Inactivated Sendai Virus Induces Apoptosis Mediated by Reactive Oxygen Species in Murine Melanoma Cells

Objective This paper aims to investigate the apoptotic effect of inactivated Sendai virus （hemagglutinating virus of Japan-enveloped, HVJ-E） on routine melanoma cells （B16FlO） and the possible mechanisms involved in the putative apoptotic reactions. Methods B16F10 cells were treated with HVJ-E at various multiplicities of infection （MOI）, and the reactive oxygen species （ROS）, cell viability, and apoptosis were measured. Next, the roles of ROS in the regulation of Bcl-2/Bax and the activation of mitogen-activated protein kinase （MAPK） pathways in HVJ-E-treated B16F10 cells were analyzed. To further evaluate the cytotoxic effect of HVJ-E-generated ROS on B16FlO cells, HVJ-E was intratumorally injected, both with and without N-acetyI-L-cysteine （NAC）, into melanoma tumors on BALB/c mice. Tumor volume was then monitored for 3 weeks, and the tumor proteins were separated for immunoblot assay. Results Treatment of B16F10 cells with HVJ-E resulted in a dose-dependent inhibition of cell-viability and an induction of apoptosis. The latter effect was associated with the generation of ROS. Inhibition of ROS generation by NAC resulted in a significant reduction of HVJ-E-induced Erkl/2, JNK, and p38 MAPK activation. Additionally, ROS inhibition caused a decrease in the Bcl-2/Bax ratio as well as promoting activation of apoptosis both in vitro and in vivo. Conclusion These results suggest that HVJ-E possesses potential anticancer activity in B16F10 cells through ROS-mediated mitochondrial dysfunction involving the MAPK pathway.

Chaetocin reactivates the lytic replication of Epstein-Barr virus from latency via reactive oxygen species

Oxidative stress,regarded as a negative effect of free radicals in vivo,takes place when organisms suffer from harmful stimuli.Some viruses can induce the release of reactive oxygen species(ROS) in infected cells,which may be closely related with their pathogenicity.In this report,chaetocin,a fungal metabolite reported to have antimicrobial and cytostatic activity,was studied for its effect on the activation of latent Epstein-Barr virus(EBV) in B95-8 cells.We found that chaetocin remarkably up-regulated EB V lytic transcription and DNA replication at a low concentration(50 nmol L^(-1).The activation of latent EBV was accompanied by an increased cellular ROS level.N-acetyl-L-cy steine(NAC),an ROS inhibitor,suppressed chaetocin-induced EBV activation.Chaetocin had little effect on histone H3K9 methylation,while NAC also significantly reduced H3K9 methylation.These results suggested that chaetocin reactivates latent EBV primarily via ROS pathways.

HBx-induced reactive oxygen species activates hepatocellular carcinogenesis via dysregulation of PTEN/Akt pathway

AIM:To investigate the role of hepatitis B virus X-protein(HBx)-induced reactive oxygen species(ROS)on liver carcinogenesis in HBx transgenic mice and HepG2-HBx cells.METHODS:Cell growth rate was analyzed,and through western blotting,mitogenic signaling was observed.Endogenous ROS from wild and HBx transgenic mice and HepG2-Mock and HBx cells were assayed by FACS-calibur.Identification of oxidized and reduced phosphatase and tensin homolog(PTEN)was analyzed through N-ethylmaleimide alkylation,nonreducing electrophoresis.RESULTS:We observed that the cell-proliferation-related phosphoinositide 3-kinase/Akt pathway is activated by HBx in vivo and in vitro.Increased ROS were detected by HBx.Tumor suppressor PTEN,via dephosphorylation of Akt,was oxidized and inactivated by increased ROS.Increased oxidized PTEN activated the mitogenic pathway through over-activated Akt.However,treatment with ROS scavenger N-acetyl cysteine can reverse PTEN to a reduced form.Endogenously produced ROS also stimulated HBx expression.CONCLUSION:HBx induced ROS promoted Akt pathways via oxidized inactive PTEN.HBx and ROS maintained a positive regulatory loop,which aggravated carcinogenesis.

