), left ventricular internal diameter in diastole(LVIDd), and myocardial collagen deposition, positive expression of collagens Ⅰ and Ⅲ, elevated apoptosis rate and malondialdehyde(MDA), Fe~(2+), and ROS levels, lowered superoxide dismutase(SOD) and glutathione peroxidase(GSH) levels, down-regulated protein levels of Nrf2, SLC7A11, and GPX4, and up-regulated protein level of ACSL4. Compared with the model group, the above indicators were restored by FYD. Moreover, ML385 reversed the protective effect of FYD on myocardial fibrosis in HF rats. In conclusion, FYD can inhibit ferroptosis by activating the Nrf2/GPX4 pathway, thereby ameliorating myocardial fibrosis in HF rats.

[Ginsenoside Re regulates mitochondrial biogenesis through Nrf2/HO-1/PGC-1α pathway to reduce hypoxia/reoxygenation injury in H9c2 cells].

This article explored the mechanism by which ginsenoside Re reduces hypoxia/reoxygenation(H/R) injury in H9c2 cells by regulating mitochondrial biogenesis through nuclear factor E2-related factor 2(Nrf2)/heme oxygenase-1(HO-1)/peroxisome prolife-rator-activated receptor gamma coactivator-1α(PGC-1α) pathway. In this study, H9c2 cells were cultured in hypoxia for 4 hours and then reoxygenated for 2 hours to construct a cardiomyocyte H/R injury model. After ginsenoside Re pre-administration intervention, cell activity, superoxide dismutase(SOD) activity, malondialdehyde(MDA) content, intracellular reactive oxygen species(Cyto-ROS), and intramitochondrial reactive oxygen species(Mito-ROS) levels were detected to evaluate the protective effect of ginsenoside Re on H/R injury of H9c2 cells by resisting oxidative stress. Secondly, fluorescent probes were used to detect changes in mitochondrial membrane potential(ΔΨ_m) and mitochondrial membrane permeability open pore(mPTP), and immunofluorescence was used to detect the expression level of TOM20 to study the protective effect of ginsenoside Re on mitochondria. Western blot was further used to detect the protein expression levels of caspase-3, cleaved caspase-3, Cyto C, Nrf2, HO-1, and PGC-1α to explore the specific mechanism by which ginsenoside Re protected mitochondria against oxidative stress and reduced H/R injury. Compared with the model group, ginse-noside Re effectively reduced the H/R injury oxidative stress response of H9c2 cells, increased SOD activity, reduced MDA content, and decreased Cyto-ROS and Mito-ROS levels in cells. Ginsenoside Re showed a good protective effect on mitochondria by increasing ΔΨ_m, reducing mPTP, and increasing TOM20 expression. Further studies showed that ginsenoside Re promoted the expression of Nrf2, HO-1, and PGC-1α proteins, and reduced the activation of the apoptosis-related regulatory factor caspase-3 to cleaved caspase-3 and the expression of Cyto C protein. In summary, ginsenoside Re can significantly reduce I/R injury in H9c2 cells. The specific mechanism is related to the promotion of mitochondrial biogenesis through the Nrf2/HO-1/PGC-1α pathway, thereby increasing the number of mitochondria, improving mitochondrial function, enhancing the ability of cells to resist oxidative stress, and alleviating cell apoptosis.

[Tanshinone â ¡_A inhibits ferroptosis via Nrf2 signaling pathway to protect liver in rats of non-alcoholic fatty liver disease].

This study investigated the protective effect of tanshinone Ⅱ_A(TSⅡ_A) on the liver in the rat model of non-alcoholic fatty liver disease(NAFLD) and the mechanism of TSⅡ_A in regulating ferroptosis via the nuclear factor E2-related factor 2(Nrf2) signaling pathway. The rat model of NAFLD was established with a high-fat diet for 12 weeks. The successfully modeled rats were assigned into model group, low-and high-dose TSⅡ_A groups, and inhibitor group, and normal control group was set. Enzyme-linked immunosorbent assay was employed to determine the content of superoxide dismutase(SOD) and malondialdehyde(MDA) in the serum of rats in each group. A biochemical analyzer was used to measure the content of aspartate aminotransferase(AST), alaninl aminotransferase(ALT), total cholesterol(TC), and triglycerides(TG). Hematoxylin-eosin(HE) staining was used to detect pathological damage in liver tissue. Terminal-deoxynucleoitidyl transferase-mediated nick end labeling(TUNEL) was employed to examine the apoptosis of the liver tissue. Oil red O staining, MitoSOX staining, and Prussian blue staining were conducted to reveal lipid deposition, the content of reactive oxygen species(ROS), and iron deposition in liver tissue. Western blot was employed to determine the expression of Nrf2, heme oxygenase-1(HO-1), glutathione peroxidase 4(GPX4), ferroptosis suppressor protein 1(FSP1), B cell lymphoma-2(Bcl-2), and Bcl-2 associated X protein(Bax) in the liver tissue. The result showed that TSⅡ_A significantly reduced the content of MDA, AST, ALT, TC, and TG in the serum, increased the activity of SOD, decreased the apoptosis rate, lipid deposition, ROS, and iron deposition in the liver tissue, up-regulated the expression of Nrf2, HO-1, FSP1, GPX, and Bcl-2, and inhibited the expression of Bax in the liver tissue of NAFLD rats. However, ML385 partially reversed the protective effect of TSⅡ_A on the liver tissue. In conclusion, TSⅡ_A could inhibit ferroptosis in the hepatocytes and decrease the ROS and lipid accumulation in the liver tissue of NAFLD rats by activating the Nrf2 signaling pathway.

