Poplar canker,caused by the fungus Cytospora chrysosperma,results in tremendous losses in poplar plantations in China.Although NADPH oxidases(NOXs)play important roles in the development and pathogenicity of several p...Poplar canker,caused by the fungus Cytospora chrysosperma,results in tremendous losses in poplar plantations in China.Although NADPH oxidases(NOXs)play important roles in the development and pathogenicity of several pathogenic fungi,their roles in C.chrysosperma remain unclear.In this study,we characterized three NOX genes(CcNox1,CcNox2,and CcNoxR)in C.chrysosperma.All three genes were highly upregulated during poplar branch infection,and deletion of any of them severely reduced virulence on poplar branches.Furthermore,deletion of either CcNox1 or CcNoxR resulted in a significant increase in endogenous reactive oxygen species production in hyphae,enhanced influx of Ca^(2+),the disruption of redox homeostasis and compromised mitochondrial integrity.Moreover,biosynthesis and secretion of a known virulence factor oxalic acid was obviously defective and exogenous oxalic acid supplementation rescued the virulence of the mutants.Taken together,our findings reveal that NOXs play important roles in redox homeostasis,mitochondrial integrity and pathogenicity in C.chrysosperma.展开更多
Endogenous elicitor, termed cellulase-degraded cell wall (CDW), was prepared from the cell wall of suspension-cultured ginseng (Panax ginseng C.A. Meyer) cells via cellulase degradation. CDW activated the NADPH oxidas...Endogenous elicitor, termed cellulase-degraded cell wall (CDW), was prepared from the cell wall of suspension-cultured ginseng (Panax ginseng C.A. Meyer) cells via cellulase degradation. CDW activated the NADPH oxidase activity of isolated plasma membranes and stimulated in vivo H2O2 generation in ginseng cell suspensions. CDW also increased the activity of phenylalanine ammonia lyase (PAL), expression of a P. ginseng squalene epoxidase (sqe) gene and saponin synthesis. NADPH oxidase inhibitors inhibited both in vitro NADPH oxidase activity and in vivo H2O2 generation. Induction of PAL activity, saponin synthesis and sqe gene expression were all inhibited by such inhibitor treatments and reduced by incubation with catalase and HA scavengers. These data indicate that activation of NADPH oxidase and generation of H2O2 are essential signalling events mediating defence responses induced by the endogenous elicitor(s) present in CDW.展开更多
Intracerebral hemorrhage is often accompanied by oxidative stress induced by reactive oxygen species,which causes abnormal mitochondrial function and secondary reactive oxygen species generation.This creates a vicious...Intracerebral hemorrhage is often accompanied by oxidative stress induced by reactive oxygen species,which causes abnormal mitochondrial function and secondary reactive oxygen species generation.This creates a vicious cycle leading to reactive oxygen species accumulation,resulting in progression of the pathological process.Therefore,breaking the cycle to inhibit reactive oxygen species accumulation is critical for reducing neuronal death after intracerebral hemorrhage.Our previous study found that increased expression of nicotinamide adenine dinucleotide phosphate oxidase 4(NADPH oxidase 4,NOX4)led to neuronal apoptosis and damage to the blood-brain barrier after intracerebral hemorrhage.The purpose of this study was to investigate the role of NOX4 in the circle involving the neuronal tolerance to oxidative stress,mitochondrial reactive oxygen species and modes of neuronal death other than apoptosis after intracerebral hemorrhage.We found that NOX4 knockdown by adeno-associated virus(AAV-NOX4)in rats enhanced neuronal tolerance to oxidative stress,enabling them to better resist the oxidative stress caused by intracerebral hemorrhage.Knockdown of NOX4 also reduced the production of reactive oxygen species in the mitochondria,relieved mitochondrial damage,prevented secondary reactive oxygen species accumulation,reduced neuronal pyroptosis and contributed to relieving secondary brain injury after intracerebral hemorrhage in rats.Finally,we used a mitochondria-targeted superoxide dismutase mimetic to explore the relationship between reactive oxygen species and NOX4.The mitochondria-targeted superoxide dismutase mimetic inhibited the expression of NOX4 and neuronal pyroptosis,which is similar to the effect of AAV-NOX4.This indicates that NOX4 is likely to be an important target for inhibiting mitochondrial reactive oxygen species production,and NOX4 inhibitors can be used to alleviate oxidative stress response induced by intracerebral hemorrhage.展开更多
Increased reactive oxygen species by the activation of NADPH oxidase（NOX） contributes to the development of diabetic complications.Apocynin,a NOX inhibitor,increases sciatic nerve conductance and blood flow in diabe...Increased reactive oxygen species by the activation of NADPH oxidase（NOX） contributes to the development of diabetic complications.Apocynin,a NOX inhibitor,increases sciatic nerve conductance and blood flow in diabetic rats.We investigated potential protective effect of apocynin in rat diabetic neuropathy and its precise mechanism of action at molecular level.Rat models of streptozotocin-induced diabetes were treated with apocynin（30 and 100 mg/kg per day,intragastrically） for 4 weeks.Mechanical hyperalgesia and allodynia were determined weekly using analgesimeter and dynamic plantar aesthesiometer.Western blot analysis and histochemistry/immunohistochemistry were performed in the lumbar spinal cord and sciatic nerve respectively.Streptozotocin injection reduced pain threshold in analgesimeter,but not in aesthesiometer.Apocynin treatment increased pain threshold dose-dependently.Western blot analysis showed an increase in catalase and NOX-p47 phox protein expression in the spinal cord.However,protein expressions of neuronal and inducible nitric oxide synthase（n NOS,i NOS）,superoxide dismutase,glutathion peroxidase,nitrotyrosine,tumor necrosis factor-α,interleukin-6,interleukin-1β,aldose reductase,cyclooxygenase-2 or MAC-1（marker for increased microgliosis） in the spinal cord remained unchanged.Western blot analysis results also demonstrated that apocynin decreased NOX-p47 phox expression at both doses and catalase expression at 100 mg/kg per day.Histochemistry of diabetic sciatic nerve revealed marked degeneration.n NOS and i NOS immunoreactivities were increased,while S-100 immunoreactivity（Schwann cell marker） was decreased in sciatic nerve.Apocynin treatment reversed these changes dose-dependently.In conclusion,decreased pain threshold of diabetic rats was accompanied by increased NOX and catalase expression in the spinal cord and increased degeneration in the sciatic nerve characterized by increased NOS expression and Schwann cell loss.Apocynin treatment attenuates neuropathic pain by decelerating the increased oxidative stress-mediated pathogenesis in diabetic rats.展开更多
Neuroinflammation mediated by microglia and oxidative stress play pivotal roles in the development of chronic temporal lobe epilepsy(TLE).We postulated that kainic acid(KA)-Induced status epilepticus triggers microgli...Neuroinflammation mediated by microglia and oxidative stress play pivotal roles in the development of chronic temporal lobe epilepsy(TLE).We postulated that kainic acid(KA)-Induced status epilepticus triggers microglia-dependent inflammation,leading to neuronal damage,a lowered seizure threshold,and the emergence of spontaneous recurrent seizures(SRS).Extensive evidence from our laboratory suggests that dextromethorphan(DM),even in ultra-low doses,has anti-inflammatory and neuroprotective effects in many animal models of neurodegenerative disease.Our results showed that administration of DM(10 ng/kg per day;subcutaneously via osmotic minipump for 4 weeks)significantly mitigated the residual effects of KA,including the frequency of SRS and seizure susceptibility.In addition,DM-treated rats showed improved cognitive function and reduced hippocampal neuronal loss.We found suppressed microglial activation-mediated neuroinflammation and decreased expression of hippocampal gp91^(phox) and p47^(phox) proteins in KA-induced chronic TLE rats.Notably,even after discontinuation of DM treatment,ultra-low doses of DM continued to confer long-term anti-seizure and neuroprotective effects,which were attributed to the inhibition of microglial NADPH oxidase 2 as revealed by mechanistic studies.展开更多
