NQO1 mRNA Research Articles

The administration of Glycyrrhiza polysaccharides mitigates liver injury in mice caused by mancozeb via the Keap1-Nrf2/NF-κB pathway

The extensive utilization of mancozeb (MCZ) poses environmental pollution risks, threatens human health, particularly hepatotoxicity. Glycyrrhiza polysaccharides (GP) exhibit antioxidant, anti-inflammatory and other biological activities. The aim of this study is to explore the mechanism of liver injury in mice exposed to MCZ and the protective effect of GP on alleviating MCZ induced liver injury. Initially, 70 female mice were divided into 7 groups, and the optimal dose of MCZ induced liver injury in mice was screened by oral administration different doses of MCZ (0, 50, 100, 150, 200, 250 and 300 mg/kg MCZ). The results demonstrated that, compared to the blank control group, as the concentration of MCZ increased, several physiological and biochemical parameters were significantly affected. Specifically, body weight and liver index significantly decreased, while the activities of SOD and CAT also decreased. Additionally, the content of ROS increased, the levels of Keap1 and Nrf2 proteins increased, the mRNA levels of Gpx2 and HO-1decreased, and the mRNA levels of Gstt2, GcLc and NQO1 were upregulated. Based on the test data, select 100mg/kg MCZ as the optimal modeling dose for experimental animals. Sixty female mice were divided into six groups and orally administered: control group A (0.2mL deionized water), model group B (100mg/kg MCZ), positive control group F (100mg/kg MCZ+100mg/kg VC), the high-dose GP group C (100mg/kgMCZ+200mg/kgGP), the medium-dose GP group D (100mg/kg MCZ+150mg/kgGP) and the low-dose GP group E (100mg/kg MCZ+100mg/kgGP). The results showed that compared to the model group, adding GP alleviated the effects of MCZ on body weight, liver index, CAT and SOD activity, MDA content, HO-1, TNF-α, and IL-1β. Additionally, the addition of GP decreased the expression of Keap1, Nrf2, NF-kB, and NQO1, GcLc, and Gstt2 mRNA. GP ameliorated liver vacuolar degeneration, steatosis and nuclear pyknosis ameliorate by MCZ. The results show that MCZ triggers hepatotoxicity via the activation of the Keap1/Nrf2 signaling pathway, whereas GP has the potential to mitigate liver damage caused by MCZ exposure by inhibiting this pathway.

Extracellular Vesicles Derived from Adipose-Derived Mesenchymal Stem Cells Alleviate Apoptosis and Oxidative Stress of Retinal Pigment Epithelial Cells Through Activation of Nrf2 Signaling Pathway.

Purpose: To examine the potential protective effects of adipose-derived mesenchymal stem cell-derived extracellular vesicles (ASC-EVs) on ARPE-19 cells exposed to hydrogen peroxide (H2O2) stress and to evaluate their ability to delay retinal degeneration in Royal College of Surgeons (RCS) rats. Methods: ARPE-19 cells were pre-treated with ASC-EVs for 24 h, followed by exposure to 200 μM H2O2 for an additional 24 h. RCS rats received an intravitreal injection of phosphate-buffered saline in one eye and ASC-EVs in the other eye. Results: ASC-EV pretreatment significantly protected against H2O2 in the Cell Counting Kit-8 assay and was also effective in the lactate dehydrogenase-release assay. It notably reduced early apoptosis (Annexin V-fluorescein isothiocyanate/propidium iodide assay) and late apoptosis (Terminal Deoxynucleotidyl Transferase dUTP Nick End Labeling assay), while significantly decreasing intracellular reactive oxygen species, glutathione levels, and superoxide dismutase activity. NFE2L2, HMOX1, and NQO1 mRNA levels, along with Nrf2, HO-1, and NQO1 protein levels, were significantly elevated with ASC-EV pretreatment. Compared with ARPE-19-derived EVs, 11 miRNAs were upregulated and 34 were downregulated in ASC-EVs. In RCS rats, intravitreal injections of ASC-EVs led to significant preservation of the outer nuclear layer and photoreceptor segments, along with increased nuclear Nrf2 expression and elevated HO-1 and NQO1 levels in the inner retina. Eyes that received intravitreal injections of ASC-EVs demonstrated significantly preserved electroretinography a- and b-wave amplitudes at 1 week post-injection, though this effect faded by 2 weeks. Conclusions: ASC-EVs mitigated apoptosis and oxidative stress in ARPE-19 cells subjected to H2O2 exposure and temporarily slowed retinal degeneration in RCS rats via Nrf2 pathway activation by miRNAs.