Suppressed effects of Phyllanthus urinaria L.ethyl acetate extract on hepatitis B virus both in vitro and in vivo

Background:Phyllanthus urinaria L.(P.urinaria)extract(PUE)has been used to inhibit hepatitis B virus(HBV).However,the underlying mechanism remains unclear.To investigate which PUE fractions and main components lead to against HBV and approach the relevant molecular mechanisms.Methods:P.urinaria was extracted with water,and then the decoction was extracted by petroleum ether,ethyl acetate,and n-butanol in turn.The HepG2.2.15 cell was treated with aqueous fraction,petroleum ether fraction,ethyl acetate fraction and n-butanol fraction,gallic acid(GA,C7H6O5)and corilagin(CL,C27H22O18),respectively.The medium was collected for hepatitis B surface antigen(HBsAg)and hepatitis B e antigen assays.Cell counting kit-8 method was used to identify cell proliferation.Also,the levels of cellular oxygen consumption,reactive oxygen species,and reduced glutathione were detected.The HBV modeling mice were treated with ethyl acetate fraction,entecavir and physiological saline,respectively.The serum was collected for HBsAg and inflammatory cytokines assays.Liver tissue metabolites were screened by LC-MS/MS method.Results:The ethyl acetate fraction(EAF)of P.urinaria could significantly inhibit HBV secretion in HepG2.2.15(P<0.05).Furthermore,two main constitutes in ethyl acetate fraction,GA and CL,could significantly inhibit HBV secretion and reduced cell proliferation(P<0.05).Also,GA and CL could increase cellular oxygen consumption,intracellular superoxide anions level,superoxide dismutase level and glutathione depletion.Compared with the Modeling group,EAF significantly decreased the expression levels of HBsAg,IL-1β,IFN-α(P<0.05).LC-MS/MS analysis results showed that EAF dramatically up-regulate hydroxyproline,maltotriose,betaine and down-regulate glutathione disulfide,taurocholate,taurochenodeoxycholate(P<0.05).Kyoto Encyclopedia of Genes and Genomes results show that the differential metabolites were mainly enriched in ATP-binding cassette transporters pathway.Conclusions:P.urinaria exhibits suppressed effects on HBV by modulating reactive oxygen species formation or metabolomics both in vitro and in vivo.These data indicate that P.urinaria may be an alternative therapeutic agent for the treatment of HBV-related hepatitis.

Inactivated Sendai Virus Induces ROS-dependent Apoptosis and Autophagy in Human Prostate Cancer Cells

Objective The current study aims to investigate the effect of Hemagglutinating virus of Japan envelope（HVJ-E） on induction of apoptosis and autophagy in human prostate cancer PC3 cells, and the underlying mechanisms. Methods PC3 cells were treated with HVJ-E at various multiplicity of infection（MOI）, and the generated reactive oxygen species（ROS）, cell viability, apoptosis, and autophagy were detected, respectively. Next, the role of ROS played in the regulation of HVJ-E-induced apoptosis and autuphagy in PC3 cells were analysed. In the end, the relationship between HVJ-E-induced apoptosis and autuophagy was investigated by using rapamycin and chloroquine. Results Flow cytometry assay revealed that HVJ-E treatment induced dose-dependent apoptosis and that the JNK and p38 MAPK signaling pathways were involved in HVJ-E-induced apoptosis in PC3 cells. In addition, HVJ-E was able to induce autophagy in PC3 cells via the class III PI3 K/beclin-1 pathway. The data also implyed that HVJ-E-triggered autophagy and apoptosis were ROS dependent. When ROS was blocked with N-acetylcysteine（NAC）, HVJ-E-induced LC3-II conversion and apoptosis were reversed. Interestingly, HVJ-E-induced apoptosis was significantly increased by an inducer of autophagy, rapamycin pretreatment, both in vitro and in vivo. Conclusion HVJ-E exerts anticancer effects via autophagic cell death in prostate cancer cells.