A rapid and concentration-dependent generation of superoxide anion （·O2^-）, measured with a superoxide-specific Cypridina luciferin-derived chemiluminescent reagent, was observed when two lanthanide salts （LaC...A rapid and concentration-dependent generation of superoxide anion （·O2^-）, measured with a superoxide-specific Cypridina luciferin-derived chemiluminescent reagent, was observed when two lanthanide salts （LaCl3 and CdCl3 ） were added to tobacco （ Nicotiana tabacum） cell suspension culture. Addition of superoxide dismutase （480 U·ml^-1） and Tiron （5 μmol·L^-1） to cell culture suspension decreases the level of lanthanide cation-induced ·O2^- generation, suggesting that ·O2^- generation is extra-cellular. Pretreatment of the cell culture suspension with diphenyleneiodonium （10 and 50 μmol·L^-1 ）, quinacrine （ 1 and 5 mmol· L^-1 ） and imidazol （ 10 mmol· L^-1 ）, inhibitors of NADPH oxidase, notably inhibits the generation of superoxide induced by lanthanide cation, implying the possible involvement of activation of NADPH oxidase. In addition, addition of SHAM （1 and 5 mmol· L^-1）, azide （0.2 and 1 mmol· L^-1 ）, inhibitor of peroxidase, has no influence on ·O2^- generation.展开更多
Diseases like Alzheimer’s and Parkinson’s diseases are defined by inflammation and the damage neurons undergo due to oxidative stress. A primary reactive oxygen species contributor in the central nervous system, NAD...Diseases like Alzheimer’s and Parkinson’s diseases are defined by inflammation and the damage neurons undergo due to oxidative stress. A primary reactive oxygen species contributor in the central nervous system, NADPH oxidase 4, is viewed as a potential therapeutic touchstone and indicative marker for these ailments. This in-depth review brings to light distinct features of NADPH oxidase 4, responsible for generating superoxide and hydrogen peroxide, emphasizing its pivotal role in activating glial cells, inciting inflammation, and disturbing neuronal functions. Significantly, malfunctioning astrocytes, forming the majority in the central nervous system, play a part in advancing neurodegenerative diseases, due to their reactive oxygen species and inflammatory factor secretion. Our study reveals that aiming at NADPH oxidase 4 within astrocytes could be a viable treatment pathway to reduce oxidative damage and halt neurodegenerative processes. Adjusting NADPH oxidase 4 activity might influence the neuroinflammatory cytokine levels, including myeloperoxidase and osteopontin, offering better prospects for conditions like Alzheimer’s disease and Parkinson’s disease. This review sheds light on the role of NADPH oxidase 4 in neural degeneration, emphasizing its drug target potential, and paving the path for novel treatment approaches to combat these severe conditions.展开更多
Production of reactive oxygen species(ROS)is a conserved immune response primarily mediated by NADPH oxidases(NOXs),also known in plants as respiratory burst oxidase homologs(RBOHs).Most microbe-associated molecular p...Production of reactive oxygen species(ROS)is a conserved immune response primarily mediated by NADPH oxidases(NOXs),also known in plants as respiratory burst oxidase homologs(RBOHs).Most microbe-associated molecular patterns(MAMPs)trigger a very fast and transient ROS burst in plants.However,recently,we found that lipopolysaccharides(LPS),a typical bacterial MAMP,triggered a biphasic ROS burst.In this study,we isolated mutants defective in LPS-triggered biphasic ROS burst(delt)in Arabidopsis,and cloned the DELT1 gene that was shown to encode RBOHD.In the delt1-2 allele,the antepenultimate residue,glutamic acid(E919),at the C-terminus of RBOHD was mutated to lysine(K).E919 is a highly conserved residue in NADPH oxidases,and a mutation of the corresponding residue E568 in human NOX2 has been reported to be one of the causes of chronic granulomatous disease.Consistently,we found that residue E919 was indispensable for RBOHD function in the MAMP-induced ROS burst and stomatal closure.It has been suggested that the mutation of this residue in other NADPH oxidases impairs the protein’s stability and complex assembly.However,we found that the E919K mutation did not affect RBOHD protein abundance or the ability of protein association,suggesting that the residue E919 in RBOHD might have a regulatory mechanism different from that of other NOXs.Taken together,our results confirm that the antepenultimate residue E is critical for NADPH oxidases and provide a new insight into the regulatory mechanisms of RBOHD.展开更多
Mitochondrial DNA(mtDNA) common deletion(CD) plays a significant role in aging and age-related diseases.In this study,we used D-galactose(D-gal) to generate an animal model of aging and the involvement and causative m...Mitochondrial DNA(mtDNA) common deletion(CD) plays a significant role in aging and age-related diseases.In this study,we used D-galactose(D-gal) to generate an animal model of aging and the involvement and causative mechanisms of mitochondrial damage in such a model were investigated.Twenty 5-week-old male Sprague-Dawley rats were randomly divided into two groups:D-gal group(n=10) and control group(n=10).The quantity of the mtDNA CD in the hippocampus was determined using a TaqMan real-time PCR assay.Transmission electron microscopy was used to observe the mitochondrial ultrastructure in the hippocampus.Western blot was used to detect the protein levels of NADPH oxidase(NOX) and uncoupling protein 2(UCP2).We found that the level of mtDNA CD was significantly higher in the hippocampus of D-gal-induced aging rats than in control rats.In comparison with the control group,the mitochondrial ultrastructure in the hippocampus of D-gal-treated rats was damaged,and the protein levels of NOX and UCP2 were significantly increased in the hippocampus of D-gal-induced aging rats.This study demonstrated that the levels of mtDNA CD and NOX protein expression were significantly increased in the hippocampus of D-gal-induced aging rats.These findings indicate that NOX-dependent reactive oxygen species generation may contribute to D-gal-induced mitochondrial damage.展开更多
Excess production of reactive oxygen species （ROS） critically contributes to occurrence of reperfusion injury, the paradoxical response of ischemic brain tissue to restoration of cerebral blood flow. However, the en...Excess production of reactive oxygen species （ROS） critically contributes to occurrence of reperfusion injury, the paradoxical response of ischemic brain tissue to restoration of cerebral blood flow. However, the enzymatic sources of ROS generation remain to be unclear. This study examined Nox2-ontaining NADPH oxidase （Nox2） expression and its activity in ischemic brain tissue following post-ischemic reperfusion to clarify the mechanism of enzymatic reaction of ROS. Male Sprague-Dawley rats were subjected to 90-minute middle cerebral artery occlusion, followed by 3 or 22.5 hours of reperfusion. Quantitative reverse transcriptase PCR and western blot assay were performed to measure mRNA and protein expression of Nox2. Lucigenin fluorescence assays were performed to assess Nox activity. Our data showed that Nox2 mRNA and protein expression levels were significantly increased （3.7-fold for mRNA and 3.6-fold for protein） in ischemic brain tissue at 22.5 hours but not at 3 hours following post-ischemic reperfusion. Similar results were obtained for the changes of NADPH oxidase activity in ischemic cerebral tissue at the two reperfusion time points. Our results suggest that Nox2 may not contribute to the early burst of reperfusion-related ROS generation, but is rather an important source of ROS generation during prolonged reperfusion.展开更多
