鸡血藤总黄酮对猪圆环病毒2型感染小鼠脾脏氧化应激的影响

试验旨在探讨鸡血藤总黄酮（TFSD）对猪圆环病毒2型（PCV2）感染小鼠脾脏氧化应激的影响。将70只昆明小鼠随机分为7组,对照组、TFSD组（100 mg/kg体重）、PCV2组、PCV2＋维生素C （VC）组、PCV2＋不同浓度TFSD （25、50和100 mg/kg体重）组,每组10只,连续3 d采用灌胃和腹腔注射PCV2病毒液的方法建立小鼠氧化应激模型,第4~6天每天按上述分别灌胃给予生理盐水、VC或TFSD。第7天剖杀小鼠,取脾脏分析黄嘌呤氧化酶（XOD）、髓过氧化物酶（MPO）和超氧化物歧化酶（SOD）活力,并检测还原型谷胱甘肽（GSH）和氧化型谷胱甘肽（GSSG）水平,计算GSH/GSSH。结果显示,PCV2感染小鼠后,脾脏中XOD与MPO的活力及GSSG水平显著上升（P＜0.05）,SOD活力、GSH水平及GSH/GSSG显著下降（P＜0.05）。TFSD处理小鼠脾脏的SOD活力、GSH水平和GSH/GSSG比值均显著高于PCV2组（P＜0.05）,而XOD、MPO活力和GSSG水平则显著低于PCV2组（P＜0.05）,对PCV2引起的氧化应激相关酶活力与相关分子水平变化的抑制作用优于VC。结果表明,鸡血藤总黄酮对PCV2诱导的小鼠脾脏氧化应激有良好的调节作用。     

鸡血藤总黄酮对大肠杆菌败血症的治疗作用

为研究鸡血藤总黄酮对小鼠大肠杆菌败血症的治疗作用,本研究采用腹腔注射大肠杆菌菌液的方法建立小鼠大肠杆菌败血症模型,1 h后灌胃给予不同剂量鸡血藤总黄酮,12 h后观察小鼠临床表现并进行评分,然后剖杀小鼠,比较肝脏、胸腺和脾脏的外观、脏器指数和肝脏病理切片结果,检测各项血液常规指标,分析血清中谷草转氨酶(AST)、谷丙转氨酶(ALT)及乳酸脱氢酶(LDH)水平,测定血清中白细胞介素1β(IL-1β)、IL-2、IL-6、IL-10及肿瘤坏死因子α(TNF-α)等炎性细胞因子的水平。结果发现,腹腔注射大肠杆菌后引起小鼠被毛粗乱、精神沉郁、眼睛红肿、眼部分泌物增加且呼吸急促等症状,部分严重的小鼠出现抽搐、休克现象,肝脏、胸腺、脾脏出现萎缩、变小的现象,且脏器指数下降。肝脏病理切片可观察到肝索紊乱、肝窦增大、细胞边缘不清晰、部分区域炎性细胞浸润。血液中白细胞(WBC)、淋巴细胞(LYM)、血红蛋白(HGB)、红细胞(RBC)和血小板(PLT)等指标水平发生显著改变(P<0.05),且血清中ALT、AST、LDH及炎性细胞因子等水平显著升高(P<0.05)。而鸡血藤总黄酮和阿米卡星处理可改善上述临床表现,抑制相关指标的改变,使其趋于与对照组相似水平。以上结果表明,鸡血藤总黄酮可通过提高机体免疫水平及抑制炎性反应减轻大肠杆菌败血症引起的机体损伤。     