Impact of alcohol on hepatitis C virus replication and interferon signaling

Hepatitis C virus (HCV) is one of the main etiological factors responsible for liver disease worldwide. It has been estimated that there are over 170 million people infected with HCV worldwide. Of these infected individuals, approximately 75% will go on to develop a life long necroinflammatory liver disease, which over decades, can result in serious complications, such as cirrhosis and hepatocellular carcinoma. Currently there is no effective vaccine and whilst antiviral therapies have been improved, they are still only effective in approximately 50% of individuals. HCV infection stands as a major cause of global morbidity and suffering, and places a signifi cant burden on health systems. The second highest cause of liver disease in the western world is alcoholic liver disease. Frequently, HCV infected individuals consume alcohol, and the combined effect of HCV and alcohol consumption is deleterious for both liver disease and response to treatment. This review discusses the impact of alcohol metabolism on HCV replication and the negative impact on interferon (IFN)-α treatment, with a particular focus on how alcohol and HCV act synergistically to increase oxidative stress, ultimately leading to exacerbated liver disease and a reduction in the effi cacy of IFN-α treatment. A better understanding of the complicated mechanisms at play in hepatocytes infected with HCV and metabo- lizing alcohol will hopefully provide better treatment options for chronic hepatitis C individuals that consume alcohol.

Activation of AMPK/MnSOD signaling mediates anti-apoptotic effect of hepatitis B virus in hepatoma cells

AIM: To investigate the anti-apoptotic capability of the hepatitis B virus(HBV) in the HepG2 hepatoma cell line and the underlying mechanisms.METHODS: Cell viability and apoptosis were measured by MTT assay and flow cytometry, respectively. Targeted knockdown of manganese superoxide dismutase(Mn SOD), AMP-activated protein kinase(AMPK) and hepatitis B virus X protein(HBx) genes as well as AMPK agonist AICAR and antagonist compound C were employed to determine the correlations of expression of these genes.RESULTS: HBV markedly protected the hepatoma cells from growth suppression and cell death in the condition of serum deprivation. A decrease of superoxide anion production accompanied with an increase of Mn SOD expression and activity was found in Hep G2.215 cells. Moreover, AMPK activation contributed to the up-regulation of Mn SOD. HBx protein was identified to induce the expression of AMPK and Mn SOD. CONCLUSION: Our results suggest that HBV suppresses mitochondrial superoxide level and exerts an antiapoptotic effect by activating AMPK/Mn SOD signaling pathway, which may provide a novel pharmacological strategy to prevent HCC.

S-adenosyl-L-methionine modifies antioxidant-enzymes,glutathione-biosynthesis and methionine adenosyltransferases-1/2 in hepatitis C virus-expressing cells