The antibody against AT1-EC2 plays a role in some kinds of inflammatory vascular diseases including malignant hypertension,preeclampsia,and renal-allograft rejection,but the detailed mechanisms remain unclear.In order...The antibody against AT1-EC2 plays a role in some kinds of inflammatory vascular diseases including malignant hypertension,preeclampsia,and renal-allograft rejection,but the detailed mechanisms remain unclear.In order to investigate the changes of NADPH oxidase and reactive oxygen species in the aorta in a mouse model which can produce AT1-EC2 antibody by active immunization with AT1-EC2 peptide,15 mice were divided into three groups:control group,AT1-EC2-immunized group,and AT1-EC2-immunized and valsartan-treated group.In AT1-EC2-immunized group and AT1-EC2-immunized and valsartan-treated group,the mice were immunized by 50 μg peptide subcutaneously at multiple points for 4 times:0,5,10,and 15 days after the experiment.In AT1-EC2-immunized and valsartan-treated group,valsartan was given at a dose of 100 mg/kg every day for 20 days.After the experiment,the mice were sacrificed under anesthesia and the aortas were obtained and frozen in liquid nitrogen for the preparation of frozen section slides and other experiments.The titer of AT1-EC2 was assayed by using ELISA.The level of NOX1 mRNA in the aorta was determined by using RT-PCR.The expression of NOX1 was detected by using Western blotting.Confocal scanning microscopy was used to assay the α-actin and NOX1 expression in the aortic tissue.The O 2.production was detected in situ after DHE staining.The mice produced high level antibody against AT1-EC2 in AT1-EC2-immunized group and AT1-EC2-immunized and valsartan-treated group,and the level of NOX1 mRNA in the aortic tissues was 1.6±0.4 times higher and the NOX1 protein expression was higher in AT1-EC2-immunized group than in control group.There were no significant differences in the level of NOX1 mRNA and protein expression between control group and AT1-EC2-immunized and valsartan-treated group.The expression and co-localization of α-actin and NOX1 in AT1-EC2-immunized group increased significantly as compared with those in control group,and the O 2.production increased about 2.7 times as compared with control group.There were no significant differences between control group and AT1-EC2-immunized and valsartan-treated group.It is concluded that active immunization with AT1-EC2 can activate NOX1-ROS,and increase vascular inflammation,which can be inhibited by AT1 receptor blocker valsartan.This may partially explain the mechanism of the pathogenesis of inflammatory vascular diseases related to antibody against AT1-EC2.展开更多
Objective The roles of cerebrovascular oxidative stress in vascular functional remodeling have been described in hindlimb-unweighting （HU） rats. However, the underlying mechanism remains to be established. Methods W...Objective The roles of cerebrovascular oxidative stress in vascular functional remodeling have been described in hindlimb-unweighting （HU） rats. However, the underlying mechanism remains to be established. Methods We investigated the generation of vascular reactive oxygen species （ROS）, Nox2/Nox4 protein and mRNA levels, NADPH oxidase activity, and manganese superoxide dismutase （MnSOD） and glutathione peroxidase-1 （GPx-1） mRNA levels in cerebral and mesenteric smooth muscle cells （VSMCs） of HU rats. Results ROS production increased in cerebral but not in mesenteric VSMCs of HU rats compared with those in control rats. Nox2 and Nox4 protein and mRNA levels were increased significantly but MnSOD/GPx-1 mRNA levels decreased in HU rat cerebral arteries but not in mesenteric arteries. NADPH oxidases were activated significantly more in cerebral but not in mesenteric arteries of HU rats. NADPH oxidase inhibition with apocynin attenuated cerebrovascular ROS production and partially restored Nox2/Nox4 protein and mRNA levels, NADPH oxidase activity, and MnSOD/GPx-1 mRNA levels in cerebral VSMCs of HU rats. Conclusion These results suggest that vascular NADPH oxidases regulate cerebrovascular redox status and participate in vascular oxidative stress injury during simulated microgravity.展开更多
OBJECTIVE The Ca2+-activated Cl-channel(Ca CC)plays a crucial role in various physiological functions.Recent evidences suggest TMEM16A encodes CaC C in various cells,including endothelial cells.However,the role of TME...OBJECTIVE The Ca2+-activated Cl-channel(Ca CC)plays a crucial role in various physiological functions.Recent evidences suggest TMEM16A encodes CaC C in various cells,including endothelial cells.However,the role of TMEM16A in the vascular endothelial dysfunction in hypertension is unclear.METHODS In the study,RT-PCR,Western blotting,co-immunopricipitation,confocal imaging,patch-clamp,and endothelial-specific TMEM16A transgenic and knockout mice were employed.RESULTS We found that TMEM16A was expressed abundantly and functioned as Ca CC in endothelial cells.AngiotensinⅡ(AngⅡ)induced endothelial dysfunction with an increase in TMEM16A expression,which was alleviated by TMEM16A inhibitor.Further studies revealed that TMEM16A endothelial-specific knockout significantly lowered the blood pressure and ameliorated endothelial dysfunction in AngⅡ-induced hypertension,whereas,TMEM16A endothelial-specific overexpression showed the opposite effects.These results were related to the increased reactive oxygen species(ROS)generation,NADPH oxidase activation,and Nox2,p22phox expression facilitated by TMEM16A upon AngⅡ-induced hypertensive challenges.Moreover,TMEM16A directly interacted with Nox2 monomer and reduced the degradation of Nox2 through the proteasome-dependent endoplasmic recticulum-associated degradation pathway.TMEM16A also potentiated the translocation of p47phox and p67phox from cytosol to cell membrane and the subsequent interaction with Nox2.CONCLUSION Our results demonstrated that TMEM16A,as Ca CC,is a positive regulator of ROS generation via upregulating the activation of Nox2 NADPH oxidase in the vascular endothelium,and therefore facilitates endothelial dysfunction and hypertension.Modification of TMEM16A may be a novel therapeutic strategy for endothelial dysfunction-associated cardiovascular diseases.展开更多
Neuroinflammation,mediated by the nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing-3(NLRP3)inflammasome,is a significant contributor to the pathogenesis of neurodegenerative disea...Neuroinflammation,mediated by the nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing-3(NLRP3)inflammasome,is a significant contributor to the pathogenesis of neurodegenerative diseases(NDDs).Reynos-in,a natural sesquiterpene lactone(SL),exhibits a broad spectrum of pharmacological effects,suggesting its potential therapeutic value.However,the effects and mechanism of reynosin on neuroinflammation remain elusive.The current study explores the effects and mechanisms of reynosin on neuroinflammation using mice and BV-2 microglial cells treated with lipopolysaccharide(LPS).Our findings reveal that reynosin effectively reduces microglial inflammation in vitro,as demonstrated by decreased CD11b expression and lowered interleukin-1 beta(IL-1β)and interleukin-18(IL-18)mRNA and protein levels.Correspondingly,in vivo,results showed a re-duction in the number of Iba-1 positive cells and alleviation of morphological alterations,alongside decreased expressions of IL-1βand IL-18.Further analysis indicates that reynosin inhibits NLRP3 inflammasome activation,evidenced by reduced transcription of NLRP3 and caspase-1,diminished NLRP3 protein expression,inhibited apoptosis-associated speck-like protein containing a CARD(ASC)oli-gomerization,and decreased caspase-1 self-cleavage.Additionally,reynosin curtailed the activation of nicotinamide adenine dinuc-leotide phosphate(NADPH)oxidase,demonstrated by reduced NADP^(+)and NADPH levels,downregulation of gp91^(phox) mRNA,pro-tein expression,suppression of p47^(phox) expression and translocation to the membrane.Moreover,reynosin exhibited a neuroprotective effect against microglial inflammation in vivo and in vitro.These collective findings underscore reynosin’s capacity to mitigate mi-croglial inflammation by inhibiting the NLRP3 inflammasome,thus highlighting its potential as a therapeutic agent for managing neuroinflammation.展开更多
Transcription of human immunodeficiency virus (HIV-1) is activated by viral Tat protein which regulates HIV-long terminal reapaet (LTR) transcription and elongation.