An insight into role of amino acids as antioxidants via NRF2 activation.

Oxidative stress can affect the protein, lipids, and DNA of the cells and thus, play a crucial role in several pathophysiological conditions. It has already been established that oxidative stress has a close association with inflammation via nuclear factor erythroid 2-related factor 2 (NRF2) signaling pathway. Amino acids are notably the building block of proteins and constitute the major class of nitrogen-containing natural products of medicinal importance. They exhibit a broad spectrum of biological activities, including the ability to activate NRF2, a transcription factor that regulates endogenous antioxidant responses. Moreover, amino acids may act as synergistic antioxidants as part of our dietary supplementations. This has aroused research interest in the NRF2-inducing activity of amino acids. Interestingly, amino acids' activation of NRF2-Kelch-like ECH-associated protein 1 (KEAP1) signaling pathway exerts therapeutic effects in several diseases. Therefore, the present review will discuss the relationship between different amino acids and activation of NRF2-KEAP1 signaling pathway pinning their anti-inflammatory and antioxidant properties. We also discussed amino acids formulations and their applications as therapeutics. This will broaden the prospect of the therapeutic applications of amino acids in a myriad of inflammation and oxidative stress-related diseases. This will provide an insight for designing and developing new chemical entities as NRF2 activators.

Glutathione Supplementation Prevents Neonatal Parenteral Nutrition-Induced Short- and Long-Term Epigenetic and Transcriptional Disruptions of Hepatic H2O2 Metabolism in Guinea Pigs.

The parenteral nutrition (PN) received by premature newborns is contaminated with peroxides that induce global DNA hypermethylation via oxidative stress. Exposure to peroxides could be an important factor in the induction of chronic diseases such as those observed in adults who were born preterm. As endogenous H2O2 is a major regulator of glucose-lipid metabolism, our hypothesis was that early exposure to PN induces permanent epigenetic changes in H2O2 metabolism. Three-day-old guinea pigs were fed orally (ON), PN or glutathione-enriched PN (PN+GSSG). GSSG promotes endogenous peroxide detoxification. After 4 days, half the animals were sacrificed, and the other half were fed ON until 16 weeks of age. The liver was harvested. DNA methylation and mRNA levels were determined for the SOD2, GPx1, GCLC, GSase, Nrf2 and Keap1 genes. PN induced GPx1 hypermethylation and decreased GPx1, GCLC and GSase mRNA. These findings were not observed in PN+GSSG. PN+GSSG induced Nrf2 hypomethylation and increased Nrf2 and SOD2 mRNA. These observations were independent of age. In conclusion, in neonatal guinea pigs, PN induces epigenetic changes, affecting the expression of H2O2 metabolism genes. These changes persist for at least 15 weeks after PN. This disruption may signify a permanent reduction in the capacity to detoxify peroxides.

Integrated network pharmacology and pharmacological investigations to explore the potential mechanism of Ding-Chuan-Tang against chronic obstructive pulmonary disease.