AIM: To elucidate the mechanism(s) by which S-adenosyl-L-methionine(SAM) decreases hepatitis C virus(HCV) expression.METHODS: We examined the effects of SAM on viral expression using an HCV subgenomic replicon cell culture system. Huh7 HCV-replicon cells were treated with 1 mmol/L SAM for different times(24-72 h), then total RNA and proteins were isolated. c DNA was synthesized and real time-PCR was achieved to quantify HCV-RNA, superoxide dismutase 1 and 2(SOD-1, SOD-2) catalase, thioredoxin 1, methionine adenosyltransferase 1A and 2A(MAT1A, MAT2A) expression, and GAPDH and RPS18 as endogenous genes. Expression of cellular and viral protein was evaluated by western-blot analysis using antibodies vs HCV-NS5 A, SOD-1, SOD-2, catalase, thioredoxin-1, MAT1 A, MAT2 A, GAPDH and actin. Total glutathione levels were measured at different times by Ellman's recycling method(0-24 h). Reactive oxidative species(ROS) levels were quantified by the dichlorofluorescein assay(0-48 h); Pyrrolidin dithiocarbamate(PDTC) was tested as an antioxidant control and H2O2 as a positive oxidant agent.RESULTS: SAM exposition decreased HCV-RNA levels 50%-70% compared to non-treated controls(24-72 h). SAM induced a synergic antiviral effect with standard IFN treatment but it was independent of IFN signaling. In addition, 1 mmol/L SAM exposition did not modify viral RNA stability, but it needs cellular translation machinery in order to decrease HCV expression. Total glutathione levels increased upon SAM treatment in HCV-replicon cells. Transcriptional antioxidant enzyme expression(SOD-1, SOD-2 and thioredoxin-1) was increased at different times but interestingly, there was no significant change in ROS levels upon SAM treatment, contrary to what was detected with PDTC treatment, where an average 40% reduction was observed in exposed cells. There was a turnover from MAT1A/MAT2 A, since MAT1 A expression was increased(2.5 fold-times at 48 h) and MAT2 A was diminished(from 24 h) upon SAM treatment at both the transcriptional and translational level. CONCLUSION: A likely mechanism(s) by which SAM diminish HCV expression could involve modulating antioxidant enzymes, restoring biosynthesis of glutathione and switching MAT1/MAT2 turnover in HCV expressing cells.

Oxidative stress modulation in hepatitis C virus infected cells

Hepatitis C virus(HCV) replication is associated with the endoplasmic reticulum, where the virus can induce cellular stress. Oxidative cell damage plays an important role in HCV physiopathology. Oxidative stress is triggered when the concentration of oxygen species in the extracellular or intracellular environment exceeds antioxidant defenses. Cells are protected and modulate oxidative stress through the interplay of intracellular antioxidant agents, mainly glutathione system(GSH) and thioredoxin; and antioxidant enzyme systems such as superoxide dismutase, catalase, GSH peroxidase, and heme oxygenase-1. Also, the use of natural and synthetic antioxidants(vitamin C and E, N-acetylcysteine, glycyrrhizin, polyenylphosphatidyl choline, mitoquinone, quercetin, S-adenosylmethionine and silymarin) has already shown promising results as co-adjuvants in HCV therapy. Despite all the available information, it is not known how different agents with antiviral activity can interfere with the modulation of the cell redox state induced by HCV and decrease viral replication. This review describes an evidence-based consensus on molecular mechanisms involved in HCV replication and their relationship with cell damage induced by oxidative stress generated by the virus itself and cell antiviral machinery. It also describes some molecules that modify the levels of oxidative stress in HCV-infected cells.

Investigation of the mechanism of adult-stage resistance to barley yellow dwarf virus associated with awheat-Thinopyrum intermedium translocation

Barley yellow dwarf virus(BYDV)can infect wheat(Triticum aestivum L.),leading to yield loss.Among four BYDV strains(GAV,GPV,PAV,and RMV)identified in China,BYDV-GAV is the prevailing isolate.YW642,a wheat–Thinopyrum intermedium translocation line,is resistant to BYDV isolates at both seedling and adult stages.Zhong 8601 is the wheat recurrent parent of YW642 and is susceptible to BYDV.In this study,we investigated the adult-stage resistance mechanism of YW642,measured BYDV titer and hydrogen peroxide(H_2O_2) in adult-stage leaves of YW642 and Zhong 8601 inoculated with BYDV-GAV,and identified transcriptional differences between YW642 and Zhong 8601 using microarray-based comparative transcriptomics.Enzyme-linked immunosorbent assay and H_2O_2assay showed that both BYDV titer and H_2O_2 content were markedly lower in YW642 than in Zhong 8601 at 21,28,35,and 40 days post-inoculation(dpi).The transcriptomic comparison revealed that many types of genes were significantly up-regulated at 35 dpi in adult-stage leaves of YW642 compared to Zhong 8601.The important up-regulated genes associated with the adult-stage resistance encoded 15 resistance-like proteins,pathogenesis-related proteins(such as defensin and lipid transfer proteins),protein kinase homologs,transcription factors,reactive oxygen species scavenging-related proteins,and jasmonic acid and gibberellic acid biosynthesis enzymes.These results suggest that precise expression regulation of these proteins plays a crucial role in adult-stage resistance of YW642 against BYDV infection.