Parkinson's disease is primarily caused by the loss of dopaminergic neurons in the substantia nigra compacta.Ferroptosis,a novel form of regulated cell death characterized by iron accumulation and lipid peroxidati...Parkinson's disease is primarily caused by the loss of dopaminergic neurons in the substantia nigra compacta.Ferroptosis,a novel form of regulated cell death characterized by iron accumulation and lipid peroxidation,plays a vital role in the death of dopaminergic neurons.However,the molecular mechanisms underlying ferroptosis in dopaminergic neurons have not yet been completely elucidated.NADPH oxidase 4 is related to oxidative stress,however,whether it regulates dopaminergic neuronal ferroptosis remains unknown.The aim of this study was to determine whether NADPH oxidase 4 is involved in dopaminergic neuronal ferroptosis,and if so,by what mechanism.We found that the transcriptional regulator activating transcription factor 3 increased NADPH oxidase 4 expression in dopaminergic neurons and astrocytes in an 1-methyl-4-phenyl-1,2,3,6 tetrahydropyridine-induced Parkinson's disease model.NADPH oxidase 4 inhibition improved the behavioral impairments observed in the Parkinson's disease model animals and reduced the death of dopaminergic neurons.Moreover,NADPH oxidase 4 inhibition reduced lipid peroxidation and iron accumulation in the substantia nigra of the Parkinson's disease model animals.Mechanistically,we found that NADPH oxidase 4 interacted with activated protein kinase Cαto prevent ferroptosis of dopaminergic neurons.Furthermore,by lowering the astrocytic lipocalin-2 expression,NADPH oxidase 4 inhibition reduced 1-methyl-4-phenyl-1,2,3,6 tetrahydropyridine-induced neuroinflammation.These findings demonstrate that NADPH oxidase 4 promotes ferroptosis of dopaminergic neurons and neuroinflammation,which contribute to dopaminergic neuron death,suggesting that NADPH oxidase 4 is a possible therapeutic target for Parkinson's disease.展开更多
The incomplete degradation of tumour cells by macrophages(Mϕ)is a contributing factor to tumour progression and metastasis,and the degradation function of Mϕis mediated through phagosomes and lysosomes.In our prelimin...The incomplete degradation of tumour cells by macrophages(Mϕ)is a contributing factor to tumour progression and metastasis,and the degradation function of Mϕis mediated through phagosomes and lysosomes.In our preliminary experiments,we found that overactivation of NADPH oxidase 2(NOX2)reduced the ability of Mϕto degrade engulfed tumour cells.Above this,we screened out liquiritin from Glycyrrhiza uralensis Fisch,which can significantly inhibit NOX2 activity and inhibit tumours,to elucidate that suppressing NOX2 can enhance the ability of Mϕto degrade tumour cells.We found that the tumour environment could activate the NOX2 activity in Mϕphagosomes,causing Mϕto produce excessive reactive oxygen species(ROS),thus prohibiting the formation of phagolysosomes before degradation.Conversely,inhibiting NOX2 in Mϕby liquiritin can reduce ROS and promote phagosome-lysosome fusion,therefore improving the enzymatic degradation of tumour cells after phagocytosis,and subsequently promote T cell activity by presenting antigens.We further confirmed that liquiritin down-regulated the expression of the NOX2 specific membrane component protein gp91 phox,blocking its binding to the NOX2 cytoplasmic component proteins p67 phox and p47 phox,thereby inhibiting the activity of NOX2.This study elucidates the specific mechanism by which Mϕcannot degrade tumour cells after phagocytosis,and indicates that liquiritin can promote the ability of Mϕto degrade tumour cells by suppressing NOX2.展开更多
The human endogenous retrovirus type W envelope glycoprotein(ERVWE1),located at chromosome 7q21–22,has been implicated in the pathophysiology of schizophrenia.Our previous studies have shown elevated ERVWE1 expressio...The human endogenous retrovirus type W envelope glycoprotein(ERVWE1),located at chromosome 7q21–22,has been implicated in the pathophysiology of schizophrenia.Our previous studies have shown elevated ERVWE1 expression in schizophrenia patients.Growing evidence suggests that autophagy dysfunction contributes to schizophrenia,yet the relationship between ERVWE1 and autophagy remains unclear.In this study,bioinformatics analysis of the human prefrontal cortex RNA microarray dataset(GSE53987)revealed that differentially expressed genes were predominantly enriched in autophagy-related pathways.Clinical data further demonstrated that serum levels of microtubuleassociated protein 1 light chain 3β(LC3B),a key marker of macroautophagy,were significantly elevated in schizophrenia patients compared to controls,and positively correlated with ERVWE1 expression.Cellular and molecular experiments suggested that ERVWE1 promoted macroautophagy by increasing the LC3B II/I ratio,enhancing autophagosome formation,and reducing sequestosome 1(SQSTM1)expression via upregulation of NADPH oxidase activator 1(NOXA1).Concurrently,NOXA1 downregulated the expression of key micromitophagy-related genes,including PTEN-induced kinase 1(PINK1),Parkin RBR E3 ubiquitin-protein ligase(Parkin),and the pyruvate dehydrogenase E1 subunitα1(PDHA1).As a result,ERVWE1,via NOXA1,inhibited micromitophagy by suppressing the expression of PINK1,Parkin,and PDHA1,thereby leading to impaired production of mitochondrialderived vesicles(MDVs).Mechanistically,ERVWE1 enhanced NOXA1 transcription by upregulating upstream transcription factor 2(USF2).In conclusion,ERVWE1 promotes macroautophagy and inhibits micromitophagy through USF2-NOXA1 axis,providing novel mechanistic insight into the role autophagy dysregulation in schizophrenia.These findings suggest that targeting autophagy pathways may offer novel therapeutic strategies for schizophrenia treatment.展开更多
In order to reveal the signaling pathways triggered by elicitor in plant-microbe interactions, the mechanisms of hypersensitive necrosis responses in Nicotiana tabacum L. cv. Gexin III induced by palmin were studied a...In order to reveal the signaling pathways triggered by elicitor in plant-microbe interactions, the mechanisms of hypersensitive necrosis responses in Nicotiana tabacum L. cv. Gexin III induced by palmin were studied at molecular and cellular level. The burst of superoxide, intercellular diffusion of hydrogen peroxide and process of cell death induced by palmin were investigated in tobacco plants by biochemical methods and Confocal microscopy. The results showed that a large amount of O-2(.-) was rapidly generated in tobacco cell elicited by palmin as a result of activation of NADPH oxidase, and the O-2(.-) was dismutated into H2O2 immediately by superoxide dismutase (SOD). Accumulation and intercellular diffusion of H2O2 were shown to be a trigger for hypersensitive cell death; and Ca2+ and some specific protein kinase were also shown to be involved in the activation of oxidative burst in tobacco cell induced by palmin.展开更多
OBJECTIVE:To assess the effect of Jianpi Huazhuo Tiaozhi granules(JHTG)on oxidative stress damage to macrophages and explore the relationship between the levels of this damage and the nicotinamide adenine dinucleotide...OBJECTIVE:To assess the effect of Jianpi Huazhuo Tiaozhi granules(JHTG)on oxidative stress damage to macrophages and explore the relationship between the levels of this damage and the nicotinamide adenine dinucleotide phosphate oxidase(NOX)/reactive oxygen species(ROS)-nuclear transcription factor kappa B(NF-κB)signaling pathway.METHODS:Macrophages cultured in vitro were divided into seven groups:control,model control,inhibitor,positive control,2.5%JHTG,5%JHTG,and10%JHTG.An oxidative stress injury model was established by stimulating macrophages with oxidized low-density lipoprotein.Cell survival and apoptosis were detected by 3-(4,5)-dimethylthiahiazo(-z-y1)-3,5-di-phenytetrazoliumromide and flow cytometry assays,respectively.Malondialdehyde and superoxide dismutase levels were detected by enzyme-linked immunosorbent assays,while ROS levels were detected using a fluorescence probe.Proteins and m RNAs associated with the NOX/ROS-NF-κB pathway,including NOX4,p22 phox,inhibitor of NF-κB kinase-α(IKK-α),inhibitor of NF-κB kinase-β(IKK-β),and NF-κB were detected by Western blot and PCR,respectively.RESULTS:After JHTG treatment,there were fewer damaged and apoptotic macrophages,while superoxide dismutase levels were elevated.The JHTG-treated groups also showed reduced ROS levels.The molecular changes following JHTG treatment included decreased expression of NOX4 and p22 phox at the protein level and decreased IKK-α,IKK-β,and NF-κB expression at the m RNA level.All of these effects were correlated with the JHTG concentration.CONCLUSION:These results demonstrated that the molecular mechanism underlying the antioxidant activity of JHTG in macrophages is through inhibiting the NOX/ROS-NF-κB pathway.展开更多
基金supported by the National Key R&D Program of China(2022YFD1401000).
