Sinapic acid ameliorates D-galactosamine/lipopolysaccharideinduced fulminant hepatitis in rats:Role of nuclear factor erythroidrelated factor 2/heme oxygenase-1 pathways

BACKGROUND Sinapic acid(SA)has been shown to have various pharmacological properties such as antioxidant,antifibrotic,anti-inflammatory,and anticancer activities.Its mechanism of action is dependent upon its ability to curb free radical production and protect against oxidative stress-induced tissue injuries.AIM To study the hepatoprotective effects of SA against lipopolysaccharide(LPS)/Dgalactosamine(D-GalN)-induced acute liver failure(ALF)in rats.METHODS Experimental ALF was induced with an intraperitoneal(i.p.)administration of 8μg LPS and 800 mg/kg D-GalN in normal saline.SA was administered orally once daily starting 7 d before LPS/D-GalN treatment.RESULTS Data showed that SA ameliorates acute liver dysfunction,decreases serum levels of alanine transaminase(ALT),and aspartate aminotransferase(AST),as well as malondialdehyde(MDA)and NO levels in ALF model rats.However,pretreatment with SA(20 mg/kg and 40 mg/kg)reduced nuclear factor kappalight-chain-enhancer of activated B cells(NF-κB)activation and levels of inflammatory cytokines(tumor necrosis factor-αand interleukin 6).Also,SA increased the activity of the nuclear factor erythroid-related factor 2/heme oxygenase-1(Nrf2/HO-1)signaling pathway.CONCLUSION In conclusion,SA offers significant protection against LPS/D-GalN-induced ALF in rats by upregulating Nrf2/HO-1 and downregulating NF-κB.

Neuroprotective effects of salidroside on focal cerebral ischemia/reperfusion injury involve the nuclear erythroid 2-related factor 2 pathway

Salidroside,the main active ingredient extracted from Rhodiola crenulata,has been shown to be neuroprotective in ischemic cerebral injury,but the underlying mechanism for this neuroprotection is poorly understood.In the current study,the neuroprotective effect of salidroside on cerebral ischemia-induced oxidative stress and the role of the nuclear factor erythroid 2-related factor 2（Nrf2）pathway was investigated in a rat model of middle cerebral artery occlusion.Salidroside（30 mg/kg）reduced infarct size,improved neurological function and histological changes,increased activity of superoxide dismutase and glutathione-S-transferase,and reduced malon-dialdehyde levels after cerebral ischemia and reperfusion.Furthermore,salidroside apparently increased Nrf2 and heme oxygenase-1 expression.These results suggest that salidroside exerts its neuroprotective effect against cerebral ischemia through anti-oxidant mechanisms and that activation of the Nrf2 pathway is involved.The Nrf2/antioxidant response element pathway may become a new therapeutic target for the treatment of ischemic stroke.

Etomidate protects retinal ganglion cells from hydrogen peroxide-induced injury via Nrf2/HO-1 pathway