ETHNOPHARMACOLOGICAL RELEVANCE: Ding-Chuan-Tang (Abbreviated as DCT) is frequently prescribed for treatment of respiratory diseases, including chronic obstructive pulmonary disease (COPD), which is characterized by coughing, wheezing, and chest tightness in traditional Chinese medicine (TCM). However, the potential mechanism of DCT has not been investigated. AIM OF STUDY: The aim of the study is to explore the efficiency of DCT in the treatment of COPD in vivo and in vitro, and to illustrate the possible mechanism against COPD. METHODS: COPD model was induced by exposure of mice to cigarette smoke (CS) for 16 weeks. Enzyme-linked immunosorbent assay (ELISA), immunofluorescence assay, Western blot, etc., were used to explore the efficiency and mechanisms of DCT. Network pharmacology analysis, including Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis, etc., was performed to explore the potential targets in the treatment of DCT on COPD. RESULTS: DCT significantly alleviated pulmonary pathological changes in mouse COPD model, and inhibited inflammatory response induced by CS and LPS in vivo and in vitro. Network pharmacology analysis suggested that DCT alleviated COPD via inhibiting inflammation by regulating PI3K-AKT pathway. In cell-based models, DCT suppressed the phosphorylation of PI3K and AKT, which further regulated its downstream targets Nrf2 and NF-κB, and inhibited inflammatory response. CONCLUSIONS: DCT effectively attenuated COPD in the mouse model induced by CS. The therapeutic mechanism of DCT against COPD was closely associated with the regulation of PI3K-AKT pathway and its downstream transcription factors, Nrf2 and NF-κB.

Effect of electroacupuncture on brain-gut oxidative stress in Parkinson's disease mice. / ç”µé’ˆå¯¹å¸•é‡‘æ£®ç— å°é¼ è„‘è‚ æ°§åŒ–åº”æ¿€çš„å½±å“.

OBJECTIVES: To observe the effect of electroacupuncture (EA) on behavior, oxidative stress factors in colon and substantia nigra of Parkinson's disease (PD) mice, so as to explore the mechanism of EA in treating PD. METHODS: C57BL/6 mice were randomly divided into blank, model and EA groups, with 12 mice in each group. The PD mouse model was established by continuous gavage of rotenone for 4 weeks. Mice in the EA group received EA (2 Hz/15 Hz) at "Baihui" (GV20), "Quchi" (LI11) and "Zusanli" (ST36) for 20 min, 5 days a week for 2 weeks. After intervention, gait analysis was used to evaluate the motor ability and motor coordination. Ink propulsion rate was used to evaluate the intestinal transport function. The level of reactive oxygen species (ROS) in the colon was detected by flow cytometry. The contents of total protein (TP), malondialdehyde (MDA) and activities of glutathione peroxidase (GSH-Px), superoxide dismutase (SOD) in colon and substantia nigra were detected by ELISA. The expression of nuclear factor E2-related factor 2 (Nrf2) in substantia nigra was detected by immunofluorescence staining. RESULTS: Compared with the blank group, the average speed, step rate, normal step ratio, distance between the front and hind feet, stride length, swing speed and maximum intensity of the maximum contact area of mice in the model group were decreased (P<0.000 1, P<0.01, P<0.001), the maximum change rate of gait was increased (P<0.001) in the model group. The intestinal propulsion rate, the activities of GSH-Px and SOD in the colon and substantia nigra, and the positive expression of Nrf2 in substantia nigra were decreased (P<0.000 1, P<0.01, P<0.05), while the fluorescence intensity of ROS in the colon, the contents of MDA in colon and substantia nigra were increased (P<0.01). Compared with the model group, the average speed, step rate, normal step ratio, distance between the front and hind feet, stride length, swing speed, and maximum intensity of the maximum contact area of the mice in the EA group were increased (P<0.01, P<0.05, P<0.001, P<0.000 1), the maximum change rate of gait was decreased (P<0.01). The intestinal propulsion rate, the activities of GSH-Px and SOD in the colon and substantia nigra, the positive expression of Nrf2 in substantia nigra were increased (P<0.001, P<0.05, P<0.000 1), while the ROS fluorescence intensity in the colon, the MDA contents in the colon and substantia nigra were decreased (P<0.01). CONCLUSIONS: EA can improve the movement disorder, gait disorder and intestinal motor function of PD mice, and protect dopaminergic neurons from damage, which may be related to its effect in antagonistic brain-gut oxidative stress.

Therapeutic potential of salidroside in preserving rat cochlea organ of corti from gentamicin-induced injury through modulation of NRF2 signaling and GSK3ß/NF-κB pathway.

Salidroside (SAL) is a phenol glycoside compound found in plants of the Rhodiola genus which has natural antioxidant and free radical scavenging properties. SAL are able to protect against manganese-induced ototoxicity. However, the molecular mechanism by which SAL reduces levels of reactive oxygen species (ROS) is unclear. Here, we established an in vitro gentamicin (GM) ototoxicity model to observe the protective effect of SAL on GM-induced hair cells (HC) damage. Cochlear explants of postnatal day 4 rats were obtained and randomly divided into six groups: two model groups (treatment with 0.2 mM or 0.4 mM GM for 24 h); two 400 µmol/L SAL-pretreated groups pretreatment with SAL for 3 h followed by GM treatment (0.2 mM or 0.4 mM) for 24 h; 400 µmol/L SAL group (treatment with SAL for 24 h); control group (normal cultured cochlear explants). The protective effects of SAL on GM-induced HC damage, and on mRNA and protein levels of antioxidant enzymes were observed. HC loss occurred after 24 h of GM treatment. Pretreatment with SAL significantly reduced GM-induced OHC loss. In cochlear tissues, mRNA and protein levels of NRF2 and HO-1 were enhanced in the GM alone group compared with the SAL pretreatment GM treatment group. SAL may protect against GM-induced ototoxicity by regulating the antioxidant defense system of cochlear tissues; SAL can activate NRF2/HO-1 signaling, inhibit NF-κB activation, activate AKT, and increase inhibitory phosphorylation of GSK3ß to decrease GSK3 activity, all of which exert antioxidant effects.