转录因子NbMYB1R1通过促进活性氧积累抑制病毒侵染

[背景]黄瓜绿斑驳花叶病毒(cucumber green mottle mosaic virus,CGMMV)是我国重要的检疫性植物病毒,严重降低了世界范围内蔬菜以及瓜类的产量。MYB蛋白家族庞大,功能多样,存在于所有真核生物中。大多数MYB作为转录因子控制植物的发育和代谢,并对植物应对生物和非生物胁迫反应起重要的调控作用。前期研究显示CGMMV侵染后可以显著上调寄主转录因子基因NbMYB1R1的表达。[目的]明确NbMYB1R1参与CGMMV侵染的机制,为CGMMV病害防控提供理论依据。[方法]运用MEGA 7.0构建系统进化树,对NbMYB1R1的氨基酸序列进行系统进化分析;通过构建NbMYB1R1的荧光表达载体,转化GV3101农杆菌后浸润本氏烟叶片,激光共聚焦显微镜观察其亚细胞定位;利用qRT-PCR技术分析NbMYB1R1在CGMMV侵染不同时期的转录水平及NbMYB1R1沉默烟草植株中活性氧(reactive oxygen species,ROS)相关基因的转录变化;通过沉默NbMYB1R1及下游调控基因NbAOX1a和NbAOX1b,分析NbMYB1R1及下游调控基因NbAOX1a和NbAOX1b在CGMMV侵染过程中的作用;瞬时过表达NbMYB1R1和NbMYB1R1关键氨基酸突变体,分析NbMYB1R1对CGMMV侵染的影响;使用台盼蓝染色以及DAB染色观察瞬时过表达NbMYB1R1造成的细胞死亡是否与程序性细胞死亡(programmed cell death,PCD)以及ROS的积累有关;运用酵母双杂交技术验证NbMYB1R1是否与CGMMV相关蛋白互作。[结果]系统进化树分析表明,NbMYB1R1属于1R MYB大类并与多种烟草的MYB转录因子同源性极高;亚细胞定位结果显示NbMYB1R1定位于细胞核;在CGMMV侵染的烟草植株中,NbMYB1R1的转录水平对比健康植株有明显变化,在CGMMV侵染8、12 d时NbMYB1R1的转录水平显著上调;在沉默内源基因NbMYB1R1的植株上接种CGMMV,3 d后NbMYB1R1沉默植株系统叶出现斑驳、卷曲症状,而对照植株在3.5 d时才出现症状;同时CGMMV CP mRNA水平和蛋白水平检测结果也表明沉默NbMYB1R1可以有效促进CGMMV的积累;在本氏烟叶片瞬时过表达NbMYB1R1及其突变体3 d时检测CGMMV蛋白水平表达量,结果显示过表达NbMYB1R1可以有效抑制CGMMV的侵染,DNA结合结构域缺失后会减轻NbMYB1R1对CGMMV的抑制;台盼蓝和DAB染色结果表明,在瞬时过表达NbMYB1R1蛋白后导致ROS积累并引起细胞死亡;在沉默内源基因NbMYB1R1的植株叶片上检测ROS相关基因的转录水平,发现交替氧化酶基因AOX1a、AOX1b转录水平显著上调;在沉默内源基因NbAOX1a和NbAOX1b的植株上接种CGMMV,4 d后NbAOX1a和NbAOX1b沉默植株系统叶出现斑驳、卷曲症状,而对照植株在3.5 d时就出现症状;同时CGMMV CP mRNA水平和蛋白水平检测结果也表明沉默NbAOX1a和NbAOX1b可以有效抑制CGMMV的积累;酵母双杂交结果显示NbMYB1R1不与CGMMV编码蛋白直接相互作用。[结论]随着CGMMV侵染,防御相关基因NbMYB1R1表达上调,从而抑制下游基因AOX1a、AOX1b的表达并激活细胞内产生ROS抑制病毒侵染,但NbMYB1R1不是通过与CGMMV病毒蛋白直接互作而产生此作用。由此可见,NbMYB1R1在CGMMV侵染过程中发挥了重要作用。