AIM:To determine whether etomidate(ET)has a protective effect on retinal ganglion cells(RGCs)injured with hydrogen peroxide(H_(2)O_(2))and to explore the potential mechanism underlying the antioxidative stress effect of ET.METHODS:Cultured RGCs were identified by double immunofluorescent labeling of microtubule-associated protein 2 and Thy1.1.An injury model of H_(2)O_(2)-induced RGCs oxidative stress was established in vitro.Cells were pretreated with different concentrations of ET(1,5,and 10μmol/L)for 4h,followed by further exposure to H_(2)O_(2)at 1000μmol/L.Cell counting kit 8 and Annexin V/propidium iodide assays were applied to detect the viabilities and apoptosis rates of the RGCs at 12,24,and 48h after H_(2)O_(2)stimulation.The levels of nitric oxide,malondialdehyde,and glutathione in culture media were measured at these time points.Quantitative reverse transcription polymerase chain reaction(qRT-PCR)and Western blot were performed to observe the effects of ET on the messenger RNA and protein expression of inducible nitric oxide synthase(iNOS),nuclear factor erythroid 2-related factor 2(Nrf2),heme oxygenase 1(HO-1),glutathione peroxidase 1 and the level of conjugated acrolein in RGCs at 12,24,and 48h after H_(2)O_(2)stimulation and in the retina at 12h after optic nerve transection(ONT).RESULTS:The applications of 5 and 10μmol/L of ET significantly increased the viability of RGCs.Results from qRT-PCR indicated a decrease in the expression of iNOS and an increase in the expressions of Nrf2 and HO-1 in ETpretreated RGCs at 12,24 and 48h after H_(2)O_(2)stimulation,as well as in ET-treated retinas at 12h after ONT.Western blot analysis revealed a decrease in the expression of iNOS and levels of conjugated acrolein,along with an increase in the expressions of Nrf2 and HO-1 in ET-pretreated RGCs in vitro and ET-treated retinas in vivo.CONCLUSION:ET is a neuroprotective agent in primary cultured RGCs injured by H_(2)O_(2).The effect of ET is dosedependent with the greatest effect being at 10μmol/L.ET plays an antioxidant role by inhibiting iNOS,up-regulating Nrf2/HO-1,decreasing the production of acrolein,and increasing the scavenge of acrolein.

Functionalized selenium nanoparticles ameliorated acetaminophen-induced hepatotoxicity through synergistically triggering PKCδ/Nrf2 signaling pathway and inhibiting CYP 2E1

Selenium nanoparticles(SeNPs)have been demonstrated potential for use in diseases associated with oxidative stress.Functionalized SeNPs with lower toxicity and higher biocompatibility could bring better therapeutic activity and clinical application value.Herein,this work was conducted to investigate the protective effect of Pleurotus tuber-regium polysaccharide-protein complex funtionnalized SeNPs(PTR-SeNPs)against acetaminophen(APAP)-induced oxidative injure in HepG2 cells and C57BL/6J mouse liver.Further elucidation of the underlying molecular mechanism,in particular their modulation of Nrf2 signaling pathway was also performed.The results showed that PTR-SeNPs could significantly ameliorate APAP-induced oxidative injury as evidenced by a range of biochemical analysis,histopathological examination and immunoblotting study.PTR-SeNPs could hosphorylate and activate PKCδ,depress Keap1,and increase nuclear accumulation of Nrf2,resulting in upregulation of GCLC,GCLM,HO-1 and NQO-1 expression.Besides,PTR-SeNPs suppressed the biotransformation of APAP to generate intracellular ROS through CYP 2E1 inhibition,restoring the mitochondrial morphology.Furthermore,the protective effect of PTR-SeNPs against APAP induced hepatotoxicity was weakened as Nrf2 was depleted in vivo,indicating the pivotal role of Nrf2 signaling pathway in PTR-SeNPs mediated hepatoprotective efficacy.Being a potential hepatic protectant,PTR-SeNPs could serve as a new source of selenium supplement for health-promoting and biomedical applications.

1,2,3,4,6-penta-O-galloyl-β-D-glucose Protects PC12 Cells from MPP^+-mediated Cell Death by Inducing Heme Oxygenase-1 in an ERK- and Akt-dependent Manner