Corynoline Suppresses Osteoclastogenesis and Attenuates ROS Activities by Regulating NF-κB/MAPKs and Nrf2 Signaling Pathways.

Declining estrogen production in postmenopausal females causes osteoporosis in which the resorption of bone exceeds the increase in bone formation. Although clinical drugs are currently available for the treatment of osteoporosis, sustained medication use is accompanied by serious side effects. Corydalis bungeana Herba, a famous traditional Chinese herb listed in the Chinese Pharmacopoeia Commission, constitutes various traditional Chinese Medicine prescriptions, which date back to thousands of years. One of the primary active components of C. bungeana Turcz. is Corynoline (Cor), a plant isoquinoline alkaloid derived from the Corydalis species, which possesses bone metabolism disease therapeutic potential. The study aimed at exploring the effects as well as mechanisms of Cor on osteoclast formation and bone resorption. TRAcP staining, F-actin belt formation, and pit formation were employed for assessing the osteoclast function. Western blot, qPCR, network pharmacology, and docking analyses were used for analyzing the expression of osteoclast-associated genes and related signaling pathways. The study focused on investigating how Cor affected OVX-induced trabecular bone loss by using a mouse model. Cor could weaken osteoclast formation and function by affecting the biological receptor activators of NF-κB and its ligand at various concentrations. Mechanistically, Cor inhibited the NF-κB activation, and the MAPKs pathway stimulated by RANKL. Besides, Cor enhanced the protein stability of the Nrf2, which effectively abolished the RANKL-stimulated ROS generation. According to an OVX mouse model, Cor functions in restoring bone mass, improving microarchitecture, and reducing the ROS levels in the distal femurs, which corroborated with its in vitro antiosteoclastogenic effect. The present study indicates that Cor may restrain osteoclast formation and bone loss by modulating NF-κB/MAPKs and Nrf2 signaling pathways. Cor was shown to be a potential drug candidate that can be utilized for the treatment of osteoporosis.

Curcumin alleviates cecal oxidative injury in diquat-induced broilers by regulating the Nrf2/ARE pathway and microflora.

This study evaluated the alleviative effect of curcumin (CUR) on the diquat (DQ)-induced cecal injury in broilers. A total of 320 one-day-old Cobb broilers were selected and randomly divided into 4 treatments, namely control, DQ, CUR 100, and CUR150 groups. The control and DQ groups were fed a basal diet, while the CUR 100 and CUR150 groups were fed the basal diet supplemented with 100 and 150 mg/kg CUR, respectively. Each group had 8 replicates, with 10 broilers per replicate. On day 21 of the experiment, 1 broiler was selected from each replicate and intraperitoneally injected 20 mg/kg body weight of DQ for DQ, CUR 100, and CUR 150 groups. Broilers in control group received equivalent volume of saline. Broilers were euthanized 48h postinjection for tissue sampling. The results showed that DQ injection could cause oxidative stress and inflammatory reactions in the cecum, affecting the fatty acid production and flora structure, thus leading to cecum damage. Compared with the DQ group, the activity of superoxide dismutase, the level of interleukin 10, acetic acid, and total volatile fatty, and the abundance of nuclear factor E2-related factor 2, copper and zinc superoxide dismutase and catalase mRNA in the cecal mucosa of broilers in the CUR group increased significantly (P < 0.05). However, the levels of malondialdehyd, reactive oxygen species, tumor necrosis factor-alpha, and the expression of cysteine-aspartic acid protease-3 and tumor necrosis factor-alpha decreased significantly (P < 0.05) in the CUR group. In addition, CUR treatment alleviated the damage to the cecum and restored the flora structure, and Lactobacillus and Lactobacillaceae promoted the alleviative effect of CUR on DQ. In summary, CUR could alleviate the cecal injury caused by DQ-induced oxidative damage and inflammatory reactions by regulating the Nrf2-ARE signaling pathway and intestinal flora, thus protecting the cecum.