Interplay between fluorine and cadmium on intestinal accumulation, oxidative stress, permeability and inflammatory response in rats

Fluorine (F) and Cadmium (Cd) have given rise to public concern regarding their adverse impacts on the environment and human beings. Yet, the toxic interplay between F and Cd on the intestine is still vague. Aiming to investigate the role of F on Cd-damaged intestine, a total of five groups of 30 SD rats were picked at random to be gavaged for 90 days: Control group (Ultra-pure water), Cd (Cd 1 mg/kg), Cd+LF (Cd 1 mg/kg+F 15 mg/kg), Cd+MF (Cd 1 mg/kg+F 45 mg/kg), and Cd+HF (Cd 1 mg/kg+F 75 mg/kg). It demonstrated that Cd enriched in the intestine and disordered intestinal barrier of rats. Interestingly, two side effects of F were observed resisting to the Cd toxicity. The Cd levels in colon contents were attenuated by 45.45 %, 28.11 %, and 19.54 % by F supplement, respectively. In the Cd+LF group, SOD, GSH-Px, and CAT activities elevated by 0.93, 1.76, and 1.78 times, respectively, and the MDA content reduced 0.67 times; the expressions of NQO1, SOD2, and GSH-Px mRNA markedly enhanced, as well as the Keap1 mRNA significantly decreased. Nevertheless, all indexes above in the Cd+HF group showed the opposite trends. Furthermore, LPS levels decreased by 45.93 % for the Cd+LF group and increased by 12.70 % in that the Cd+HF group. The ZO-1 expression in the Cd+LF group increased, whereas the Cd+HF group's expressions of Claudin-1, Occludin, and ZO-1 were all diminished by 35.46 %, 27.23 %, and 16.32 %, respectively. Moreover, the levels of TNF-α, IL-1β and TLR-4 decreased and IL-10 level promoted, while all showed opposite trends in the Cd+HF group. Collectively, it indicated there is a twofold interplay between F and Cd on intestinal damage and mainly depends on F dosages.

Resolvin D5 Protects Female Hairless Mouse Skin from Pathological Alterations Caused by UVB Irradiation.

Resolvin D5 (RvD5) is a lipid mediator that has been reported to present anti-inflammatory and pro-resolution properties. Evidence also supports its capability to enhance reactive oxygen species (ROS) production during bacterial infections, which would be detrimental in diseases driven by ROS. The biological activity of RvD5 and mechanisms against UVB irradiation skin pathology have not been investigated so far. Female hairless mice were treated intraperitoneally with RvD5 before UVB stimulus. RvD5 reduced skin edema in a dose-dependent manner as well as oxidative stress by increasing antioxidants (endogenous tissue antioxidant scavenging of cationic radical, iron reduction, catalase activity and reduced glutathione levels) and decreasing pro-oxidants (superoxide anion and lipid peroxidation). RvD5 antioxidant activity was accompanied by enhancement of Nrf2, HO-1 and NQO1 mRNA expression. RvD5 reduced the production of IL-1β, TNF-α, TGF-β, and IL-10. RvD5 also reduced the inflammatory cell counts, including mast cells and neutrophils/macrophages. The reduction of oxidative stress and inflammation resulted in diminished matrix metalloproteinase 9 activity, collagen degradation, epidermal thickening and sunburn cell development. Therefore, this study demonstrates, to our knowledge, the first body of evidence that RvD5 can be used to treat UVB skin pathology and unveils, at least in part, its mechanisms of action.

Effects of intradialytic bicycle ergometer exercise on transcription factors NF-ĸB and Nrf2 in patients with chronic kidney disease: A randomized crossover clinical trial

PurposeTo evaluate the effects of an intradialytic aerobic exercise training program on the expression of transcription factors nuclear factor κappa B (NF-κB) and nuclear factor erythroid 2-related factor 2 (Nrf2), related to inflammatory and antioxidant pathways, respectively, in patients with chronic kidney disease (CKD) on hemodialysis. MethodsThis was a longitudinal, randomized clinical trial with a washout period and crossover performed with 33 patients randomized into two groups: Exercise (individualized intradialytic aerobic exercise on an adapted stationary exercise bike) three times per week for three months and control (without exercise). After the washout period (1 month), the exercise group became the control, and the other group performed the exercises for another three months. Blood sample collection, food intake, and anthropometry were evaluated at the beginning and end of each study phase. Nrf2, its target gene NAD(P)H quinone oxidoreductase 1 (NQO1), and NF-κB transcription factors were evaluated by quantitative real-time polymerase chain reaction (qRT-PCR) and the inflammatory cytokines tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) by ELISA assay. ResultsEighteen patients [11 men, 44.1 ± 8.4 years, 17.3 (6.6–124) months on HD] completed all the study. The obtained data revealed that the intervention did not affect Nrf2, NQO1, and NF-κB mRNA expression. Also, TNF-α levels were not changed. However, IL-6 showed a tendency to decrease after the exercise intervention (p = 0.054). ConclusionIn hemodialysis patients, three months of intradialytic aerobic exercise did not modulate the transcription factors associated with inflammation (NF-κB) and antioxidant activity (Nrf2 and NQO1). Clinical trials registration numberNCT04375553.

Role of NQO1 Gene Involvement and Susceptibility of T2DM Among Saudi Arabia Population.