This study examined the ability of 1,2,3,4,6-penta-O-galloyl-β-D-glucose (β-PGG) to induce the expression of heme oxygenase-1 (HO-1) in the PC12 cells and its regulation in the PC12 cells.One week before treatment with the drug,nerve growth factor (NGF) was added to the cultures at a final concentration of 50 ng/mL to induce neuronal differentiation.After drug treatment,HO-1 gene transcription was analyzed by reverse transcription polymerase chain reaction (RT-PCR).Expression of HO-1 and NF-E2-related factor2 (Nrf2) and activation of extracellular signal-regulated kinase (ERK) and Akt were detected by Western blotting.The viability of the PC12 cells treated with different medicines was examined by MTT assay.The oxidative stress in the PC12 cells was evaluated qualitatively and quantitatively by DCFH-DA.The results showed that β-PGG up-regulated HO-1 expression and this increased expression provided neuroprotection against MPP+-induced oxidative injury.Moreover,β-PGG induced Nrf2 nuclear translocation,which was found to be upstream of β-PGG-induced HO-1 expression,and the activation of ERK and Akt,a pathway that is involved in β-PGG-induced Nrf2 nuclear translocation,HO-1 expression and neuroprotection.In conclusion,β-PGG up-regulates HO-1 expression by stimulating Nrf2 nuclear translocation in an ERK-and Akt-dependent manner,and HO-1 expression by β-PGG may provide the PC12 cells with an acquired antioxidant defense capacity to survive the oxidative stress.

Ketogenic diet alleviates cognitive dysfunction and neuroinflammation in APP/PS1 mice via the Nrf2/HO-1 and NF-κB signaling pathways

Alzheimer's disease is a progressive neurological disorder characterized by cognitive decline and chronic inflammation within the brain.The ketogenic diet,a widely recognized therapeutic intervention for refractory epilepsy,has recently been proposed as a potential treatment for a variety of neurological diseases,including Alzheimer's disease.However,the efficacy of ketogenic diet in treating Alzheimer's disease and the underlying mechanism remains unclear.The current investigation aimed to explore the effect of ketogenic diet on cognitive function and the underlying biological mechanisms in a mouse model of Alzheimer's disease.Male amyloid precursor protein/presenilin 1(APP/PS1)mice were randomly assigned to either a ketogenic diet or control diet group,and received their respective diets for a duration of 3 months.The findings show that ketogenic diet administration enhanced cognitive function,attenuated amyloid plaque formation and proinflammatory cytokine levels in APP/PS1 mice,and augmented the nuclear factor-erythroid 2-p45 derived factor 2/heme oxygenase-1 signaling pathway while suppressing the nuclear factor-kappa B pathway.Collectively,these data suggest that ketogenic diet may have a therapeutic potential in treating Alzheimer's disease by ameliorating the neurotoxicity associated with Aβ-induced inflammation.This study highlights the urgent need for further research into the use of ketogenic diet as a potential therapy for Alzheimer's disease.

Upregulation of CDGSH iron sulfur domain 2 attenuates cerebral ischemia/reperfusion injury

CDGSH iron sulfur domain 2 can inhibit ferroptosis,which has been associated with cerebral ischemia/reperfusion,in individuals with head and neck cancer.Therefore,CDGSH iron sulfur domain 2 may be implicated in cerebral ischemia/reperfusion injury.To validate this hypothesis in the present study,we established mouse models of occlusion of the middle cerebral artery and HT22 cell models of oxygen-glucose deprivation and reoxygenation to mimic cerebral ischemia/reperfusion injury in vivo and in vitro,respectively.We found remarkably decreased CDGSH iron sulfur domain 2 expression in the mouse brain tissue and HT22 cells.When we used adeno-associated virus and plasmid to up-regulate CDGSH iron sulfur domain 2 expression in the brain tissue and HT22 cell models separately,mouse neurological dysfunction was greatly improved;the cerebral infarct volume was reduced;the survival rate of HT22 cells was increased;HT22 cell injury was alleviated;the expression of ferroptosis-related glutathione peroxidase 4,cystine-glutamate antiporter,and glutathione was increased;the levels of malondialdehyde,iron ions,and the expression of transferrin receptor 1 were decreased;and the expression of nuclear-factor E2-related factor 2/heme oxygenase 1 was increased.Inhibition of CDGSH iron sulfur domain 2 upregulation via the nuclear-factor E2-related factor 2 inhibitor ML385 in oxygen-glucose deprived and reoxygenated HT22 cells blocked the neuroprotective effects of CDGSH iron sulfur domain 2 up-regulation and the activation of the nuclear-factor E2-related factor 2/heme oxygenase 1 pathway.Our data indicate that the up-regulation of CDGSH iron sulfur domain 2 can attenuate cerebral ischemia/reperfusion injury,thus providing theoretical support from the perspectives of cytology and experimental zoology for the use of this protein as a therapeutic target in patients with cerebral ischemia/reperfusion injury.