Amelioration of nitroglycerin-induced migraine in mice via Wuzhuyu decoction: Inhibition of the MZF1/PGK1 pathway and activation of NRF2 antioxidant response.

ETHNOPHARMACOLOGICAL RELEVANCE: Migraine, a chronic and intricate disorder, manifests as recurrent episodic headaches accompanied by various neurological symptoms. Wuzhuyu Decoction (WZYD) is a traditional Chinese medical formula with promising effects in treating migraines; however, its underlying mechanisms have not yet been clarified. AIM OF STUDY: The study aimed to evaluate WZYD's effectiveness in migraine treatment and investigate the potential mechanism of WZYD's effects on migraine and oxidative stress. MATERIALS AND METHODS: Behavior tests and immunofluorescence assay for the intensity of migraine markers to assess the migraine-relieving effect of WZYD after chronic migraine model induced by nitroglycerin in mice. The impacts of WZYD on oxidative stress-related markers, including reactive oxygen species (ROS), malondialdehyde (MDA), superoxide dismutase (SOD), nuclear factor erythroid 2-related factor 2 (NRF2), heme oxygenase 1 (HO1), and NAD (P)H quinone oxidoreductase 1 (NQO1) in brain tissue were examined. In addition, protein expression or mRNA levels of the MZF1/PGK1 were detected using Western blot or PCR, respectively. Finally, the MZF1 overexpression vector was constructed to the higher level of MZF1. The MZF1/PGK1 signaling pathway expression was evaluated by markers of oxidative stress including NRF2 and others in this series of experiments. RESULTS: Through murine model experimentation, we observed that WZYD effectively alleviates migraine symptoms, signifying its therapeutic efficacy. Mechanistically, WZYD emerges as a potent activator of the NRF2, acting as a robust defense against oxidative stress. In vitro investigations demonstrated that WZYD combats oxidative stress and curbs cell apoptosis induced by these detrimental conditions. Furthermore, by suppressing the transcriptional expression of PGK1, an influential player in the NRF2 pathway, WZYD effectively activates NRF2 signaling. Intriguingly, we have identified MZF1 as the mediator orchestrating the regulation of the PGK1/NRF2 pathway by WZYD. CONCLUSION: The study confirms the effectiveness of WZYD in alleviating migraine symptoms. Mechanistically, WZYD activated the NRF2 signaling pathway; moreover, the action of WZYD involved the down-regulation of PGK1 mediated by MZF1, which promoted the activation of the NRF2 pathway. This study advances our understanding of the intricate mechanisms driving WZYD's efficacy, paving the way for novel treatments in migraine management.

RNA-seq combined network pharmacology reveals that Fu-Gan-Wan (FGW) inhibits liver fibrosis via NF-κB/CCL2/CCR2 and lipid peroxidation via Nrf2/HMOX1 signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Liver fibrosis is a serious complication of liver disease characterized by excessive collagen deposition, without effective therapeutic agents in the clinic. Fu-Gan-Wan (FGW) is an empirical formula used for the clinical treatment of hepatitis and cirrhosis. It has been shown to reverse experimental liver fibrosis. However, its corresponding mechanisms remain unclear. AIM OF THE REVIEW: This study aimed to elucidate the key pathways and target genes of FGW in attenuating liver fibrosis. MATERIALS AND METHODS: The therapeutic effects of different doses of FGW on liver fibrosis were investigated using a 2 mL/kg 15% CCl4-induced mouse model. Then, RNA-seq combined with network pharmacology was used to analyze the key biological processes and signaling pathways underlying the anti-liver fibrosis exertion of FGW. These findings were validated in a TGF-ß1-induced model of activation and proliferation of mouse hepatic stellate cell line JS-1. Finally, the key signaling pathways and molecular targets were validated using animal tissues, and the effect of FGW on tissue lipid peroxidation was additionally observed. RESULTS: We found that 19.5 g/kg FGW significantly down-regulated CCl4-induced elevation of hepatic ALT and AST, decreased collagen deposition, and inhibited the expression of pro-fibrotic factors α-SMA, COL1α1, CTGF, TIMP-1, as well as pro-inflammatory factor TGF-ß1. Additionally, FGW at doses of 62.5, 125, and 250 µg/mL dose-dependently blocked JS-1 proliferation, migration, and activation. Furthermore, RNA-seq identified the NF-κB signaling pathway as a key target molecular pathway for FGW against liver fibrosis, and network pharmacology combined with RNA-seq focused on 11 key genes. Significant changes were identified in CCL2 and HMOX1 by tissue RT-PCR, Western blot, and immunohistochemistry. We further demonstrated that FGW significantly attenuated CCl4-induced increases in p-p65, CCL2, CCR2, and HMOX1, while significantly elevating Nrf2. Finally, FGW significantly suppressed the accumulation of lipid peroxidation products MDA and 4-HNE and reconfigured the oxidation-reduction balance, including promoting the increase of antioxidants GPx, GSH, and SOD, and the decrease of peroxidation products ROS and GSSG. CONCLUSIONS: This study demonstrated that FGW exhibits potential in mitigating CCl4-induced hepatic fibrosis, lipid peroxidation, and iron metabolism disorders in mice. This effect may be mediated through the NF-κB/CCL2/CCR2 and Nrf2/HMOX1 pathways.