辣蓼黄酮对脂多糖诱导下RAW264.7细胞活性氧及炎性因子分泌的影响

采用酶解-超声偶联法提取辣蓼全草中的总黄酮,并经过萃取分离结合大孔树脂纯化富集获得辣蓼乙酸乙酯部分黄酮(FEA)与辣蓼正丁醇部分黄酮(FNB),并通过MTT法测定药物安全浓度。采用不同剂量的FEA与FNB作用于脂多糖(LPS)刺激所诱导的RAW264.7细胞炎症体外模型,测定细胞内活性氧(ROS)、一氧化氮(NO)及炎症因子TNF-α、IL-1β、IL-6、IL-8、IL-10水平。结果表明,FEA与FNB能明显减少LPS诱导的ROS释放量;不同浓度的FEA与FNB可以抑制LPS诱导的RAW264.7细胞分泌NO水平的升高。FEA与FNB均能减少LPS刺激所诱导的促炎因子TNF-α、IL-1β、IL-6、IL-8释放,FEA能促进抗炎因子IL-10的生成。说明一定浓度的FEA和FNB具有显著的抗炎效果,且其抗炎作用的产生可能与抗氧化途径有关。     

鸡血藤黄酮乙酸乙酯部位对H_2O_2诱导RAW264.7细胞氧化应激的调节作用

为了探讨鸡血藤黄酮乙酸乙酯部位(the ethyl acetate fraction of spatholobus suberectdunn flavonoid,EFSF)的抗氧化应激作用,试验采用双氧水(H_2O_2)处理RAW264.7细胞,建立氧化应激模型,EFSF处理氧化应激细胞后,利用分光光度法检测一氧化氮(NO)分泌水平和细胞内活性氧自由基(ROS)水平的变化。结果表明:200μg/m L及以下浓度的EFSF对RAW264.7细胞无毒性作用;25,50,100μg/m L的EFSF均能明显降低H_2O_2诱导氧化应激细胞的NO分泌水平及细胞内ROS的含量。说明EFSF对H_2O_2处理RAW264.7细胞引起的氧化应激损伤具有一定的保护作用。     

正交试验优选鸡血藤总黄酮提取工艺

为对鸡血藤总黄酮的提取工艺方法进行优化选择,以单因素考察为试验基础,选用四因素三水平的正交试验方案,对乙醇浓度、提取时间、提取温度、料液比四个因素对鸡血藤总黄酮提取率的影响进行了研究。结果表明:试验条件范围内鸡血藤总黄酮的最佳提取工艺为:乙醇提取液浓度60%、提取温度70℃、提取时间2.5 h、提取料液比1:20,此条件下鸡血藤总黄酮的提取率为34.90%。同时试验分析结果表明,四个影响因素中提取时间对提取率影响最大。综上,此提取工艺方法绿色、简单,可为鸡血藤在饲料添加剂中的应用提供参考。     

鸡血藤总黄酮对环磷酰胺所致小鼠肝损伤的保护作用

目的 研究鸡血藤总黄酮对环磷酰胺所致小鼠肝损伤的保护作用。方法 将60只昆明系小鼠随机分为6组,每组10只,试验周期为7 d。对照组小鼠试验全程给予生理盐水。环磷酰胺（CTX）组小鼠1~4 d灌胃给予生理盐水;5~7 d腹腔注射80 mg/（kg·BW）CTX,每天给药1次。鸡血藤总黄酮（TFSD）100组、CTX+TFSD25组、CTX+TFSD50组、CTX+TFSD100组小鼠1~7 d分别灌胃给予100、25、50、100 mg/（kg·BW）TFSD,每天给药1次;试验5~7 d除TFSD100组小鼠腹腔注射生理盐水外,其余试验组腹腔注射80 mg/（kg·BW）CTX,每天给药1次。试验结束后对各组小鼠进行眼球采血及剖检;检测肝组织中超氧化物歧化酶（SOD）、髓过氧化物酶（MPO）、黄嘌呤氧化酶（XOD）以及谷胱甘肽过氧化物酶（GSH-Px）活力,测定血清中谷草转氨酶（AST）、谷丙转氨酶（ALT）和碱性磷酸酶（ALP）活力,检测血清中肿瘤坏死因子-α（TNF-α）、白介素-6（IL-6）和白介素-1β（IL-1β）含量。结果 环磷酰胺可显著（P<0.05）降低小鼠肝脏中SOD、GSH-Px以及血清中ALT活力,显著（P<0.05）升高肝脏中XOD与MPO活力、血清中ALP活力以及炎症细胞因子IL-1β和IL-6释放水平。25、50、100 mg/（kg·BW）的TFSD处理均可显著（P<0.05）抑制由CTX引起的SOD活力降低,25 mg/（kg·BW）的TFSD处理可显著（P<0.05）提高CTX处理小鼠的GSH-Px活力;50 mg/（kg·BW）的TFSD处理可显著（P<0.05）抑制由CTX引起的XOD和MPO活力升高,且可显著（P<0.05）降低CTX处理小鼠血清中的AST活力;25、50、100 mg/（kg·BW）的TFSD处理可显著（P<0.05）抑制由CTX引起的血清中ALP活力升高,同时降低ALT活力。25、50、100 mg/（kg·BW）的TFSD处理均可显著（P<0.05）抑制由CTX引起的IL-1β水平升高。结论 鸡血藤总黄酮可通过调节小鼠炎症因子释放以及肝组织中氧化还原酶水平对环磷酰胺所致肝损伤提供保护,推荐剂量为25 mg/（kg·BW）。     