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

AIM: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.METHODS: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.RESULTS: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.CONCLUSION: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.

Rhamnus crenata leaf extracts exhibit anti-inflammatory activity via modulating the Nrf2/HO-1 and NF-κB/MAPK signaling pathways

Objective:To elucidate the potential anti-inflammatory mechanisms of Rhamnus crenata leaf extracts using RAW264.7 cells.Methods:We used 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide assay to measure cell viability.Nitric oxide(NO)production was measured using Griess reagent.Western blotting and RT-PCR assays were carried out for analyzing the protein and gene expressions of pro-inflammatory mediators,respectively.Moreover,PD98059(ERK1/2 inhibitor),SB203580(p38 inhibitor),SP600125(JNK inhibitor),and BAY11-7082(NF-κB inhibitor)were used to evaluate the anti-inflammatory mechanism of Rhamnus crenata leaf extract.Results:Rhamnus crenata leaf extracts significantly inhibited the production of the pro-inflammatory mediators such as NO,iNOS,COX-2,IL-1β,and TNF-αin lipopolysaccharide(LPS)-stimulated RAW264.7 cells.Rhamnus crenata leaf extracts also suppressed LPS-induced degradation of IκB-αand nuclear accumulation of p65,which resulted in the inhibition of NF-κB activation in RAW264.7 cells.Additionally,the extracts attenuated the phosphorylation of p38,ERK1/2,and JNK in LPS-stimulated RAW264.7 cells.Moreover,HO-1 expression induced by Rhamnus crenata leaf extracts was significantly downregulated by SB230580,PD98059,SP600125 and BAY11-7082.Conclusions:Rhamnus crenata leaf extract may upregulate HO-1 expression through inhibition of p38,ERK1/2,and NF-κB activation,which may contribute to the anti-inflammatory activity of the extracts.Rhamnus crenata leaf extracts may have great potential for the development of anti-inflammatory drugs to treat acute and chronic inflammatory diseases.

Therapeutic Role of Chinese Medicine Targeting Nrf2/HO-1 Signaling Pathway in Myocardial Ischemia/Reperfusion Injury

Acute myocardial infarction(AMI),characterized by high incidence and mortality rates,poses a significant public health threat.Reperfusion therapy,though the preferred treatment for AMI,often exacerbates cardiac damage,leading to myocardial ischemia/reperfusion injury(MI/RI).Consequently,the development of strategies to reduce MI/RI is an urgent priority in cardiovascular therapy.Chinese medicine,recognized for its multi-component,multi-pathway,and multi-target capabilities,provides a novel approach for alleviating MI/RI.A key area of interest is the nuclear factor E2-related factor 2(Nrf2)/heme oxygenase-1(HO-1)pathway.This pathway is instrumental in regulating inflammatory responses,oxidative stress,apoptosis,endoplasmic reticulum stress,and ferroptosis in MI/RI.This paper presents a comprehensive overview of the Nrf2/HO-1 signaling pathway's structure and its influence on MI/RI.Additionally,it reviews the latest research on leveraging Chinese medicine to modulate the Nrf2/HO-1 pathway in MI/RI treatment.