文摘Poplar canker,caused by the fungus Cytospora chrysosperma,results in tremendous losses in poplar plantations in China.Although NADPH oxidases(NOXs)play important roles in the development and pathogenicity of several pathogenic fungi,their roles in C.chrysosperma remain unclear.In this study,we characterized three NOX genes(CcNox1,CcNox2,and CcNoxR)in C.chrysosperma.All three genes were highly upregulated during poplar branch infection,and deletion of any of them severely reduced virulence on poplar branches.Furthermore,deletion of either CcNox1 or CcNoxR resulted in a significant increase in endogenous reactive oxygen species production in hyphae,enhanced influx of Ca^(2+),the disruption of redox homeostasis and compromised mitochondrial integrity.Moreover,biosynthesis and secretion of a known virulence factor oxalic acid was obviously defective and exogenous oxalic acid supplementation rescued the virulence of the mutants.Taken together,our findings reveal that NOXs play important roles in redox homeostasis,mitochondrial integrity and pathogenicity in C.chrysosperma.
文摘Endogenous elicitor, termed cellulase-degraded cell wall (CDW), was prepared from the cell wall of suspension-cultured ginseng (Panax ginseng C.A. Meyer) cells via cellulase degradation. CDW activated the NADPH oxidase activity of isolated plasma membranes and stimulated in vivo H2O2 generation in ginseng cell suspensions. CDW also increased the activity of phenylalanine ammonia lyase (PAL), expression of a P. ginseng squalene epoxidase (sqe) gene and saponin synthesis. NADPH oxidase inhibitors inhibited both in vitro NADPH oxidase activity and in vivo H2O2 generation. Induction of PAL activity, saponin synthesis and sqe gene expression were all inhibited by such inhibitor treatments and reduced by incubation with catalase and HA scavengers. These data indicate that activation of NADPH oxidase and generation of H2O2 are essential signalling events mediating defence responses induced by the endogenous elicitor(s) present in CDW.
基金supported by the National Natural Science Foundation of China,No.81671125the Natural Science Foundation of Guangdong Province,No.2021A1515011115Guangzhou Science and Technology Project,No.202102010346(all to YZC)。
文摘Intracerebral hemorrhage is often accompanied by oxidative stress induced by reactive oxygen species,which causes abnormal mitochondrial function and secondary reactive oxygen species generation.This creates a vicious cycle leading to reactive oxygen species accumulation,resulting in progression of the pathological process.Therefore,breaking the cycle to inhibit reactive oxygen species accumulation is critical for reducing neuronal death after intracerebral hemorrhage.Our previous study found that increased expression of nicotinamide adenine dinucleotide phosphate oxidase 4(NADPH oxidase 4,NOX4)led to neuronal apoptosis and damage to the blood-brain barrier after intracerebral hemorrhage.The purpose of this study was to investigate the role of NOX4 in the circle involving the neuronal tolerance to oxidative stress,mitochondrial reactive oxygen species and modes of neuronal death other than apoptosis after intracerebral hemorrhage.We found that NOX4 knockdown by adeno-associated virus(AAV-NOX4)in rats enhanced neuronal tolerance to oxidative stress,enabling them to better resist the oxidative stress caused by intracerebral hemorrhage.Knockdown of NOX4 also reduced the production of reactive oxygen species in the mitochondria,relieved mitochondrial damage,prevented secondary reactive oxygen species accumulation,reduced neuronal pyroptosis and contributed to relieving secondary brain injury after intracerebral hemorrhage in rats.Finally,we used a mitochondria-targeted superoxide dismutase mimetic to explore the relationship between reactive oxygen species and NOX4.The mitochondria-targeted superoxide dismutase mimetic inhibited the expression of NOX4 and neuronal pyroptosis,which is similar to the effect of AAV-NOX4.This indicates that NOX4 is likely to be an important target for inhibiting mitochondrial reactive oxygen species production,and NOX4 inhibitors can be used to alleviate oxidative stress response induced by intracerebral hemorrhage.
基金supported by the Research Fund of Ege University(Project No.2010-TIP-076)
文摘Increased reactive oxygen species by the activation of NADPH oxidase（NOX） contributes to the development of diabetic complications.Apocynin,a NOX inhibitor,increases sciatic nerve conductance and blood flow in diabetic rats.We investigated potential protective effect of apocynin in rat diabetic neuropathy and its precise mechanism of action at molecular level.Rat models of streptozotocin-induced diabetes were treated with apocynin（30 and 100 mg/kg per day,intragastrically） for 4 weeks.Mechanical hyperalgesia and allodynia were determined weekly using analgesimeter and dynamic plantar aesthesiometer.Western blot analysis and histochemistry/immunohistochemistry were performed in the lumbar spinal cord and sciatic nerve respectively.Streptozotocin injection reduced pain threshold in analgesimeter,but not in aesthesiometer.Apocynin treatment increased pain threshold dose-dependently.Western blot analysis showed an increase in catalase and NOX-p47 phox protein expression in the spinal cord.However,protein expressions of neuronal and inducible nitric oxide synthase（n NOS,i NOS）,superoxide dismutase,glutathion peroxidase,nitrotyrosine,tumor necrosis factor-α,interleukin-6,interleukin-1β,aldose reductase,cyclooxygenase-2 or MAC-1（marker for increased microgliosis） in the spinal cord remained unchanged.Western blot analysis results also demonstrated that apocynin decreased NOX-p47 phox expression at both doses and catalase expression at 100 mg/kg per day.Histochemistry of diabetic sciatic nerve revealed marked degeneration.n NOS and i NOS immunoreactivities were increased,while S-100 immunoreactivity（Schwann cell marker） was decreased in sciatic nerve.Apocynin treatment reversed these changes dose-dependently.In conclusion,decreased pain threshold of diabetic rats was accompanied by increased NOX and catalase expression in the spinal cord and increased degeneration in the sciatic nerve characterized by increased NOS expression and Schwann cell loss.Apocynin treatment attenuates neuropathic pain by decelerating the increased oxidative stress-mediated pathogenesis in diabetic rats.
基金supported by the National Major Scientific and Technological Special Project for Significant New Drugs Development(2019zx09301102)the Project of Liaoning Provincial Department of Education(LJKZ0826)the Open Project of National and Local Joint Engineering Research Center for Drug Research and Development of Neurodegenerative Diseases(2022GCYJZX-YB02).
文摘Neuroinflammation mediated by microglia and oxidative stress play pivotal roles in the development of chronic temporal lobe epilepsy(TLE).We postulated that kainic acid(KA)-Induced status epilepticus triggers microglia-dependent inflammation,leading to neuronal damage,a lowered seizure threshold,and the emergence of spontaneous recurrent seizures(SRS).Extensive evidence from our laboratory suggests that dextromethorphan(DM),even in ultra-low doses,has anti-inflammatory and neuroprotective effects in many animal models of neurodegenerative disease.Our results showed that administration of DM(10 ng/kg per day;subcutaneously via osmotic minipump for 4 weeks)significantly mitigated the residual effects of KA,including the frequency of SRS and seizure susceptibility.In addition,DM-treated rats showed improved cognitive function and reduced hippocampal neuronal loss.We found suppressed microglial activation-mediated neuroinflammation and decreased expression of hippocampal gp91^(phox) and p47^(phox) proteins in KA-induced chronic TLE rats.Notably,even after discontinuation of DM treatment,ultra-low doses of DM continued to confer long-term anti-seizure and neuroprotective effects,which were attributed to the inhibition of microglial NADPH oxidase 2 as revealed by mechanistic studies.