NQO1 disruption enhances susceptibility to oxidative stress during hyperglycemia and is a significant contributor to the development and progression of diabetes. Oxidative stress has been linked to several symptoms, including hyperglycemia, reactive oxygen species buildup, high blood pressure, and the expression of inflammatory markers. Therefore, the present research aimed to evaluate the genetic abnormality of NQO1 (rs1800566, C609T) gene polymorphism, expression, and vitamin-D level assessment among Type 2 diabetes mellitus (T2DM) patients. The study included 100 newly diagnosed T2DM cases and 100 healthy individuals as healthy controls. Total RNA was extracted from the whole blood using the TRIzol method, and further cDNA was synthesized, and expression was evaluated. There is a significant difference in NQO1 (rs1800566, C609T) genotype distribution among the T2DM patients and healthy controls (p = 0.04). Compared with the NQO1 CC wild-type genotype, the NQO1 CT heterozygous genotype had an odds ratio of 1.96 (1.08-3.55), and the NQO1 TT mutant type genotype had an odds ratio of 3.31 (0.61-17.77). Significantly decreased expression of NQO1 mRNA was observed with heterozygous CT (p < 0.0001) and homozygous mutant TT genotype (p = 0.0004), compared with homozygous wild-type CC genotype. NQO1 mRNA expression level was also compared with vitamin D levels among the T2DM patients. T2DM patients with vitamin D deficiency had 1.83-fold NQO1 mRNA expression, while vitamin D insufficient and sufficient T2DM cases had 3.31-fold (p < 0.0001) and 3.70-fold (p < 0.0001) NQO1 mRNA expression. It was concluded that NQO1 (rs1800566, C609T) CT and TT genotypes played a significant role in the worseness of type II diabetes mellitus, and decreased expression of NQO1 mRNA expression could be an essential factor for disease worseness as well as hypermethylation could be a factor for reduced expression leading to disease severity. The decreased NQO1 mRNA expression with heterozygous CT and mutant TT genotype associated with vitamin D deficiency may contribute to disease progression.

Effect of iron administration on the aortic iron content and vascular calcification in phosphorus-loaded chronic kidney disease rats

BackgroundCardiovascular disease (CVD) is a major cause of morbidity and mortality in patients with chronic kidney disease (CKD) and could be related to oxidative stress. Vascular calcification (VC) has been established as a critical risk factor for accelerated CVD. In CKD, phosphorus (Pi), iron (Fe) and Nrf2 are modulators of VC and important agonists and antagonists of oxidative stress. The aim of this study was to determine whether Fe administration, which is commonly used to treat renal anemia, affects aortic Fe overload and VC, and whether Nrf2 and its related genes, ferritin H and HIF-1α, are involved in the development of VC.MethodsA CKD model was created in rats by administering adenine and simultaneously feeding a high-Pi diet. In addition to control and CKD rats without Fe administration (No-Fe group), Fe was administered orally (PO-Fe group) or intraperitoneally (IP-Fe group) to CKD animals to clarify the effects of Fe administration on the aortic Fe and calcium (Ca) contents and the involvement of Nrf2 and its induced antioxidative proteins, ferritin H and HIF-1α, in VC.ResultsThe aortic Fe content increased significantly in the IP-Fe group, which was closely correlated with liver HAMP (hepcidin) expression in all animals. Fe administration had no significant effect on the aortic Ca and Pi contents regardless of the route of Fe administration. The aortic mRNA level of Nrf2 was significantly increased in the IP-Fe group and correlated with serum Pi levels and aortic Fe contents, which could respond to oxidative stress. Notably, the mRNA level of Nrf2 was also significantly correlated with the mRNA levels of ferritin H and HIF-1α. Since we could not measure Nrf2 protein levels in this study, we confirmed the upregulation of HMOX1 and NQO1 mRNA expression in parallel with Nrf2 mRNA.ConclusionParenteral Fe administration increased aortic Fe in parallel with the liver HAMP mRNA level but did not affect VC. Aortic Nrf2 mRNA levels correlated significantly with aortic Fe and serum Pi levels and with aortic mRNA levels of ferritin H and HIF-1α as well as HMOX1 and NQO1.

Effects of rutin supplementation on intestinal morphology, antioxidant capacity, immunity, and gut microbiota of laying hens fed a diet containing stored soybean meal

Protein oxidation of soybean meal (SBM) during storage may have adverse effects on the intestinal health of laying hens. Moreover, rutin has anti-inflammatory and antioxidant properties which might be used as a feed additive to mitigate the intestinal damage caused by oxidised protein of SBM. This study aimed to investigate the effects of rutin supplementation on intestinal morphology, antioxidant capacity, immunity and caecal microbiota in laying hens fed a diet containing stored SBM. A total of 384 Hy-Line Brown laying hens (220 days) were randomly allocated into four groups with eight replicates of 12 laying hens each according to a 2 × 2 factorial arrangement with 2 types of SBM (FSM: SBM was stored in the cold storage warehouses at −20 °C for 45 days, and was considered as fresh and control SBM; RTSM: SBM was stored in room temperature warehouse (15 °C to 25 °C), average temperature was 20 °C for 45 days) and 2 levels of rutin (0 and 500 mg/kg). The results showed that the RTSM diet decreased the ileal glutathione peroxidase (GSH-Px) activity, the jejunal superoxide dismutase 2 (SOD2) and ileal NAD(P)H: quinone oxidoreductase 1 (NQO1) mRNA expression levels (p < 0.05), and tended to decrease the jejunal superoxide dismutase 1 (SOD1), ileal glutathione peroxidase 1 (GPX1) and increase the jejunal interleukin-1β (IL-1β) and interleukin-4 (IL-4) mRNA expression levels. Dietary rutin decreased the jejunal crypt depth (CD) (p < 0.05), increased the jejunal total antioxidant capacity (T-AOC) and GSH-Px activities (p < 0.05), decreased the content of interferon-γ (IFN-γ) in the jejunum (p < 0.05), and significantly reduced levels of immunoglobulin G (IgG), IL-1β, tumour necrosis factor-α (TNF-α), IFN-γ, and IL-4 in the ileal mucosa (p < 0.05). Dietary rutin increased SOD2, nuclear factor E2-related factor 2 (Nrf2) and NQO1 mRNA expression levels in the jejunum and GPX1, Nrf2 and NQO1 mRNA expression levels in the ileum, and decreased nuclear factor-κB (NF-κB), IL-1β, IFN-γ and IL-4 mRNA expression in the jejunum and NF-κB, IL-1β, TNF-α, IFN-γ and IL-4 mRNA expressions in the ileum. What’s more, dietary rutin changed the caecal microbiota. PCoA analysis indicated significant structural differences among four groups (p < 0.05), and SBM × Rutin interactions were found in Actinobacteriota and unclassifiedk_norank_d_ Bacteria at the phylum level (p < 0.05). These results suggested that RTSM had slight adverse effects on the intestinal health, and dietary rutin improved intestinal morphology, exerted antioxidant and anti-inflammatory effects via Nrf2 and NF-κB signal pathways, and changed the composition of caecal microbiota. Highlights The RTSM diet has adverse effects on the intestinal health of laying hens compared with the FSM diet. Dietary rutin improved the intestinal health by increasing the intestinal morphology, antioxidant capacity and anti-inflammatory effects. Dietary soybean meal and rutin changed the composition of caecal microbiota.