D-GalN/LPS诱导大鼠急性肝衰竭中髓过氧化物酶和 Nrf2/HO-1信号通路的变化

目的 探讨D-氨基半乳糖(D-GalN)/脂多糖(LPS)诱导急性肝衰竭(acute liver failure,ALF)大鼠中髓过氧化物酶(myeloperoxidase,MPO)和核因子E2相关因子2(Nrf2)/血红素氧合酶-1(HO-1)信号通路的变化及ALF的发生机制。方法 SD大鼠随机分为对照组和ALF组。ALF组:将D-GalN 800 mg/kg和LPS 8μg/只同时腹腔注射,于注射后6、12和24 h检测血清总胆红素(TBiL)、丙氨酸氨基转移酶(ALT)、天门冬氨酸氨基转移酶(AST)含量。ELISA法检测血清MPO、肿瘤坏死因子-α(TNF-α)和白细胞介素-6(IL-6)水平,比色法测定MPO活性,RT-PCR检测肝组织Nrf2和HO-1的mRNA水平,Western blot法检测肝组织MPO、Nrf2和HO-1蛋白水平。结果 与对照组比较,ALF组血清MPO水平于造模后6 h升高,12 h为最高,24 h开始下降(23.33±2.06 vs.33.00±3.16,65.75±7.02,53.92±5.63,P<0.05);血清TNF-α(11.30±3.26 vs.102.17±14.80,83.33±11.22,64.25±9.29,P<0.01)和IL-6(10.83±2.92 vs.89.25±10.86,77.33±8.02,65.58±7.31,P<0.01)于造模后6 h升高最明显,12 h后逐渐下降,差异均有统计学意义。ALF组6、12、24 h肝组织MPO活性高于对照组,差异均有统计学意义(15.5±1.51,28.08±4.65,22.92±1.93 vs.12.17±1.27,P<0.05);ALF组肝组织6、12、24 h Nrf2 mRNA(1.59±0.13,3.65±0.11,2.35±0.11 vs.1.04±1.01,P<0.01)和HO-1 mRNA(2.44±0.19,4.77±0.18,3.82±0.17 vs.1.12±0.06,P<0.01)水平高于对照组,差异均有统计学意义。ALF组肝组织MPO(4.10±0.70,9.77±1.15,7.23±0.40 vs.2.07±0.42,P<0.01)、Nrf2(4.03±0.80,9.03±0.50,6.07±0.47 vs.2.63±0.38,P<0.01)和HO-1(1.73±0.21,5.17±0.51,3.03±0.32 vs.0.97±0.21,P<0.01)蛋白表达于12 h达最高值,ALF组各时间点与对照组比较均差异有统计学意义。结论 MPO可能通过氧化应激和炎症反应影响Nrf2/HO-1信号通路,在ALF中发挥重要作用。

胃癌组织中NrF-2和Ho-1的表达及临床意义

目的研究胃癌组织中核因子E-2相关因子2（nuclear factor E-2 related factor,Nr F-2）和血红素氧合酶-1（heme oxygenase-1,Ho-1）的表达并探讨其临床意义。方法选取从2016年2月-2016年6月,杭州市肿瘤医院病理科存档的人胃组织蜡块176例,其中胃癌78例,正常组织78例。分别采用免疫组织化学法检测2组人胃组织中Nr F-2和Ho-1的表达情况并进行分析。结果胃癌组Nr F-2和Ho-1水平分别为（0.752±0.098）、（0.862±0.081）,均显著高于正常组的（0.381±0.068）、（0.412±0.083）;Nr F-2、Ho-1阳性表达率在胃壁侵犯程度为T3＋T4、伴有淋巴结转移、NTM分期为Ⅲ、Ⅳ期胃癌组织中显著增高,上述差异均有统计学意义（均P〈0.05）。结论 Nr F-2和Ho-1在胃癌的发生发展中具有重要作用,且Nr F-2、Ho-1在胃癌组织中的阳性表达率与胃壁侵犯程度、有无淋巴结转移、NTM分期存在密切相关。提示了临床诊断中若联合检测Nr F-2和Ho-1,有利于对胃癌的早期诊断以及预后。

Iron increases HMOX1 and decreases hepatitis C viral expression in HCV-expressing cells