文摘A rapid and concentration-dependent generation of superoxide anion （·O2^-）, measured with a superoxide-specific Cypridina luciferin-derived chemiluminescent reagent, was observed when two lanthanide salts （LaCl3 and CdCl3 ） were added to tobacco （ Nicotiana tabacum） cell suspension culture. Addition of superoxide dismutase （480 U·ml^-1） and Tiron （5 μmol·L^-1） to cell culture suspension decreases the level of lanthanide cation-induced ·O2^- generation, suggesting that ·O2^- generation is extra-cellular. Pretreatment of the cell culture suspension with diphenyleneiodonium （10 and 50 μmol·L^-1 ）, quinacrine （ 1 and 5 mmol· L^-1 ） and imidazol （ 10 mmol· L^-1 ）, inhibitors of NADPH oxidase, notably inhibits the generation of superoxide induced by lanthanide cation, implying the possible involvement of activation of NADPH oxidase. In addition, addition of SHAM （1 and 5 mmol· L^-1）, azide （0.2 and 1 mmol· L^-1 ）, inhibitor of peroxidase, has no influence on ·O2^- generation.
基金supported by the National Research Foundation of the Republic of Korea 2018R1D1A3B07047960the Soonchunhyang University Research Fund(to SSY).
文摘Diseases like Alzheimer’s and Parkinson’s diseases are defined by inflammation and the damage neurons undergo due to oxidative stress. A primary reactive oxygen species contributor in the central nervous system, NADPH oxidase 4, is viewed as a potential therapeutic touchstone and indicative marker for these ailments. This in-depth review brings to light distinct features of NADPH oxidase 4, responsible for generating superoxide and hydrogen peroxide, emphasizing its pivotal role in activating glial cells, inciting inflammation, and disturbing neuronal functions. Significantly, malfunctioning astrocytes, forming the majority in the central nervous system, play a part in advancing neurodegenerative diseases, due to their reactive oxygen species and inflammatory factor secretion. Our study reveals that aiming at NADPH oxidase 4 within astrocytes could be a viable treatment pathway to reduce oxidative damage and halt neurodegenerative processes. Adjusting NADPH oxidase 4 activity might influence the neuroinflammatory cytokine levels, including myeloperoxidase and osteopontin, offering better prospects for conditions like Alzheimer’s disease and Parkinson’s disease. This review sheds light on the role of NADPH oxidase 4 in neural degeneration, emphasizing its drug target potential, and paving the path for novel treatment approaches to combat these severe conditions.
基金the National Natural Science Foundation of China(No.31622006)the Postdoctoral Science Foundation of China(Nos.2018M630683 and 2018T110601)
文摘Production of reactive oxygen species(ROS)is a conserved immune response primarily mediated by NADPH oxidases(NOXs),also known in plants as respiratory burst oxidase homologs(RBOHs).Most microbe-associated molecular patterns(MAMPs)trigger a very fast and transient ROS burst in plants.However,recently,we found that lipopolysaccharides(LPS),a typical bacterial MAMP,triggered a biphasic ROS burst.In this study,we isolated mutants defective in LPS-triggered biphasic ROS burst(delt)in Arabidopsis,and cloned the DELT1 gene that was shown to encode RBOHD.In the delt1-2 allele,the antepenultimate residue,glutamic acid(E919),at the C-terminus of RBOHD was mutated to lysine(K).E919 is a highly conserved residue in NADPH oxidases,and a mutation of the corresponding residue E568 in human NOX2 has been reported to be one of the causes of chronic granulomatous disease.Consistently,we found that residue E919 was indispensable for RBOHD function in the MAMP-induced ROS burst and stomatal closure.It has been suggested that the mutation of this residue in other NADPH oxidases impairs the protein’s stability and complex assembly.However,we found that the E919K mutation did not affect RBOHD protein abundance or the ability of protein association,suggesting that the residue E919 in RBOHD might have a regulatory mechanism different from that of other NOXs.Taken together,our results confirm that the antepenultimate residue E is critical for NADPH oxidases and provide a new insight into the regulatory mechanisms of RBOHD.
基金supported by grants from the Natural Science Foundation of Hubei province (No. 2010CDB08005)the National Natural Science Foundation of China (No. 30730094 and81000409)Special Funds for State Key Development Program for Basic Research of China (973 Program) (No.2011CB504504)
文摘Mitochondrial DNA(mtDNA) common deletion(CD) plays a significant role in aging and age-related diseases.In this study,we used D-galactose(D-gal) to generate an animal model of aging and the involvement and causative mechanisms of mitochondrial damage in such a model were investigated.Twenty 5-week-old male Sprague-Dawley rats were randomly divided into two groups:D-gal group(n=10) and control group(n=10).The quantity of the mtDNA CD in the hippocampus was determined using a TaqMan real-time PCR assay.Transmission electron microscopy was used to observe the mitochondrial ultrastructure in the hippocampus.Western blot was used to detect the protein levels of NADPH oxidase(NOX) and uncoupling protein 2(UCP2).We found that the level of mtDNA CD was significantly higher in the hippocampus of D-gal-induced aging rats than in control rats.In comparison with the control group,the mitochondrial ultrastructure in the hippocampus of D-gal-treated rats was damaged,and the protein levels of NOX and UCP2 were significantly increased in the hippocampus of D-gal-induced aging rats.This study demonstrated that the levels of mtDNA CD and NOX protein expression were significantly increased in the hippocampus of D-gal-induced aging rats.These findings indicate that NOX-dependent reactive oxygen species generation may contribute to D-gal-induced mitochondrial damage.
基金financially supported by grants from Shenzhen Science and Technology Innovation Commission of China,No.JCYJ20150330102401097,KQCX20140521101427034,JCYJ20140414170821291China Postdoctoral Science Foundation,No.2015M572388
文摘Excess production of reactive oxygen species （ROS） critically contributes to occurrence of reperfusion injury, the paradoxical response of ischemic brain tissue to restoration of cerebral blood flow. However, the enzymatic sources of ROS generation remain to be unclear. This study examined Nox2-ontaining NADPH oxidase （Nox2） expression and its activity in ischemic brain tissue following post-ischemic reperfusion to clarify the mechanism of enzymatic reaction of ROS. Male Sprague-Dawley rats were subjected to 90-minute middle cerebral artery occlusion, followed by 3 or 22.5 hours of reperfusion. Quantitative reverse transcriptase PCR and western blot assay were performed to measure mRNA and protein expression of Nox2. Lucigenin fluorescence assays were performed to assess Nox activity. Our data showed that Nox2 mRNA and protein expression levels were significantly increased （3.7-fold for mRNA and 3.6-fold for protein） in ischemic brain tissue at 22.5 hours but not at 3 hours following post-ischemic reperfusion. Similar results were obtained for the changes of NADPH oxidase activity in ischemic cerebral tissue at the two reperfusion time points. Our results suggest that Nox2 may not contribute to the early burst of reperfusion-related ROS generation, but is rather an important source of ROS generation during prolonged reperfusion.
基金supported by National Natural Science Foundation of China (No. 30871069)
文摘The antibody against AT1-EC2 plays a role in some kinds of inflammatory vascular diseases including malignant hypertension,preeclampsia,and renal-allograft rejection,but the detailed mechanisms remain unclear.In order to investigate the changes of NADPH oxidase and reactive oxygen species in the aorta in a mouse model which can produce AT1-EC2 antibody by active immunization with AT1-EC2 peptide,15 mice were divided into three groups:control group,AT1-EC2-immunized group,and AT1-EC2-immunized and valsartan-treated group.In AT1-EC2-immunized group and AT1-EC2-immunized and valsartan-treated group,the mice were immunized by 50 μg peptide subcutaneously at multiple points for 4 times:0,5,10,and 15 days after the experiment.In AT1-EC2-immunized and valsartan-treated group,valsartan was given at a dose of 100 mg/kg every day for 20 days.After the experiment,the mice were sacrificed under anesthesia and the aortas were obtained and frozen in liquid nitrogen for the preparation of frozen section slides and other experiments.The titer of AT1-EC2 was assayed by using ELISA.The level of NOX1 mRNA in the aorta was determined by using RT-PCR.The expression of NOX1 was detected by using Western blotting.Confocal scanning microscopy was used to assay the α-actin and NOX1 expression in the aortic tissue.The O 2.production was detected in situ after DHE staining.The mice produced high level antibody against AT1-EC2 in AT1-EC2-immunized group and AT1-EC2-immunized and valsartan-treated group,and the level of NOX1 mRNA in the aortic tissues was 1.6±0.4 times higher and the NOX1 protein expression was higher in AT1-EC2-immunized group than in control group.There were no significant differences in the level of NOX1 mRNA and protein expression between control group and AT1-EC2-immunized and valsartan-treated group.The expression and co-localization of α-actin and NOX1 in AT1-EC2-immunized group increased significantly as compared with those in control group,and the O 2.production increased about 2.7 times as compared with control group.There were no significant differences between control group and AT1-EC2-immunized and valsartan-treated group.It is concluded that active immunization with AT1-EC2 can activate NOX1-ROS,and increase vascular inflammation,which can be inhibited by AT1 receptor blocker valsartan.This may partially explain the mechanism of the pathogenesis of inflammatory vascular diseases related to antibody against AT1-EC2.