慢性乙型肝炎病毒感染患者中性粒细胞活性氧的生成特点及临床意义

目的分析慢性乙型肝炎病毒(HBV)感染患者中性粒细胞活性氧(ROS)的生成特点并探讨其临床意义。方法选取2022年4-9月解放军总医院第五医学中心收治的88例慢性HBV感染者,其中58例未接受抗病毒治疗者纳入未治疗组,30例已进行核苷(酸)类似物(NAs)治疗者纳入治疗组,同时选取20名健康人作为对照组,比较各组临床指标及病毒学指标的差异。采用二氢罗丹明123(DHR 123)荧光染料与中性粒细胞进行孵育,通过流式细胞仪检测各组中性粒细胞自发ROS生成水平,并分析其与HBV DNA载量、乙肝表面抗原(HBsAg)定量、白细胞计数(WBC)、C反应蛋白(CRP)、谷丙转氨酶(ALT)及谷草转氨酶(AST)的相关性。分别使用完全培养基、HepG2细胞上清液及HepG2.2.15细胞上清液与健康人中性粒细胞共孵育,体外验证HBV病毒颗粒及蛋白对中性粒细胞自发ROS生成水平的影响。比较脂多糖(LPS)刺激后各组中性粒细胞继发ROS的生成水平。结果与对照组比较,未治疗组患者中性粒细胞自发ROS生成水平增高(P<0.05),且与HBV DNA载量、HBsAg定量呈正相关(r=0.315,P=0.016;r=0.326,P=0.013),但与WBC、CRP、ALT及AST无明显相关性(P>0.05)。HepG2.2.15上清液可诱导中性粒细胞自发ROS生成水平增高(P<0.05)。LPS刺激后,未治疗组及治疗组患者中性粒细胞继发ROS生成水平均较对照组降低(P<0.05)。结论HBV抗病毒治疗在降低病毒载量的同时可降低患者中性粒细胞自发ROS生成水平,但不能有效恢复LPS刺激后中性粒细胞继发ROS的生成能力。

In vitro and in vivo evaluation of cerium oxide nanoparticles in respiratory syncytial virus infection

Respiratory syncytial virus(RSV)is the most common cause of viral bronchiolitis among children worldwide,yet there is no vaccine for RSV disease.This study investigates the potential of cube and sphere-shaped cerium oxide nanoparticles(CNP)to modulate reactive oxygen(ROS)and nitrogen(RNS)species and immune cell phenotypes in the presence of RSV infection in vitro and in vivo.Cube and sphere-shaped CNP were synthesized by hydrothermal and ultrasonication methods,respectively.Physico-chemical characterization confirmed the shape of sphere and cube CNP and effect of various parameters on their particle size distribution and zeta potential.In vitro results revealed that sphere and cube CNP differentially modulated ROS and RNS levels in J774 macrophages.Specifically,cube CNP significantly reduced RSV-induced ROS levels without affecting RNS levels while sphere CNP increased RSV-induced RNS levels with minimal effect on ROS levels.Cube CNP drove an M1 phenotype in RSV-infected macrophages in vitro by increasing macrophage surface expression of CD80 and CD86 with a concomitant increase in TNFαand IL-12p70,while simultaneously decreasing M2 CD206 expression.Intranasal administration of sphere and cube-CNP were well-tolerated with no observed toxicity in BALB/c mice.Notably,cube CNP preferentially accumulated in murine alveolar macrophages and induced their activation,avoiding enhanced uptake and activation of other inflammatory cells such as neutrophils,which are associated with RSV-mediated inflammation.In conclusion,we report that sphere and cube CNP modulate macrophage polarization and innate cellular responses during RSV infection.