AIM： To investigate effects of iron on oxidative stress, heme oxygenase-1 （HMOX1） and hepatitis C viral （HCV） expression in human hepatoma ceils stably expressing HCV proteins. METHODS： Effects of iron on oxidative stress, HMOX1, and HCV expression were assessed in CON1 cells. Measurements included mRNA by quantitative reverse transcription-polymerase chain reaction, and protein levels by Western blots. RESULTS： Iron, in the form of ferric nitrilotriacetate,increased oxidative stress and upegulated HMOX1 gene expression. Iron did not affect mRNA or protein levels of Bach1, a repressor of HMOXl. Silencing the up-regulation of HMOXl nuclear factor-erythroid 2-related factor 2 （Nrf2） by Nrf2-siRNA decreased FeNTA-mediated up-regulation of HMOXl mRNA levels. These iron effects were completely blocked by deferoxamine （DFO）. Iron also significantly decreased levels of HCV core mRNA and protein by 80%-90%, nonstructural 5A mRNA by 90% and protein by about 50% in the Con1 full length HCV replicon cells, whereas DFO increased them. CONCLUSION： Excess iron up-regulates HMOXl and down-regulates HCV gene expression in hepatoma cells. This probably mitigates liver injury caused by combined iron overload and HCV infection.

Withaferin A inhibits ferroptosis and protects against intracerebral hemorrhage

Recent studies have indicated that suppressing oxidative stress and ferroptosis can considerably improve the prognosis of intracerebral hemorrhage(ICH).Withaferin A(WFA),a natural compound,exhibits a positive effect on a number of neurological diseases.However,the effects of WFA on oxidative stress and ferroptosis-mediated signaling pathways to ICH remain unknown.In this study,we investigated the neuroprotective effects and underlying mechanism for WFA in the regulation of ICH-induced oxidative stress and ferroptosis.We established a mouse model of ICH by injection of autologous tail artery blood into the caudate nucleus and an in vitro cell model of hemin-induced ICH.WFA was injected intracerebroventricularly at 0.1,1 or 5μg/kg once daily for 7 days,starting immediately after ICH operation.WFA markedly reduced brain tissue injury and iron deposition and improved neurological function in a dose-dependent manner 7 days after cerebral hemorrhage.Through in vitro experiments,cell viability test showed that WFA protected SH-SY5Y neuronal cells against hemin-induced cell injury.Enzyme-linked immunosorbent assays in vitro and in vivo showed that WFA markedly decreased the level of malondialdehyde,an oxidative stress marker,and increased the activities of anti-oxidative stress markers superoxide dismutase and glutathione peroxidase after ICH.Western blot assay,quantitative polymerase chain reaction and immunofluorescence results demonstrated that WFA activated the nuclear factor E2-related factor 2(Nrf2)/heme oxygenase-1(HO-1)signaling axis,promoted translocation of Nrf2 from the cytoplasm to nucleus,and increased HO-1 expression.Silencing Nrf2 with siRNA completely reversed HO-1 expression,oxidative stress and protective effects of WFA.Furthermore,WFA reduced hemin-induced ferroptosis.However,after treatment with an HO-1 inhibitor,the neuroprotective effects of WFA against hemin-induced ferroptosis were weakened.MTT test results showed that WFA combined with ferrostatin-1 reduced hemin-induced SH-SY5Y neuronal cell injury.Our findings reveal that WFA treatment alleviated ICH injury-induced ferroptosis and oxidative stress through activating the Nrf2/HO-1 pathway,which may highlight a potential role of WFA for the treatment of ICH.