水栀子多糖提取物对感染猪圆环病毒2型的RAW264.7细胞氧化应激的调节作用研究

为了研究水栀子多糖提取物(Gardenia jasminoide var.radicans polysaccharide extract, GJPE)对猪圆环病毒2型(PCV-2)感染的RAW264.7细胞氧化应激的调节作用,试验采用ELISA法测定了GJPE对RAW264.7细胞的安全浓度;将RAW264.7细胞分为空白对照组、维生素C组、不同浓度(安全浓度范围)GJPE组,分别加入到96孔细胞培养板(1×10⁶ 个/mL)和24孔细胞培养板(2×10⁵ 个/mL)中,除空白对照组加基础培养基外,其他各组分别加入1×10³ TCID₅₀ PCV-2病毒液,并设置PCV-2组(只添加PCV-2病毒液)作为病毒对照组,100μL/孔(96孔细胞培养板)或500μL/孔(24孔细胞培养板),培养24 h;吸弃上清液,用PBS洗涤3遍,空白对照组和PCV-2组分别加入基础培养基,维生素C组加入200μmol/mL维生素C,不同浓度GJPE组分别加入相应浓度GJPE[100μL/孔(96孔细胞培养板)或50...     

谷氨酰胺对脂多糖诱导的断奶仔猪氧化应激的影响

本试验以大肠杆菌型脂多糖(LPS)建立氧化应激模型,探讨了谷氨酰胺(GLN)对断奶仔猪氧化应激的影响。选用24头28日龄的健康三元(杜×长×大)断奶仔猪,随机分成3组,每组8个重复,每个重复1头猪。对照组和应激组饲喂基础饲粮,GLN组饲粮在基础饲粮中添加1%GLN,试验期为30 d。在试验第22、25、28和30天,应激组和GLN组仔猪分别按每千克体重腹腔注射100μg LPS,对照组仔猪腹腔注射相同剂量的灭菌生理盐水,第30天进行前腔静脉采血并屠宰采取所需肠道样品,检测氧化应激相关指标。结果显示:1)LPS攻毒前各组血清抗氧化能力指标均无显著差异(P0.05)。LPS攻毒后,应激组血清丙二醛(MDA)含量显著高于对照组(P0.05);GLN组血清MDA含量和超氧化物歧化酶(SOD)活性显著低于应激组和对照组(P0.05)。2)LPS攻毒后,在十二指肠黏膜中,GLN组过氧化氢酶(CAT)和锌铜超氧化物歧化酶(Cu Zu SOD)基因相对表达量显著高于应激组(P0.05)。在空肠黏膜中,GLN组CAT、锰超氧化物歧化酶(Mn SOD)、谷胱甘肽过氧化物酶1(GPX1)和谷胱甘肽过氧化物酶4(GPX4)基因相对表达量显著高于对照组和应激组(P0.05),对照组GPX4基因相对表达量显著高于应激组(P0.05)。在回肠黏膜中,GLN组和应激组CAT基因相对表达量显著低于对照组(P0.05),GPX4基因相对表达量显著高于对照组(P0.05);GLN组Mn SOD基因相对表达量显著高于对照组和应激组(P0.05),Cu Zn SOD基因相对表达量显著高于对照组(P0.05)。结果表明,GLN在一定程度上可以缓解断奶仔猪因LPS引起的氧化应激,以期为实际生产中减少氧化应激提供一定的理论基础。     