Effect of miR-27b-3p and Nrf2 in human retinal pigment epithelial cell induced by high-glucose.

To determine whether the microRNA-27b-3p (miR-27b-3p)/NF-E2-related factor 2 (Nrf2) pathway plays a role in human retinal pigment epithelial (hRPE) cell response to high glucose, how miR-27b-3p and Nrf2 expression are regulated, and whether this pathway could be specifically targeted. hRPE cells were cultured in normal glucose or high glucose for 1, 3, or 6d before measuring cellular proliferation rates using cell counting kit-8 and reactive oxygen species (ROS) levels using a dihydroethidium kit. miR-27b-3p, Nrf2, NAD(P)H quinone oxidoreductase 1 (NQO1) and heme oxygenase-1 (HO-1) mRNA and protein levels were analyzed using reverse transcription quantitative polymerase chain reaction (RT-qPCR) and immunocytofluorescence (ICF), respectively. Western blot analyses were performed to determine nuclear and total Nrf2 protein levels. Nrf2, NQO1, and HO-1 expression levels by RT-qPCR, ICF, or Western blot were further tested after miR-27b-3p overexpression or inhibitor lentiviral transfection. Finally, the expression level of those target genes was analyzed after treating hRPE cells with pyridoxamine. Persistent exposure to high glucose gradually suppressed hRPE Nrf2, NQO1, and HO-1 mRNA and protein levels and increased miR-27b-3p mRNA levels. High glucose also promoted ROS release and inhibited cellular proliferation. Nrf2, NQO1, and HO-1 mRNA levels decreased after miR-27b-3p overexpression and, conversely, both mRNA and protein levels increased after expressing a miR-27b-3p inhibitor. After treating hRPE cells exposed to high glucose with pyridoxamine, ROS levels tended to decreased, proliferation rate increased, Nrf2, NQO1, and HO-1 mRNA and protein levels were upregulated, and miR-27b-3p mRNA levels were suppressed. Nrf2 is a downstream target of miR-27b-3p. Furthermore, the miR-27b-3p inhibitor pyridoxamine can alleviate high glucose injury by regulating the miR-27b-3p/Nrf2 axis.

Nicotinamide mononucleotide inhibits oxidative stress-induced damage in a SIRT1/NQO-1-dependent manner

Oxidative stress causes endothelial dysfunction, which is associated with vascular cellular aging and is causally related to cardiovascular disease pathogenesis. Preclinical studies indicate that a nicotinamide adenine dinucleotide (NAD+) precursor, nicotinamide mononucleotide (NMN), alleviates oxidative stress in aged vessels, granting vasoprotective effects. However, the associated cellular mechanism remains largely unclear. In this study, we used human umbilical vein endothelial cells (HUVECs) to demonstrate that NMN inhibits oxidative stress-induced damage by activating the sirtuin 1 (SIRT1)/NAD(P)H: quinone oxidoreductase 1 (NQO-1) axis. We found that NMN inhibited H2O2-induced cytotoxicity and senescence-associated protein expression, such as p16 and p21. Furthermore, NMN prevented H2O2-induced actin cytoskeletal disorganization via inhibiting reactive oxygen species (ROS) production. NMN increased NQO-1 mRNA and protein expression that in turn was abrogated by SIRT1 inhibition, suggesting that NMN-inducible NQO-1 was associated with SIRT1 activity. SIRT1 and NQO-1 inhibition attenuated the inhibitory effect of NMN on H2O2-inducible cytotoxicity, senescence-related protein upregulation, and actin cytoskeletal disorganization. Our findings provide new insights into the mechanism by which NMN exerts protective effects against vascular oxidative stress.