Resveratrol reduces brain injury after subarachnoid hemorrhage by inhibiting oxidative stress and endoplasmic reticulum stress

Previous studies have shown that resveratrol,a bioactive substance found in many plants,can reduce early brain injury after subarachnoid hemorrhage,but how it acts is still unclear.This study explored the mechanism using the experimental subarachnoid hemorrhage rat model established by injecting autologous blood into the cerebellomedullary cistern.Rat models were treated with an intraperitoneal injection of 60 mg/kg resveratrol 2,6,24 and 46 hours after injury.At 48 hours after injury,their neurological function was assessed using a modified Garcia score.Brain edema was measured by the wet-dry method.Neuronal apoptosis in the prefrontal cortex was detected by terminal deoxyribonucleotidyl transferase-mediated biotin-16-dUTP nick-end labeling assay.Levels of reactive oxygen species and malondialdehyde in the prefrontal cortex were determined by colorimetry.CHOP,glucose-regulated protein 78,nuclear factor-erythroid2-related factor 2 and heme oxygenase-1 mRNA expression levels in the prefrontal cortex were measured by reverse transcription polymerase chain reaction.Tumor necrosis factor-alpha content in the prefrontal cortex was detected by enzyme linked immunosorbent assay.Immunohistochemical staining was used to detect the number of positive cells of nuclear factor-erythroid 2-related factor 2,heme oxygenase 1,glucose-regulated protein 78,CHOP and glial fibrillary acidic protein.Western blot assay was utilized to analyze the expression levels of nuclear factor-erythroid 2-related factor 2,heme oxygenase 1,glucose-regulated protein 78 and CHOP protein expression levels in the prefrontal cortex.The results showed that resveratrol treatment markedly alleviated neurological deficits and brain edema in experimental subarachnoid hemorrhage rats,and reduced neuronal apoptosis in the prefrontal cortex.Resveratrol reduced the levels of reactive oxygen species and malondialdehyde,and increased the expression of nuclear factor-erythroid 2-related factor 2,heme oxygenase-1 mRNA and protein in the prefrontal cortex.Resveratrol decreased glucose-regulated protein 78,CHOP mRNA and protein expression and tumor necrosis factor-alpha level.It also activated astrocytes.The results suggest that resveratrol exerted neuroprotective effect on subarachnoid hemorrhage by reducing oxidative damage,endoplasmic reticulum stress and neuroinflammation.The study was approved by the Animals Ethics Committee of Shandong University,China on February 22,2016(approval No.LL-201602022).

酸枣仁黄酮减轻全氟辛烷磺酸诱导小鼠肝脏氧化损伤

目的通过全氟辛烷磺酸(PFOS)诱导小鼠肝脏氧化损伤模型来研究酸枣仁黄酮(ZSSF)对其的保护作用。方法将小鼠随机分为空白组、造模组、维生素C组和ZSSF高、中、低剂量组,每组10只。灌胃给药同时用PFOS(10 mg/kg)造模,连续给药30 d。末次给药6 h后,取小鼠眼球血,取肝脏加入9倍体积的0.9%氯化钠溶液于匀浆器中制成10%的组织匀浆液,采用ELLSA法对血清中的丙转氨酶(ALT)、过氧化氢酶(CAT)、谷胱甘肽过氧化物酶(GSH-Px)、谷草转氨酶(AST)、丙二醛(MDA)、总抗氧化能力(T-AOC)和超氧化物歧化酶(SOD)等因子进行定量分析;同时应用RT-qPCR技术对肝组织中Nrf2、HO-1 mRNA的表达进行检测。结果造模组与空白组相比,ALT、AST、MDA含量升高,SOD、GSH-Px、CAT和T-AOC含量降低(P<0.01,P<0.05);给药干预组与模型组比较,减少了AST、ALT和MDA的含量,提高了SOD、CAT、GSH-Px、T-AOC含量以及肝组织中Nrf2、HO-1 mRNA的表达(P<0.01,P<0.05)。结论ZSSF具有改善PFOS诱导的小鼠氧化性肝损伤的作用,其可能的作用机制是通过调节Nrf2/HO-1途径增加体内抗氧化酶的活性,从而抑制过氧化脂质的合成。