活性氧参与脂多糖诱导小鼠巨噬细胞RAW 264.7活化诱导凋亡

研究活性氧(ROS)与脂多糖(LPS)诱导后巨噬细胞活化诱导凋亡的关系。体外培养小鼠巨噬细胞RAW264.7,采用不同浓度LPS诱导细胞,添加100μmol/L H2O2增加细胞内ROS,或用姜黄素(7.5μmol/L)降低细胞内ROS;流式细胞术分析细胞凋亡、线粒体膜电位(MMP)和ROS。结果显示,RAW264.7细胞凋亡和细胞内ROS水平均随LPS浓度增加而增加,高浓度的LPS导致MMP下降;与LPS(8μg/m L)单独作用组比,H2O2增加ROS,并导致凋亡细胞百分率增加;姜黄素降低LPS诱导的细胞凋亡,同时减少细胞内ROS。表明活性氧参与LPS诱导小鼠巨噬细胞RAW264.7活化诱导凋亡。     

替唑尼特对脂多糖诱导RAW264.7细胞氧化应激及炎症因子的影响

本研究通过脂多糖(LPS)诱导大鼠腹腔巨噬细胞(RAW264.7)建立氧化应激模型,探讨替唑尼特(TIZ)的抗氧化和抗炎效果。培养RAW264.7细胞,给予LPS(1μg/m L)刺激并用TIZ(25、50、75、100μmol/L)作用。MTT法检测细胞存活率;DCFH-DA荧光探针法检测TIZ对ROS生成的影响;同时,检测MDA的生成量以评估TIZ对脂质氧化的影响;检测GSH的含量以及CAT、SOD、GSH-Px酶活性以评估TIZ对细胞抗氧化系统的影响;检测IL-6的分泌和COX-2的表达量以评估TIZ对炎症细胞因子的作用。结果显示LPS刺激后细胞表现出显著性的氧化应激效应,而给予TIZ处理后,ROS和MDA生成量呈剂量相关性的显著下降;细胞IL-6的分泌和COX-2的表达量也随TIZ处理而显著下降,而GSH含量以及CAT、SOD、GSH-Px活性在TIZ处理后却呈剂量相关性的显著性上升。因此,TIZ可抑制LPS诱导的RAW264.7细胞氧化应激和炎症反应,这为TIZ的作用机制研究以及应用奠定了基础。     

Effect of Total Flavonoids of Spatholobus suberectus Dunn on Oxidative Stress in Mice Spleen Induced by PCV2