Herbal cake-separated moxibustion relieves aspirin induced gastric mucosal injury by activating Nrf2/ARE/HO-1 signaling in rats

To investigate whether herbal cake-separated moxibustion can activate nuclear factor erythroid 2-related factor 2 （Nrf2） / antioxidant responsive element （ARE） /hemeoxygenase-1 （HO-1） signaling pathway to repair aspirin induced gastric mucosal injury （GMI） in rats. SD rats （half male and half female） were randomly divided into blank control, model, moxibustion, inhibitor of HO-1 （inhibitor）, model+inhibitor, moxibustion + inhibitor groups, with 20 rats in each group. The GMI model was established by gavage of aspirin 150 mg/kg（1 mL/100 g）. Herbal cake-separated moxibustion was alternatively applied to bilateral "Zusanli" （ST36） and "Zhongwan" （CV12） and bilateral "Pishu" （BL20） and "Weishu" （BL21） for 30 min, once daily for 8 days. The rats in the three inhibitor groups received intraperitoneal injection of HO-1 inhibitor zinc protoporphyrin （5 mg/kg）. The rats' behavior score, emotional response score, skin hair score, diet score and stool state score were given. The GMI index was calculated according to Guth's methods. Histopathological changes of gastric mucosa were observed by H.E. staining. The activity of superoxide dismutase （SOD） and the content of malondialdehyde （MDA） in the gastric mucosal tissue and serum were detected by ELISA. The levels of Nrf2/ARE/HO-1 signaling pathway-related factors Keap1, Nrf2, HO-1, CAT, GST, and NQO1 in the gastric mucosal tissue were detected by quantitative real-time PCR and Western blot, separately. Compared with the blank control group, the behavior score, emotional response score, skin hair score, diet score and stool state score, GMI index, MDA contents of gastric mucosal tissue and serum, expression le-vels of Keap1 mRNA and protein were significantly increased （P<0.01）, while the activity of gastric mucosal and serum SOD, the expression levels of Nrf2, HO-1, CAT, GST and NQO1 mRNAs and proteins were considerably decreased （P<0.01） in the model group. Compared with the model group, moxibustion obviously reversed the increase of emotional response score, skin hair score, stool state score, GMI index, MDA levels of gastric mucosal tissue and serum, and expression levels of Keap1 mRNA and protein （P<0.01, P<0.05）, and the decrease of activity of SOD of gastric mucosal and serum, and the expression levels of Nrf2, HO-1, CAT, GST and NQO1 mRNAs and proteins （P<0.01, P<0.05）. After administration of antagonist of HO-1, the effects of moxibustion were eliminated or weakened pronouncedly in reducing skin hair score, GMI index, contents of gastric mucosal and serum MDA, and expression of Keap1 mRNA and protein, and in up-regulating gastric mucosal and serum SOD, and expression of HO-1, CAT, GST and NQO1 mRNAs and proteins （P<0.01, P<0.05）. Herbal-cake separated moxibustion can improve the GMI in rats, which may be associated with its effects in reducing oxidative stress and activating Nrf2/ARE/HO-1 signaling pathway.

Herbal cake-separated moxibustion relieves aspirin induced gastric mucosal injury by activating Nrf2/ARE/HO-1 signaling in rats

To investigate whether herbal cake-separated moxibustion can activate nuclear factor erythroid 2-related factor 2 （Nrf2） / antioxidant responsive element （ARE） /hemeoxygenase-1 （HO-1） signaling pathway to repair aspirin induced gastric mucosal injury （GMI） in rats. SD rats （half male and half female） were randomly divided into blank control, model, moxibustion, inhibitor of HO-1 （inhibitor）, model+inhibitor, moxibustion + inhibitor groups, with 20 rats in each group. The GMI model was established by gavage of aspirin 150 mg/kg（1 mL/100 g）. Herbal cake-separated moxibustion was alternatively applied to bilateral "Zusanli" （ST36） and "Zhongwan" （CV12） and bilateral "Pishu" （BL20） and "Weishu" （BL21） for 30 min, once daily for 8 days. The rats in the three inhibitor groups received intraperitoneal injection of HO-1 inhibitor zinc protoporphyrin （5 mg/kg）. The rats' behavior score, emotional response score, skin hair score, diet score and stool state score were given. The GMI index was calculated according to Guth's methods. Histopathological changes of gastric mucosa were observed by H.E. staining. The activity of superoxide dismutase （SOD） and the content of malondialdehyde （MDA） in the gastric mucosal tissue and serum were detected by ELISA. The levels of Nrf2/ARE/HO-1 signaling pathway-related factors Keap1, Nrf2, HO-1, CAT, GST, and NQO1 in the gastric mucosal tissue were detected by quantitative real-time PCR and Western blot, separately. Compared with the blank control group, the behavior score, emotional response score, skin hair score, diet score and stool state score, GMI index, MDA contents of gastric mucosal tissue and serum, expression le-vels of Keap1 mRNA and protein were significantly increased （P<0.01）, while the activity of gastric mucosal and serum SOD, the expression levels of Nrf2, HO-1, CAT, GST and NQO1 mRNAs and proteins were considerably decreased （P<0.01） in the model group. Compared with the model group, moxibustion obviously reversed the increase of emotional response score, skin hair score, stool state score, GMI index, MDA levels of gastric mucosal tissue and serum, and expression levels of Keap1 mRNA and protein （P<0.01, P<0.05）, and the decrease of activity of SOD of gastric mucosal and serum, and the expression levels of Nrf2, HO-1, CAT, GST and NQO1 mRNAs and proteins （P<0.01, P<0.05）. After administration of antagonist of HO-1, the effects of moxibustion were eliminated or weakened pronouncedly in reducing skin hair score, GMI index, contents of gastric mucosal and serum MDA, and expression of Keap1 mRNA and protein, and in up-regulating gastric mucosal and serum SOD, and expression of HO-1, CAT, GST and NQO1 mRNAs and proteins （P<0.01, P<0.05）. Herbal-cake separated moxibustion can improve the GMI in rats, which may be associated with its effects in reducing oxidative stress and activating Nrf2/ARE/HO-1 signaling pathway.