Activation of NRF2 by celastrol increases antioxidant functions and prevents the progression of osteoarthritis in mice.

Excessive oxidative stress impairs cartilage matrix metabolism balance, significantly contributing to osteoarthritis (OA) development. Celastrol (CSL), a drug derived from Tripterygium wilfordii, has recognized applications in the treatment of cancer and immune system disorders, yet its antioxidative stress mechanisms in OA remain underexplored. This study aimed to substantiate CSL's chondroprotective effects and unravel its underlying mechanisms. We investigated CSL's impact on chondrocytes under both normal and inflammatory conditions. In vitro, CSL mitigated interleukin (IL)-1ß-induced activation of proteinases and promoted cartilage extracellular matrix (ECM) synthesis. In vivo, intra-articular injection of CSL ameliorated cartilage degeneration and mitigated subchondral bone lesions in OA mice. Mechanistically, it was found that inhibiting nuclear factor erythroid 2-related factor 2 (NRF2) abrogated CSL-mediated antioxidative functions and exacerbated the progression of OA. This study is the first to elucidate the role of CSL in the treatment of OA through the activation of NRF2, offering a novel therapeutic avenue for arthritis therapy.

Moxibustion prevents tripterygium glycoside-induced oligoasthenoteratozoospermia in rats via reduced oxidative stress and modulation of the Nrf2/HO-1 signaling pathway.

Oligoasthenoteratozoospermia (OAT) decreases male fertility, seriously affecting the production of offspring. This study clarified the preventive impact of different moxibustion frequencies on OAT and selected the optimal frequency to elucidate the underlying mechanism. An OAT rat model was constructed by gavage of tripterygium glycosides (TGS) suspension. Daily moxibustion (DM) or alternate-day moxibustion (ADM) was administered on the day of TGS suspension administration. Finally, we selected DM for further study based on sperm quality and DNA fragmentation index, testicular and epididymal morphology, and reproductive hormone level results. Subsequently, the oxidative stress (OS) status was evaluated by observing the OS indices levels; malondialdehyde (MDA), 8-hydroxy-deoxyguanosine (8-OHdG), total antioxidant capacity (T-AOC), and total superoxide dismutase (T-SOD) in testicular tissue using colorimetry and enzyme-linked immunosorbent assay. Furthermore, heme oxygenase 1 (HO-1) and nuclear factor erythropoietin-2-related factor 2 (Nrf2) were evaluated using Western blotting. Immunohistochemistry was employed to locate and assess the expression of HO-1 and Nrf2 protein, while quantitative real-time polymerase chain reaction was utilized to detect their mRNA expression. MDA and 8-OHdG levels decreased following DM treatment, while T-SOD and T-AOC increased, suggesting that DM may prevent TGS-induced OAT in rats by decreasing OS in the testis. Furthermore, protein and mRNA expression of Nrf2 and HO-1 in the testis were elevated, indicating that DM may reduce OS by activating the signaling pathway of Nrf2/HO-1. Therefore, DM could prevent OAT in rats via the Nrf2/HO-1 pathway, thereby presenting a promising therapeutic approach against OAT.

Dietary conjugated linoleic acid downregulates the AlCl3-induced hyperactivation of compensatory and maladaptive signalling in the mouse brain cortex.