基金supported by the National Natural Science Foundation of China(Grant Nos.81101468&81571841)the Beijing NOVO Program(Grant No.XX2013105)
文摘Objective The roles of cerebrovascular oxidative stress in vascular functional remodeling have been described in hindlimb-unweighting （HU） rats. However, the underlying mechanism remains to be established. Methods We investigated the generation of vascular reactive oxygen species （ROS）, Nox2/Nox4 protein and mRNA levels, NADPH oxidase activity, and manganese superoxide dismutase （MnSOD） and glutathione peroxidase-1 （GPx-1） mRNA levels in cerebral and mesenteric smooth muscle cells （VSMCs） of HU rats. Results ROS production increased in cerebral but not in mesenteric VSMCs of HU rats compared with those in control rats. Nox2 and Nox4 protein and mRNA levels were increased significantly but MnSOD/GPx-1 mRNA levels decreased in HU rat cerebral arteries but not in mesenteric arteries. NADPH oxidases were activated significantly more in cerebral but not in mesenteric arteries of HU rats. NADPH oxidase inhibition with apocynin attenuated cerebrovascular ROS production and partially restored Nox2/Nox4 protein and mRNA levels, NADPH oxidase activity, and MnSOD/GPx-1 mRNA levels in cerebral VSMCs of HU rats. Conclusion These results suggest that vascular NADPH oxidases regulate cerebrovascular redox status and participate in vascular oxidative stress injury during simulated microgravity.
基金The project supported by National Natural Science Foundation of China(81230082,81302771,81525025,81573422,81500226)Natural Science Foundation of Guangdong Province(2014A030313087)by Science and Technology program of Guangzhou City(201607010255)
文摘OBJECTIVE The Ca2+-activated Cl-channel(Ca CC)plays a crucial role in various physiological functions.Recent evidences suggest TMEM16A encodes CaC C in various cells,including endothelial cells.However,the role of TMEM16A in the vascular endothelial dysfunction in hypertension is unclear.METHODS In the study,RT-PCR,Western blotting,co-immunopricipitation,confocal imaging,patch-clamp,and endothelial-specific TMEM16A transgenic and knockout mice were employed.RESULTS We found that TMEM16A was expressed abundantly and functioned as Ca CC in endothelial cells.AngiotensinⅡ(AngⅡ)induced endothelial dysfunction with an increase in TMEM16A expression,which was alleviated by TMEM16A inhibitor.Further studies revealed that TMEM16A endothelial-specific knockout significantly lowered the blood pressure and ameliorated endothelial dysfunction in AngⅡ-induced hypertension,whereas,TMEM16A endothelial-specific overexpression showed the opposite effects.These results were related to the increased reactive oxygen species(ROS)generation,NADPH oxidase activation,and Nox2,p22phox expression facilitated by TMEM16A upon AngⅡ-induced hypertensive challenges.Moreover,TMEM16A directly interacted with Nox2 monomer and reduced the degradation of Nox2 through the proteasome-dependent endoplasmic recticulum-associated degradation pathway.TMEM16A also potentiated the translocation of p47phox and p67phox from cytosol to cell membrane and the subsequent interaction with Nox2.CONCLUSION Our results demonstrated that TMEM16A,as Ca CC,is a positive regulator of ROS generation via upregulating the activation of Nox2 NADPH oxidase in the vascular endothelium,and therefore facilitates endothelial dysfunction and hypertension.Modification of TMEM16A may be a novel therapeutic strategy for endothelial dysfunction-associated cardiovascular diseases.
基金supported by the National Natural Science Foundation of China(No.82174076)the Construction Project of Liaoning Provincial Key Laboratory,China(No.2022JH13/10200026)+2 种基金the Fundamental Research Funds for the Central Universities(No.N2220002),the 111 Project(No.B16009)Scientific Research Fund of Liaoning Province Education Department(No.LJKZ0945)Natural Science Foundation of Liaoning Province(No.2022-MS-242).
文摘Neuroinflammation,mediated by the nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing-3(NLRP3)inflammasome,is a significant contributor to the pathogenesis of neurodegenerative diseases(NDDs).Reynos-in,a natural sesquiterpene lactone(SL),exhibits a broad spectrum of pharmacological effects,suggesting its potential therapeutic value.However,the effects and mechanism of reynosin on neuroinflammation remain elusive.The current study explores the effects and mechanisms of reynosin on neuroinflammation using mice and BV-2 microglial cells treated with lipopolysaccharide(LPS).Our findings reveal that reynosin effectively reduces microglial inflammation in vitro,as demonstrated by decreased CD11b expression and lowered interleukin-1 beta(IL-1β)and interleukin-18(IL-18)mRNA and protein levels.Correspondingly,in vivo,results showed a re-duction in the number of Iba-1 positive cells and alleviation of morphological alterations,alongside decreased expressions of IL-1βand IL-18.Further analysis indicates that reynosin inhibits NLRP3 inflammasome activation,evidenced by reduced transcription of NLRP3 and caspase-1,diminished NLRP3 protein expression,inhibited apoptosis-associated speck-like protein containing a CARD(ASC)oli-gomerization,and decreased caspase-1 self-cleavage.Additionally,reynosin curtailed the activation of nicotinamide adenine dinuc-leotide phosphate(NADPH)oxidase,demonstrated by reduced NADP^(+)and NADPH levels,downregulation of gp91^(phox) mRNA,pro-tein expression,suppression of p47^(phox) expression and translocation to the membrane.Moreover,reynosin exhibited a neuroprotective effect against microglial inflammation in vivo and in vitro.These collective findings underscore reynosin’s capacity to mitigate mi-croglial inflammation by inhibiting the NLRP3 inflammasome,thus highlighting its potential as a therapeutic agent for managing neuroinflammation.
基金Funded by National Natural Sciences Foundation of China (No. 30800580Beijing Nova Program (No. 2007B014)+3 种基金Beijing Natural Science Foundation (No. 5093025)Beijing Talents Foundation (No. 20071D0501500213)BJUT Science Foundation for Youths (No. 97015999200702)BJUT Scientific Research Starting Foundation for Doctor (No. 52015999200703)
文摘Transcription of human immunodeficiency virus (HIV-1) is activated by viral Tat protein which regulates HIV-long terminal reapaet (LTR) transcription and elongation.