Pan-cancer and single-cell analysis reveal the prognostic value and immune response of NQO1.

Background: Overexpression of the NAD(P)H: Quinone Oxidoreductase 1 (NQOI) gene has been linked with tumor progression, aggressiveness, drug resistance, and poor patient prognosis. Most research has described the biological function of the NQO1 in certain types and limited samples, but a comprehensive understanding of the NQO1's function and clinical importance at the pan-cancer level is scarce. More research is needed to understand the role of NQO1 in tumor infiltration, and immune checkpoint inhibitors in various cancers are needed. Methods: The NQO1 expression data for 33 types of pan-cancer and their association with the prognosis, pathologic stage, gender, immune cell infiltration, the tumor mutation burden, microsatellite instability, immune checkpoints, enrichment pathways, and the half-maximal inhibitory concentration (IC50) were downloaded from public databases. Results: Our findings indicate that the NQO1 gene was significantly upregulated in most cancer types. The Cox regression analysis showed that overexpression of the NQO1 gene was related to poor OS in Glioma, uveal melanoma, head and neck squamous cell carcinoma, kidney renal papillary cell carcinoma, and adrenocortical carcinoma. NQO1 mRNA expression positively correlated with infiltrating immune cells and checkpoint molecule levels. The single-cell analysis revealed a potential relationship between the NQO1 mRNA expression levels and the infiltration of immune cells and stromal cells in bladder urothelial carcinoma, invasive breast carcinoma, and colorectal cancer. Conversely, a negative association was noted between various drugs (17-AAG, Lapatinib, Trametinib, PD-0325901) and the NQO1 mRNA expression levels. Conclusion: NQO1 expression was significantly associated with prognosis, immune infiltrates, and drug resistance in multiple cancer types. The inhibition of the NQO1-dependent signaling pathways may provide a promising strategy for developing new cancer-targeted therapies.

Effects of sinapic acid on lead acetate-induced oxidative stress, apoptosis and inflammation in testicular tissue.

In this study, the effect of lead acetate (PbAc) and sinapic acid (SNP) administration on oxidative stress, apoptosis, inflammation, sperm quality and histopathology in testicular tissue of rats was tried to be determined. PbAc was administered at a dose of 30 mg/kg/bw for 7 days to induce testicular toxicity in rats. Oral doses of 5 and 10 mg/kg/bw SNP were administered to rats for 7 days after PbAc administration. According to our findings, while PbAc administration increased MDA content in rats, it decreased GPx, SOD, CAT activity and GSH content. NF-kB, IL-1β, TNF-α, and COX-2, which are among the inflammation parameters that increased due to PbAc, decreased with the administration of SNP. Nrf2, HO-1, and NQO1 mRNA transcript levels decreased with PbAc, but SNP treatments increased these mRNA levels in a dose-dependent manner. RAGE and NLRP3 gene expression were upregulated in PbAc treated rats. MAPK14, MAPK15, and JNK relative mRNA levels decreased with SNP treatment in PbAc treated rats. While the levels of apoptosis markers Bax, Caspase-3, and Apaf-1 increased in rats treated with PbAc, the level of Bcl-2 decreased, but SNP inhibited this apoptosis markers. PbAc caused histopathological deterioration in testis tissue and negatively affected spermatogenesis. When the sperm quality was examined, the decrease in sperm motility and spermatozoon density caused by PbAc, and the increase in the ratio of dead and abnormal spermatozoa were inhibited by SNP. As a result, while PbAc increased apoptosis and inflammation by inducing oxidative stress in testicles, SNP treatment inhibited these changes and increased sperm quality.

Vascular protection afforded by zinc supplementation in human coronary artery smooth muscle cells mediated by NRF2 signaling under hypoxia/reoxygenation