烟粉虱取食及其与TYLCCNV共侵染对烟草植株内H_(2)O_(2)的诱导响应

本研究采用DAB染色法定位检测经烟粉虱取食及其与中国番茄黄化曲叶病毒(TYLCCNV)共侵染后烟草叶片中H_(2)O_(2)的积累,并定量分析烟草叶片中H_(2)O_(2)的含量。DAB染色结果表明,2种处理的烟草叶片中均未检测到H_(2)O_(2)积累;H_(2)O_(2)的定量分析结果表明,2种处理均可诱导烟草叶片中H_(2)O_(2)含量较对照显著升高。在烟粉虱取食处理后0.5、1、3、6、12 h及1、3、5、7、9 d,烟草叶片中H_(2)O_(2)含量分别为对照的1.62、1.71、1.77、1.77、1.95、1.46、1.82、1.63、1.53和1.08倍;共侵染处理后烟草叶片中H_(2)O_(2)含量分别为对照的1.64、1.84、1.95、2.19、2.29、1.43、2.17、2.08、2.60和1.79倍。烟粉虱与TYLCCNV共侵染较烟粉虱取食可诱导烟草植株内H_(2)O_(2)含量显著升高。

Hepatitis B virus X protein induces ALDH2 ubiquitin-dependent degradation to enhance alcoholic steatohepatitis

Background:Excessive alcohol intake with hepatitis B virus(HBV)infection accelerates chronic liver disease progression and patients with HBV infection are more susceptible to alcohol-induced liver disease.Hepatitis B virus X protein(HBx)plays a crucial role in disease pathogenesis,while its specific role in alcoholic liver disease(ALD)progression has not yet been elucidated.Here,we studied the role of HBx on the development of ALD.Methods:HBx-transgenic(HBx-Tg)mice and their wild-type littermates were exposed to chronic plus binge alcohol feeding.Primary hepatocytes,cell lines,and human samples were used to investigate the interaction between HBx and acetaldehyde dehydrogenase 2(ALDH2).Lipid profiles in mouse livers and cells were assessed by using liquid chromatography–mass spectrometry.Results:We identified that HBx significantly aggravated alcohol-induced steatohepatitis,oxidative stress,and lipid peroxidation in mice.In addition,HBx induced worse lipid profiles with high lysophospholipids generation in alcoholic steatohepatitis,as shown by using lipidomic analysis.Importantly,serum and liver acetaldehyde were markedly higher in alcoholfed HBx-Tg mice.Acetaldehyde induced lysophospholipids generation through oxidative stress in hepatocytes.Mechanistically,HBx directly bound to mitochondrial ALDH2 to induce its ubiquitin–proteasome degradation,resulting in acetaldehyde accumulation.More importantly,we also identified that patients with HBV infection reduced ALDH2 protein levels in the liver.Conclusions:Our study demonstrated that HBx-induced ubiquitin-dependent degradation of mitochondrial ALDH2 aggravates alcoholic steatohepatitis.

鸡冠状病毒感染Vero细胞后NLRP3炎症小体信号通路的活化

目的探讨鸡冠状病毒[即传染性支气管炎病毒(IBV)]感染Vero细胞后,NLRP3炎症小体的变化。方法通过荧光染色、酶联免疫吸附实验、聚合酶链式反应(PCR)方法检测IBV Beaudette株感染非洲绿猴肾细胞Vero细胞不同时间点NLRP3炎症小体相关的信号分子。结果IBV感染Vero细胞后,病毒开始增殖并有明显的细胞病变效应(CPE)现象;通过荧光染色和流式细胞术检测胞内活性氧水平,发现IBV可使Vero细胞内活性氧含量随感染时间延长而增加;酶联免疫吸附实验检测细胞上清液中IL-1β与IL-18的分泌量,发现IBV能促进细胞IL-1β、IL-18的分泌;qRT-PCR检测NLRP3炎症小体相关基因(NLRP3、Caspase-1、IL-1β和IL-18)的转录水平,发现IBV感染Vero细胞后,所有检测基因mRNA表达量均在不同时间点高于对照组。结论IBV感染Vero细胞后可活化NLRP3炎症小体信号通路,并进一步促进IL-1β、IL-18的分泌。