The aim of this study was to investigate the effect of total flavonoids of Spatholobus suberectus Dunn (TFSD) on porcine circovirus type 2 (PCV2) induced oxidative stress in mice spleen.70 Kunming mice were divided into 7 groups:Control group, TFSD group (100 mg/(kg·BW)), PCV2 group, PCV2+vitamin C (VC) group, and PCV2+various concentrations of TFSD groups (25, 50 and 100 mg/(kg·BW)). Mice were continuously treated with PCV2 via both intragastric administration and intraperitoneal injection for 3 d to establish oxidative stress models. From the 4th to 6th day, mice were intragastric administrated with saline, VC or TFSD, respectively, according to the grouping method. At the 7th day, the activities of xanthine oxidase (XOD), myeloperoxidase (MPO) and superoxide dismutase (SOD), the levels of glutathione (GSH) and oxidized glutathione (GSSG), and the ratio of GSH to GSSG in the mice spleen were analyzed. The results showed that PCV2 infection significantly upregulated the XOD and MPO activities and GSSG content(P <0.05), and dramatically downregulated the SOD activity, GSH level and the ratio of GSH to GSSG (P <0.05) in the mice spleen.Compared to PCV2 group, the SOD activity, GSH content and the ratio of GSH to GSSG in mice treated with TFSD were significantly increased (P <0.05), while the activities of XOD and MPO and the level of GSSG were significantly decreased (P <0.05), showing better performance in the inhibition of PCV2 induced changes of oxidative stress associated enzyme activities and moledule levels than VC.In conclusion,TFSD had regulative effect on the oxidative stress induced by PCV2 in mouse spleen.     

钩吻素子对LPS诱导RAW264.7细胞炎症的作用

为揭示钩吻素子的体外抗炎作用机理,利用硝酸还原酶法和ELISA法研究了不同质量浓度(100,200,400mg/L)的钩吻素子对一氧化氮(NO)和细胞因子白介素-1β(IL-1β)、白介素-6(IL-6)和肿瘤坏死因子-α(TNF-α)分泌的影响,运用了QPCR的方法检测了钩吻素子对诱导型一氧化氮合成酶(iNOS)、IL-1β、IL-6和TNF-αmRNA表达水平的影响,并运用Western blot法检测了钩吻素子对iNOS蛋白表达的影响。结果显示,100,200,400 mg/L质量浓度的钩吻素子能够极显著的抑制RAW264.7细胞NO的产生以及IL-1β、IL-6和TNF-α的分泌(P<0.01),同时能够剂量依赖性的降低iNOS、IL-1β、IL-6和TNF-α的mRNA水平,抑制脂多糖(LPS)引起的iNOS蛋白表达的升高(P<0.01)。由此推论,钩吻素子可能通过抑制炎症因子的分泌,减少NO的释放,下调iNOS、IL-1β、IL-6和TNF-α的mRNA水平,抑制iNOS蛋白过度表达达到其抗炎作用。     

氨基多糖纳米微粒对 RAW264.7 细胞株TNF-α基因mRNA表达量的影响

本试验探讨氨基多糖纳米微粒(Chitosan nanoparticle,CNP)对巨噬细胞RAW264.7细胞株中肿瘤坏死因子-α(TNF-α)基因mRNA表达量的影响.以巨噬细胞株RAW264.7为腹腔巨噬细胞模型,分别作用RAW264.7细胞12、18 h及24 h后,运用RT-PCR半定量法分析TNF-α mRNA表达水平的变化.结果表明:CNP有明显的提高TNF-α mRNA表达量的作用.CNP组TNF-α mRNA表达量12、18 h及24 h分别为91%、63.2%和152.5%;对照组TNF-α mRNA表达量12、18 h及24 h分别为78.9%、51.2%及109.3%.CNP组与对照组相应时间段mRNA表达量相比均有所提高,提高了15.3%、23.4%及39.5%.其中CNP组24 h TNF-αmRNA表达量与其他各组相比差异显著(P＜0.05).CNP能提高巨噬细胞株RAW264.7 TNF-α mRNA的表达,因此发挥抗肿瘤作用.     