Zinc (Zn) has antioxidant, anti-inflammatory and anti-proliferative actions, with Zn dysregulation associated with coronary ischemia/reperfusion injury and smooth muscle cell dysfunction. As the majority of studies concerning Zn have been conducted under non-physiological hyperoxic conditions, we compare the effects of Zn chelation or supplementation on total intracellular Zn content, antioxidant NRF2 targeted gene transcription and hypoxia/reoxygenation-induced reactive oxygen species generation in human coronary artery smooth muscle cells (HCASMC) pre-adapted to hyperoxia (18 kPa O2) or normoxia (5 kPa O2). Expression of the smooth muscle marker SM22-α was unaffected by lowering pericellular O2, whereas calponin-1 was significantly upregulated in cells under 5 kPa O2, indicating a more physiological contractile phenotype under 5 kPa O2. Inductively coupled plasma mass spectrometry established that Zn supplementation (10 μM ZnCl2 + 0.5 μM pyrithione) significantly increased total Zn content in HCASMC under 18 but not 5 kPa O2. Zn supplementation increased metallothionein mRNA expression and NRF2 nuclear accumulation in cells under 18 or 5 kPa O2. Notably, NRF2 regulated HO-1 and NQO1 mRNA expression in response to Zn supplementation was only upregulated in cells under 18 but not 5 kPa. Furthermore, whilst hypoxia increased intracellular glutathione (GSH) in cells pre-adapted to 18 but not 5 kPa O2, reoxygenation had negligible effects on GSH or total Zn content. Reoxygenation-induced superoxide generation in cells under 18 kPa O2 was abrogated by PEG-superoxide dismutase but not by PEG-catalase, and Zn supplementation, but not Zn chelation, attenuated reoxygenation-induced superoxide generation in cells under 18 but not 5kPaO2, consistent with a lower redox stress under physiological normoxia. Our findings highlight that culture of HCASMC under physiological normoxia recapitulates an in vivo contractile phenotype and that effects of Zn on NRF2 signaling are altered by oxygen tension.

Beneficial effects of Chrysin on Cadmium‐induced nephrotoxicity in rats: Modulating the levels of Nrf2/HO‐1, RAGE/NLRP3, and Caspase-3/Bax/Bcl-2 signaling pathways

Cadmium (Cd) is a toxic heavy metal that targets the kidney directly in the body. Chrysin (CHR) is a natural flavonoid with many properties such as antioxidant, anti-inflammatory and anti-apoptotic. The current study discloses new evidence as regards of the curative effects of CHR on Cd-induced nephrotoxicity by regulating oxidative stress, apoptosis, autophagy, and inflammation. Cd was administered orally at a dose of 25 mg/kg body weight alone or in combination with orally administered CHR (25 and 50 mg/kg body weight) for 7 days. Biochemical, molecular, and histological methods were used to investigate inflammation, apoptosis, autophagy, and oxidant pathways in renal tissue. Renal function tests were also evaluated. Cd caused an increase in serum toxicity markers, lipid peroxidation and a decrease in the activities of antioxidant enzymes. Nrf-2 triggered inflammatory responses by suppressing HO-1 and NQO1 mRNA transcripts and increasing NF-κB, TNF-α, IL-1β and iNOS mRNA transcripts. Cd caused inflammasome by increasing RAGE and NLRP3 mRNA transcripts. In addition, Cd application caused apoptosis by increasing Bax, Apaf-1 and Caspase-3 mRNA transcripts and decreasing Bcl-2 mRNA transcript level. It caused autophagy by increasing the activity of Beclin-1 level. CHR treatment had the opposite effect on all these values and reduced the damage caused by all these signal pathways. Overall, the data of this study indicate that renal damage associated with Cd toxicity could be ameliorated by CHR administration.

The coumaric acid and syringic acid ameliorate acetic acid-induced ulcerative colitis in rats via modulator of Nrf2/HO-1 and pro-inflammatory cytokines

BackgroundUlcerative colitis (UC) is an inflammatory bowel disease (IBD) that causes uncontrolled inflammation and ulcers in your digestive tract. The coumaric acid and syringic acid are phenolic derivative found in many fruits and vegetables and is widely recognized for the ability of anti-parasitic, anti-microbial, anti-viral, anti-inflammatory, and antioxidant. The purpose of this study was to investigate the anti-inflammatory and antioxidant properties of coumaric acid and syringic acid on acetic acid-induced colitis in rats. MethodsA total of 64 male Wistar rats were divided into eight equal groups (n = 8). Colitis was induced by intrarectal administration of acetic acid, and rats orally received coumaric acid (100 and 150 mg/kg), syringic acid (10, 25, and 50 mg/kg), and dexamethasone (2 mg/kg) once per day for four days after colitis induction. Then, HO-1, Nrf2, and NQO1 mRNA expression were quantified by real time-PCR. Finally, the tissue levels of TNF-α and IL-1β protein were measured by ELISA. ResultsColitis led to a decrease in HO-1, Nrf2, and NQO1 mRNA expression and an increase in the tissue levels of TNF-α and IL-1β protein in the colon tissue. Treatment with dexamethasone significantly increased HO-1, Nrf2, and NQO1 mRNA expression and decreased the tissue levels of TNF-α and IL-1β protein compared to the UC group. Treatment with 150 mg/kg of coumaric acid and 50 mg/kg of syringic acid significantly increased HO-1, Nrf2, and NQO1 mRNA expression compared to the UC group. Also, treatment with 100 and 150 mg/kg of coumaric acid and 10, 25, and 50 mg/kg of syringic acid significantly decreased the tissue levels of TNF-α and IL-1β protein compared to the UC group. ConclusionThe coumaric acid and syringic acid, especially at high doses, may be an alternative strategy for the treatment of UC by the reduction of TNF-α and IL-1β levels and upregulation of the Nrf2/HO-1 pathway.

FLLL12, a curcumin analog, activates the oxidative stress response pathway in head and neck cancer: implication for chemoprevention.