Protective Effects of Anthocyanins Extracted from Vaccinium Uliginosum on 661W Cells Against Microwave-Induced Retinal Damage

Objective: To study the protective effect of anthocyanins extracted from Vaccinium Uliginosum(VU)on retinal 661W cells against microwave radiation induced retinal injury. Methods: 661W cells were divided into 6 groups, including control, model [661W cells radiated by microwave(30 mW/cm2, 1 h)] and VU groups [661W cells pretreated with anthocyanins extracted from VU(25, 50, 100 and 200 μg/mL, respectively) for 48 h, and radiated by microwave 30 mW/cm2, 1 h]. After treatment with different interventions, the cell apoptosis index(AI)was determined using Heochst staining;contents of malonaldehyde(MDA), glutataione(GSH), and activity of superoxide dismutase(SOD) were measured. mRNA expressions of nuclear factor erythroid 2-related factor 2(Nrf2) and heme oxygenase 1(HO-1) were detected by real time quantitative polymerase chain reaction, and the expression of HO-1 protein was examined by Western blot analysis. Nucleus and cytoplasm were separated and Nrf2 protein expression was further verified by Western blot analysis. Results: There was significant difference in AI among the groups(F=322.83, P<0.05). Compared with the control group, AI was significantly higher in the model group and was lower in 4 VU-pretreated groups(P<0.05). Linear regression analysis showed the decline of AI was in a dose-dependent manner with VU treatment(r=0.8419, P<0.05). The MDA and GSH contents of 661W cells in VU-treated groups were significantly lower than the model group(P<0.05). Compared with the model group, the SOD activity in the VU-treated groups(50, 100 and 200 μg/mL) was significantly higher(all P<0.05). The Nrf2 and HO-1 mRNA expressions were slightly increased after irradiation, and obviously increased in 100 μg/mL VU-treated group. After irradiation, the relative expressions of HO-1 and Nrf2 proteins in nucleus were slightly increased(P<0.05), and the changes in cytoplasm were not obvious,whereas it was significantly increased in both nucleus and cytoplasm in the VU treatment groups. Conclusions:Anthocyanins extracted from VU could reduce apoptosis, stabilize cell membrane, and alleviate oxidant injury of mouse retinal photoreceptor 661W cells. The mechanism might be through activating Nrf2/HO-1 signal pathway and inducing HO-1 transcription and translation.

Antihepatofibrotic effect of Guizhifuling pill(桂枝茯苓丸) on carbon tetrachloride-induced liver fibrosis in mice

OBJECTIVE: To evaluate the protective effects and the underlying mechanism of Guizhifuling pill(桂枝茯苓丸, GZFL) on carbon tetrachloride(CCl4)-induced hepatic fibrosis in mice. METHODS: Male ICR mice by intraperitoneally administered with 20% CCl4(mixed 1∶4 in soybean oil) to induce liver fibrosis. Mice that underwent CCl4 were orally with GZFL. Using hematoxylin and eosin and Masson staining to examine the pathological changes in liver tissue. Serum biochemical parameters, antioxidant enzyme activity and proinflammatory cytokines was assessed. Nuclear factor-kappa B(NF-κB) pathway and nuclear factor-erythroid 2-related factor 2(Nrf2) family members were evaluated by Western blotting. RESULTS: Our findings indicated that GZFL could effectively suppress the progression of liver fibrosis in mice, which was determined based on the improvement in liver function and reduction of collagen deposition. GZFL treatment also decreased the level of cytokines and increased the activity of antioxidant enzymes in liver tissue. Moreover, GZFL exerted anti-inflammatory and antioxidant effects through regulating the Nrf2-mediated antioxidant system and inhibiting the NF-κB pathway. CONCLUSIONS: GZFL may prevent the progression of liver fibrosis by regulating the Nrf2/heme oxygenase-1 and NF-κB signaling pathways, thereby highlighting its role in the management of liver fibrosis.