基金supported by the National Natural Science Foundation of China,Nos.82271444(to JP),82271268(to BZ),and 82001346(to YL)the National Key Research and Development Program of China,No.2022YFE0210100(to BZ)。
文摘Parkinson's disease is primarily caused by the loss of dopaminergic neurons in the substantia nigra compacta.Ferroptosis,a novel form of regulated cell death characterized by iron accumulation and lipid peroxidation,plays a vital role in the death of dopaminergic neurons.However,the molecular mechanisms underlying ferroptosis in dopaminergic neurons have not yet been completely elucidated.NADPH oxidase 4 is related to oxidative stress,however,whether it regulates dopaminergic neuronal ferroptosis remains unknown.The aim of this study was to determine whether NADPH oxidase 4 is involved in dopaminergic neuronal ferroptosis,and if so,by what mechanism.We found that the transcriptional regulator activating transcription factor 3 increased NADPH oxidase 4 expression in dopaminergic neurons and astrocytes in an 1-methyl-4-phenyl-1,2,3,6 tetrahydropyridine-induced Parkinson's disease model.NADPH oxidase 4 inhibition improved the behavioral impairments observed in the Parkinson's disease model animals and reduced the death of dopaminergic neurons.Moreover,NADPH oxidase 4 inhibition reduced lipid peroxidation and iron accumulation in the substantia nigra of the Parkinson's disease model animals.Mechanistically,we found that NADPH oxidase 4 interacted with activated protein kinase Cαto prevent ferroptosis of dopaminergic neurons.Furthermore,by lowering the astrocytic lipocalin-2 expression,NADPH oxidase 4 inhibition reduced 1-methyl-4-phenyl-1,2,3,6 tetrahydropyridine-induced neuroinflammation.These findings demonstrate that NADPH oxidase 4 promotes ferroptosis of dopaminergic neurons and neuroinflammation,which contribute to dopaminergic neuron death,suggesting that NADPH oxidase 4 is a possible therapeutic target for Parkinson's disease.
基金supported by National Natural Science Foundation of China(Grant No.:82074092)Natural Science Foundation of Guangdong Province,China(Grant No.:2023A1515011141)+1 种基金Research projects in Key Fields of Universities in Guangdong Province,China(Grant No.:2023ZDZX2020)Guangzhou Municipal Bureau of Science and Technology,China(Grant No.:202201011631).
文摘The incomplete degradation of tumour cells by macrophages(Mϕ)is a contributing factor to tumour progression and metastasis,and the degradation function of Mϕis mediated through phagosomes and lysosomes.In our preliminary experiments,we found that overactivation of NADPH oxidase 2(NOX2)reduced the ability of Mϕto degrade engulfed tumour cells.Above this,we screened out liquiritin from Glycyrrhiza uralensis Fisch,which can significantly inhibit NOX2 activity and inhibit tumours,to elucidate that suppressing NOX2 can enhance the ability of Mϕto degrade tumour cells.We found that the tumour environment could activate the NOX2 activity in Mϕphagosomes,causing Mϕto produce excessive reactive oxygen species(ROS),thus prohibiting the formation of phagolysosomes before degradation.Conversely,inhibiting NOX2 in Mϕby liquiritin can reduce ROS and promote phagosome-lysosome fusion,therefore improving the enzymatic degradation of tumour cells after phagocytosis,and subsequently promote T cell activity by presenting antigens.We further confirmed that liquiritin down-regulated the expression of the NOX2 specific membrane component protein gp91 phox,blocking its binding to the NOX2 cytoplasmic component proteins p67 phox and p47 phox,thereby inhibiting the activity of NOX2.This study elucidates the specific mechanism by which Mϕcannot degrade tumour cells after phagocytosis,and indicates that liquiritin can promote the ability of Mϕto degrade tumour cells by suppressing NOX2.
基金supported by the National Natural Science Foundation of China(No.82272321)the Stanley Foundation from the Stanley Medical Research Institute(SMRI),United States(No.06R-1366)。
文摘The human endogenous retrovirus type W envelope glycoprotein(ERVWE1),located at chromosome 7q21–22,has been implicated in the pathophysiology of schizophrenia.Our previous studies have shown elevated ERVWE1 expression in schizophrenia patients.Growing evidence suggests that autophagy dysfunction contributes to schizophrenia,yet the relationship between ERVWE1 and autophagy remains unclear.In this study,bioinformatics analysis of the human prefrontal cortex RNA microarray dataset(GSE53987)revealed that differentially expressed genes were predominantly enriched in autophagy-related pathways.Clinical data further demonstrated that serum levels of microtubuleassociated protein 1 light chain 3β(LC3B),a key marker of macroautophagy,were significantly elevated in schizophrenia patients compared to controls,and positively correlated with ERVWE1 expression.Cellular and molecular experiments suggested that ERVWE1 promoted macroautophagy by increasing the LC3B II/I ratio,enhancing autophagosome formation,and reducing sequestosome 1(SQSTM1)expression via upregulation of NADPH oxidase activator 1(NOXA1).Concurrently,NOXA1 downregulated the expression of key micromitophagy-related genes,including PTEN-induced kinase 1(PINK1),Parkin RBR E3 ubiquitin-protein ligase(Parkin),and the pyruvate dehydrogenase E1 subunitα1(PDHA1).As a result,ERVWE1,via NOXA1,inhibited micromitophagy by suppressing the expression of PINK1,Parkin,and PDHA1,thereby leading to impaired production of mitochondrialderived vesicles(MDVs).Mechanistically,ERVWE1 enhanced NOXA1 transcription by upregulating upstream transcription factor 2(USF2).In conclusion,ERVWE1 promotes macroautophagy and inhibits micromitophagy through USF2-NOXA1 axis,providing novel mechanistic insight into the role autophagy dysregulation in schizophrenia.These findings suggest that targeting autophagy pathways may offer novel therapeutic strategies for schizophrenia treatment.
文摘In order to reveal the signaling pathways triggered by elicitor in plant-microbe interactions, the mechanisms of hypersensitive necrosis responses in Nicotiana tabacum L. cv. Gexin III induced by palmin were studied at molecular and cellular level. The burst of superoxide, intercellular diffusion of hydrogen peroxide and process of cell death induced by palmin were investigated in tobacco plants by biochemical methods and Confocal microscopy. The results showed that a large amount of O-2(.-) was rapidly generated in tobacco cell elicited by palmin as a result of activation of NADPH oxidase, and the O-2(.-) was dismutated into H2O2 immediately by superoxide dismutase (SOD). Accumulation and intercellular diffusion of H2O2 were shown to be a trigger for hypersensitive cell death; and Ca2+ and some specific protein kinase were also shown to be involved in the activation of oxidative burst in tobacco cell induced by palmin.
基金Supported by National Natural Science Foundation of China Jianpi Huazhuo Tiaozhi Granules Based on the Theory of Turbidity Adjust and Control NOX/ROS-NF-k B Signaling Pathway and the Mechanism for the Treatment of Atherosclerosis(No.81660781)the Innovation Special Fund Project of Graduate StudentsJiangxi Province Study on the Mechanism of Treating Atherosclerosis by Jianpi Huazhuo Tiaozhi Granules(No.YC2017-B085)。
文摘OBJECTIVE:To assess the effect of Jianpi Huazhuo Tiaozhi granules(JHTG)on oxidative stress damage to macrophages and explore the relationship between the levels of this damage and the nicotinamide adenine dinucleotide phosphate oxidase(NOX)/reactive oxygen species(ROS)-nuclear transcription factor kappa B(NF-κB)signaling pathway.METHODS:Macrophages cultured in vitro were divided into seven groups:control,model control,inhibitor,positive control,2.5%JHTG,5%JHTG,and10%JHTG.An oxidative stress injury model was established by stimulating macrophages with oxidized low-density lipoprotein.Cell survival and apoptosis were detected by 3-(4,5)-dimethylthiahiazo(-z-y1)-3,5-di-phenytetrazoliumromide and flow cytometry assays,respectively.Malondialdehyde and superoxide dismutase levels were detected by enzyme-linked immunosorbent assays,while ROS levels were detected using a fluorescence probe.Proteins and m RNAs associated with the NOX/ROS-NF-κB pathway,including NOX4,p22 phox,inhibitor of NF-κB kinase-α(IKK-α),inhibitor of NF-κB kinase-β(IKK-β),and NF-κB were detected by Western blot and PCR,respectively.RESULTS:After JHTG treatment,there were fewer damaged and apoptotic macrophages,while superoxide dismutase levels were elevated.The JHTG-treated groups also showed reduced ROS levels.The molecular changes following JHTG treatment included decreased expression of NOX4 and p22 phox at the protein level and decreased IKK-α,IKK-β,and NF-κB expression at the m RNA level.All of these effects were correlated with the JHTG concentration.CONCLUSION:These results demonstrated that the molecular mechanism underlying the antioxidant activity of JHTG in macrophages is through inhibiting the NOX/ROS-NF-κB pathway.