Carcinogenesis is initiated with irreversible genetic mutation due to endogenously derived or carcinogen-induced oxidative stress. Activation of the antioxidant pathways ameliorates oxidative stress, protects cells from carcinogenic insults, and plays a pivotal role in chemoprevention. We have previously reported that FLLL12 is a potent curcumin analog, possesses in vitro and in vivo anticancer activity, and has better pharmacokinetic profiles than curcumin. The current study aims to identify novel pathways activated by FLLL12. MDA686, a head and neck cancer cell line, was treated with FLLL12 for 24h. Total RNA was used for RNA-Seq analysis. 2-fold differentially expressed genes were used for Ingenuity Pathway Analysis to identify the most significantly affected pathways. Real-time qPCR and western blotting were used to confirm the expression of the genes and their protein products, respectively, in normal, premalignant, and malignant cell lines. RNASeq identified 641 genes. Ingenuity Pathway Analysis identified the ferroptosis, the tumor microenvironment, and the oxidative response pathway as the top three most significantly affected pathways. We confirmed the activation of HMOX-1, NQO1, SLC7A11, and GCLC mRNA and proteins in normal, premalignant, and malignant cells. Although these genes are common for ferroptosis and the oxidative response pathway, treatment of cells with ferroptosis inhibitors, ferrostatin-1, and deferoxamine, did not affect FLLL12-induced cell death, suggesting that these genes are associated with the oxidative stress response pathway. Our results indicate that FLLL12 activates the oxidative stress response pathway in normal, premalignant, and malignant head and neck cancer cell lines and has strong promise for chemoprevention.

Regulation of CD163 Receptor in Patients with Abdominal Aortic Aneurysm and Associations with Antioxidant Enzymes HO-1 and NQO1.

Red blood cells are found within the abdominal aortic aneurysm (AAA), in the intraluminal thrombus (ILT), and in neovessels. Hemolysis promotes aortic degeneration, e.g., by heme-induced reactive oxygen species formation. To reduce its toxicity, hemoglobin is endocytosed by the CD163 receptor and heme is degraded by heme oxygenase-1 (HO-1). A soluble form (sCD163) is discussed as an inflammatory biomarker representing the activation of monocytes and macrophages. HO-1 and NAD(P)H quinone dehydrogenase 1 (NQO1) are antioxidant genes that are induced by the Nrf2 transcription factor, but their regulation in AAA is only poorly understood. The aim of the present study was to analyze linkages between CD163, Nrf2, HO-1, and NQO1 and to clarify if plasma sCD163 has diagnostic and risk stratification potential. Soluble CD163 was 1.3-fold (p = 0.015) higher in AAA compared to patients without arterial disease. The difference remained significant after adjusting for age and sex. sCD163 correlated with the thickness of the ILT (rs = 0.26; p = 0.02) but not with the AAA diameter or volume. A high aneurysmal CD163 mRNA was connected to increases in NQO1, HMOX1, and Nrf2 mRNA. Further studies are needed to analyze the modulation of the CD163/HO-1/NQO1 pathway with the overall goal of minimizing the detrimental effects of hemolysis.

Abstract 5257: Activation of NRF2-mediated oxidative stress response pathway by curcumin analog FLLL12: Implication for chemoprevention

Abstract Purpose: Carcinogenesis is a multi-step complex process initiated with irreversible genetic mutation due to endogenously derived or carcinogen-induced oxidative stress. Activation of the antioxidant pathways ameliorates oxidative stress, protects cells from carcinogenic insults, and plays a pivotal role in chemoprevention. We have previously reported that FLLL12 is a potent curcumin analog, possesses in vitro and in vivo anticancer activity, and has better pharmacokinetic profiles than curcumin. The purpose of the current study is to identify novel pathways activated by FLLL12 using RNASeq analysis. Methods: MDA686, a head and neck cancer cell line, were treated with 2 µM FLLL12 for 24h. Total RNA was isolated and used for RNASeq analysis. 2-fold differentially expressed genes were used for Ingenuity Pathway Analysis to identify the most significantly affected pathways. Real-time qPCR and western blotting were used to confirm the expression of the genes associated with the most significantly affected pathways and their protein products respectively in normal, premalignant, and malignant cell lines. Results: Whole transcriptome analysis using RNASeq identified 641 genes that were either upregulated or downregulated by 2-folds after FLLL12 treatment. Enrichment of this gene set with Ingenuity Pathway Analysis identified the ferroptosis signaling pathway, the tumor microenvironment pathway, and the NRF2-mediated oxidative response pathway as the top three most significantly affected pathways by FLLL12. We confirmed the activation of HMOX-1, NQO1, SLC7A11, and GCLC mRNA and proteins in HOK (normal), MSK-LEUK1 (premalignant), and MDA686TU (malignant) cells. Although these genes are common for the ferroptosis signaling pathway and the NRF2-mediated oxidative response pathway, treatment of cells with ferroptosis inhibitors, ferrostatin 1, and deferoxamine, had no effect on FLLL12-induced cell death suggesting that these genes are associated with oxidative stress response pathway. Conclusions: Our results strongly suggest that FLLL12 activates the oxidative stress response pathway in normal, premalignant, and malignant head and neck cancer cell lines and has strong promise for chemoprevention. Citation Format: Raji Lukmon, Adeoluwa A. Adeluola, A.R.M. Ruhul Amin. Activation of NRF2-mediated oxidative stress response pathway by curcumin analog FLLL12: Implication for chemoprevention. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5257.