Oxidative stress, hyperactivation of compensatory mechanisms (unfolded protein response, UPR; nuclear factor erythroid 2-related factor 2, Nrf2) and the stimulation of maladaptive response (inflammation/apoptosis) are interconnected pathogenic processes occurring during Alzheimer's disease (AD) progression. The neuroprotective ability of dietary Conjugated linoleic acid (CLAmix) in a mouse model of AlCl3-induced AD was recently described but, the effects of AlCl3 or CLAmix intake on these pathogenic processes are still unknown. The effects of dietary AlCl3 or CLAmix - alone and in combination - were examined in the brain cortex of twenty-eight BalbC mice divided into 4 groups (n = 7 each). The neurotoxic effects of AlCl3 were investigated in animals treated for 5 weeks with 100 mg/kg/day (AL). CLAmix supplementation (600 mg/kg bw/day) for 7 weeks (CLA) was aimed at evaluating its modulatory effects on the Nrf2 pathway while its co-treatment with AlCl3 during the last 5 weeks of CLAmix intake (CLA + AL) was used to investigate its neuroprotective ability. Untreated mice were used as controls. In the CLA group, the NADPH oxidase (NOX) activation in the brain cortex was accompanied by the modulation of the Nrf2 pathway. By contrast, in the AL mice, the significant upregulation of oxidative stress markers, compensatory pathways (UPR/Nrf2), proinflammatory cytokines (IL-6, TNFα) and the proapoptotic protein Bax levels were found as compared with control. Notably, in CLA + AL mice, the marked decrease of oxidative stress, UPR/Nrf2 markers and proinflammatory cytokines levels were associated with the significant increase of the antiapoptotic protein Bcl2. The involvement of NOX in the adaptive response elicited by CLAmix along with its protective effects against the onset of several pathogenic processes triggered by AlCl3, broadens the knowledge of the mechanism underlying the pleiotropic activity of Nrf2 activators and sheds new light on their potential therapeutic use against neurodegenerative disorders.

Carnitine functions as an enhancer of NRF2 to inhibit osteoclastogenesis via regulating macrophage polarization in osteoporosis.

Osteoporosis, which manifests as reduced bone mass and deteriorated bone quality, is common in the elderly population. It is characterized by persistent elevation of macrophage-associated inflammation and active osteoclast bone resorption. Currently, the roles of intracellular metabolism in regulating these processes remain unclear. In this study, we initially performed bioinformatics analysis and observed a significant increase in the proportion of M1 macrophages in bone marrow with aging. Further metabolomics analysis demonstrated a notable reduction in the expression of carnitine metabolites in aged macrophages, while carnitine was not detected in osteoclasts. During the differentiation process, osteoclasts took up carnitine synthesized by macrophages to regulate their own activity. Mechanistically, carnitine enhanced the function of Nrf2 by inhibiting the Keap1-Nrf2 interaction, reducing the proteasome-dependent ubiquitination and degradation of Nrf2. In silico molecular ligand docking analysis of the interaction between carnitine and Keap1 showed that carnitine binds to Keap1 to stabilize Nrf2 and enhance its function. In this study, we found that the decrease in carnitine levels in aging macrophages causes overactivation of osteoclasts, ultimately leading to osteoporosis. A decrease in serum carnitine levels in patients with osteoporosis was found to have good diagnostic and predictive value. Moreover, supplementation with carnitine was shown to be effective in the treatment of osteoporosis.

Can curcumin supplementation break the vicious cycle of inflammation, oxidative stress, and uremia in patients undergoing peritoneal dialysis?

BACKGROUND & AIMS: Turmeric (a source of curcumin) is an excellent food to modulate oxidative stress, inflammation, and gut dysbiosis in patients with chronic kidney disease (CKD). However, no studies report the benefits of curcumin in patients undergoing peritoneal dialysis (PD). This study aims to evaluate the effects of curcuminoid supplementation on oxidative stress, inflammatory markers, and uremic toxins originating from gut microbiota in patients with CKD undergoing PD. METHODS: This longitudinal, randomized, single-blind, placebo-controlled trial evaluated 48 patients who were randomized into two groups: Curcumin (three capsules of 500 mg of Curcuma longa extract, with 98.42 % total curcuminoids) or placebo (three capsules of 500 mg of starch) for twelve weeks. In the peripheral blood mononuclear cells (PBMCs), the transcriptional expression levels of Nrf2, HOX-1 and NF-κB were evaluated by quantitative real-time PCR. Oxidative stress was evaluated by malondialdehyde (MDA) and total Thiol (T-SH). TNF-α and IL-6 plasma levels were measured by ELISA. P-cresyl sulphate plasma level, a uremic toxin, was evaluated by high-performance liquid chromatography (HPLC) with fluorescent detection. RESULTS: Twenty-four patients finished the study: 10 in the curcumin group (57.5 ± 11.6 years) and 14 in the placebo group (56.5 ± 10.0 years). The plasma levels of MDA were reduced after 12 weeks in the curcumin group (p = 0.01), while the placebo group remained unchanged. However, regarding the difference between the groups at the endpoint, no change was observed in MDA. Still, there was a trend to reduce the p-CS plasma levels in the curcumin group compared to the placebo group (p = 0.07). Likewise, the concentrations of protein thiols, mRNA expression of Nrf2, HOX-1, NF-κB, and cytokines plasma levels did not show significant changes. CONCLUSION: Curcuminoid supplementation for twelve weeks attenuates lipid peroxidation and might reduce uremic toxin in patients with CKD undergoing PD. This study was registered on Clinicaltrials.gov as NCT04413266.