RIP3/CaMKⅡ信号通路对重症病毒性心肌炎小鼠心脏功能及生存曲线的影响

目的:观察RIP3/CaMKⅡ信号通路对柯萨奇病毒B3(CVB3)诱导的重症病毒性心肌炎小鼠心脏功能及生存曲线的影响,并探讨CaMKⅡ特异性抑制剂KN-93对心肌损伤的作用。方法:将60只雄性BALB/c小鼠分为正常对照组、CVB3组、CVB3+KN-93组和CVB3+KN-93+Ti(活性氧簇特异性抑制剂)组,以上小鼠腹腔及尾静脉注射药物连续7 d。采用双抗体夹心免疫法检测小鼠心肌细胞坏死标志物,心脏超声技术检测小鼠心脏结构和功能,绘制Kaplan-Meier生存曲线,观察KN-93对小鼠心肌的保护作用。结果:CVB3+KN-93组小鼠心肌细胞坏死较CVB3组显著减轻,心脏功能和生存曲线改善(P<0.01);CVB3+KN-93组小鼠心肌组织H2O2含量较CVB3组显著下降(P<0.01);加入Ti后,CVB3+KN-93+Ti组H2O2含量较CVB3+KN-93组轻度下降(P<0.05),但无论是心脏功能还是生存曲线均无显著差异。结论:RIP3/CaMKⅡ信号通路在CVB3诱导的急性重症病毒性心肌炎小鼠心肌细胞损伤中起到主导作用;KN-93能阻断这种作用并可能产生新的治疗方式,其效果可能是通过对CaMKⅡ的直接抑制和对活性氧簇的间接抑制而实现的。

氧化应激与艾滋病

日益累积的证据表明,H IV感染和艾滋病进展与氧化应激密切相关,H IV/AIDS病人处于高度氧化应激状态,抗氧化剂补充有望成为艾滋病预防和治疗的重要措施之一。通过H IV感染者的氧化应激,氧化应激在艾滋病致病机理中的作用和H IV/AIDS病人的抗氧化剂补充3个方面论述了氧化应激与艾滋病的关系。

乙肝病毒S蛋白对人精子氧化应激的影响

目的：乙肝病毒（HBV）可通过血睾屏障将病毒DNA整合到男性生殖细胞的基因组中,诱发多种染色体畸变,而且还能破坏精子线粒体功能,导致精子活力受损,受精率和受精指数下降,但其机制未明。本研究主要考察乙肝病毒S蛋白（HBs）对人精子氧化应激的影响。方法：人精子与不同浓度（0、25、50、100μg/m L）HBs共孵育3 h后,用流式细胞术检测精子内活性氧（ROS）的变化情况,应用酶标仪检测精子细胞内丙二醛（MDA）含量和总抗氧化能力（TAC）。结果：人精子与25~100μg/m L HBs共孵育3 h后,ROS阳性精子比率、人精子细胞膜MDA水平显著高于对照组,而TAC水平显著低于对照组（P〈0.05或0.01）。MDA水平以及ROS阳性精子比率的升高与HBs呈剂量依赖关系（r=0.62,r=0.75;P均〈0.01）。TAC水平的降低与HBs呈剂量依赖关系（r=-0.89;P〈0.01）。结论：HBs可诱导人精子内ROS升高、TAC降低,从而产生氧化应激,使细胞膜脂质过氧化进而影响人精子的功能。