Capsicum annuum var. abbreviatum Induces a Macrophage M2-Like Phenotype Mediated by Nrf2/HO-1 in LPS-Stimulated Macrophages.

Capsicum annuum var. abbreviatum (CAAE), which is in the genus Capsicum L. (Solanaceae), was found to be richer in polyphenols and flavonoids than other prevalent peppers of Capsicum annuum var. angulosum and Capsicum annuum. L. Yet, it is still unclear how CAAE reduces inflammation. In this study, we used the lipopolysaccharide-stimulated RAW264.7 macrophage cell line and bone marrow-derived macrophages to assess its anti-inflammatory activities. Initially, we discovered that CAAE decreased the levels of nitric oxide and inducible nitric oxide synthase. In addition, CAAE decreased the intracellular reactive oxygen species levels and increased the nuclear factor-erythroid 2-related factor 2 and heme oxygenase-1 compared with the phenotype of M2 macrophages. CAAE inhibited the activation of mitogen-activated protein kinases (MAPKs), including extracellular signal-regulated kinases, c-Jun N-terminal kinases, and p38 MAPKs. CAAE also inhibited the translocation of nuclear factor kappa B into nuclear, hence preventing the production of proinflammatory cytokines. Therefore, we suggest that CAAE might have potential as a candidate therapeutic agent for inflammatory diseases.

Effect of electroacupuncture on Nrf2/NLRP3/Caspase-1 pathway mediated-pyroptosis in mice with Parkinson's disease. / ç”µé’ˆå¯¹å¸•é‡‘æ£®ç— å°é¼ Nrf2/NLRP3/Caspase-1通路介导的细胞焦亡的影响.

OBJECTIVES: To observe the effect of electroacupuncture (EA) on nuclear transcription factor E2 related factor 2 (Nrf2)/NOD-like receptor family pyrin domain-containing protein 3 (NLRP3)/cysteine aspartic acid specific protease-1 (Caspase-1) pathway in the substantia nigra (SN) of mice with Parkinson's disease (PD), so as to explore the neuroprotective mechanism of EA. METHODS: Forty C57BL/6 male mice were randomly divided into 4 groups, namely, control, PD model, EA and sham-EA groups, with 10 mice in each group. The PD mouse model was established by gavage of rotenone for 4 weeks. Mice in the EA group were given EA stimulation (1 mA, 2 Hz) at "Fengfu" (GV36), bilateral "Taichong" (LR3) and "Zusanli" (ST36) for 30 min, once daily for 2 consecutive weeks. And mice in the sham-EA group were given acupuncture at the subcutaneous areas of the same acupoints without EA stimulation. The open-field test was used for assessment of mouse behavior. The levels of interleukin-1ß (IL-1ß) and interleukin-18 (IL-18) in the serum were detected by enzyme-linked immunosorbent assay . The positive expression of tyrosine hydroxylase (TH) in SN was determined by immunohistochemistry. The mRNA expression levels of Nrf2, NLRP3, Caspase-1, gasdermin D(GSDMD), IL-1ß, IL-18 and the protein expression levels of Nrf2, NLRP3, Caspase-1 and GSDMD in the SN were detected by quantitative real-time PCR and Western blot, separately. RESULTS: After modeling, compared with the control group, the behavioral score was increased (P<0.01), the total exercise time, the total distance and the average speed were decreased (P<0.01), and the positive expression of TH and the mRNA and protein expression levels of Nrf2 in the SN were decreased (P<0.01), while the contents of IL-1ß and IL-18 in serum, the mRNA and protein expression levels of NLRP3, Caspase-1 and GSDMD and the mRNA expression levels of IL-1ß and IL-18 in the SN were up-regulated (P<0.01) in the PD model group. Following EA intervention, the behavioral score was decreased(P<0.01), the total exercise time, total distance and average speed were increased (P<0.01), the positive expression of TH and the mRNA and protein expressions of Nrf2 in SN were up-regulated (P<0.01, P<0.05), while the contents of IL-1ß and IL-18 in serum, the mRNA and protein expression levels of NLRP3, Caspase-1 and GSDMD as well as the mRNA expression levels of IL-1ß and IL-18 in the SN were down-regulated (P<0.01, P<0.05) in the EA group relative to the PD model and sham-EA groups. There were no significant differences in the above indicators between the PD model and sham-EA groups. CONCLUSIONS: EA stimulation of GV36, LR3 and ST36 can improve motor deficits, reduce the loss of dopamine neurons in the SN, and inhibit neuroinflammatory responses in mice with PD, which may be related to its effects in regulating the Nrf2/NLRP3/Caspase-1 pathway mediated pyroptosis.