TLRs介导的信号通路在马链球菌感染RAW264.7细胞中的作用

为探究Toll样受体(TLRs)介导的信号通路在马链球菌马亚种(S.equi)感染小鼠巨噬细胞RAW264.7中的作用,收集S.equi感染后不同时间点的RAW264.7细胞,提取总RNA并反转录成cDNA,利用实时荧光定量PCR技术检测细胞Toll样受体1、2、6(TLR1、TLR2、TLR6)、接头蛋白骨髓分化蛋白88(MyD88)及细胞因子IL-1、IL-6、IL-10、IL-12、TNF-αmRNA的表达情况。结果显示,S.equi感染RAW264.7细胞后6h时,TLR1、TLR2、TLR6与MyD88mRNA水平均较对照组没有显著差异(P>0.05);感染后12h时,TLR1、TLR2和TLR6mRNA表达量未出现明显上升(P>0.05),而MyD88mRNA水平极显著升高(P<0.01);感染后24h时,TLR1、TLR2和TLR6mRNA表达水平出现极显著升高(P<0.01),MyD88mRNA表达没有显著变化(P>0.05),且IL-10和IL-12mRNA水平与对照组相比极显著升高(P<0.01),IL-1、IL-6和TNF-αmRNA水平均极显著下降(P<0.01)。结果表明,TLRs介导的信号通路参与S.equi感染RAW264.7细胞的免疫应答反应。     

Anti-inflammatory effect of sulforaphane on LPS-stimulated RAW 264.7 cells and ob/ob mice

BackgroundSulforaphane (SFN) is an isothiocyanate compound present in cruciferous vegetables. Although the anti-inflammatory effects of SFN have been reported, the precise mechanism related to the inflammatory genes is poorly understood.ObjectivesThis study examined the relationship between the anti-inflammatory effects of SFN and the differential gene expression pattern in SFN treated ob/ob mice.MethodsNitric oxide (NO) level was measured using a Griess assay. The inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) expression levels were analyzed by Western blot analysis. Pro-inflammatory cytokines (tumor necrosis factor [TNF]-α, interleukin [IL]-1β, and IL-6) were measured by enzyme-linked immunosorbent assay (ELISA). RNA sequencing analysis was performed to evaluate the differential gene expression in the liver of ob/ob mice.ResultsThe SFN treatment significantly attenuated the iNOS and COX-2 expression levels and inhibited NO, TNF-α, IL-1β, and IL-6 production in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. RNA sequencing analysis showed that the expression levels of 28 genes related to inflammation were up-regulated (> 2-fold), and six genes were down-regulated (< 0.6-fold) in the control ob/ob mice compared to normal mice. In contrast, the gene expression levels were restored to the normal level by SFN. The protein-protein interaction (PPI) network showed that chemokine ligand (Cxcl14, Ccl1, Ccl3, Ccl4, Ccl17) and chemokine receptor (Ccr3, Cxcr1, Ccr10) were located in close proximity and formed a “functional cluster” in the middle of the network.ConclusionsThe overall results suggest that SFN has a potent anti-inflammatory effect by normalizing the expression levels of the genes related to inflammation that were perturbed in ob/ob mice.     

Effect of grape polyphenols on oxidative stress in canine lens epithelial cells

OBJECTIVE: To evaluate whether the effects of oxidative stress could be attenuated in cultures of canine lens epithelial cells (LECs) by incubation with grape seed proanthocyanidin extract (GSE), resveratrol (RES), or a combination of both (GSE+RES). SAMPLE POPULATION: Primary cultures of canine LECs. PROCEDURES: LECs were exposed to 100MM tertiary butyl-hydroperoxide (TBHP) with or without GSE, RES, or GSE+RES. The dichlorofluorescein assay was used to detect production of reactive oxygen species (ROS), and immunoblot analysis was used to evaluate the expression of stress-induced cell-signaling markers (ie, the mitogen-activated protein kinase [MAPK] and phosphoinositide-3 kinase [PI3K] pathways). RESULTS: GSE and GSE+RES significantly reduced ROS production after a 30-minute exposure to TBHP. Only GSE significantly reduced ROS production after a 120-minute exposure to TBHP. Incubation with GSE reduced TBHP-induced activity of the MAPK and PI3K pathways. CONCLUSIONS AND CLINICAL RELEVANCE: GSE inhibited key components associated with cataractogenesis, ROS production, and stress-induced cell signaling. On the basis of the data reported here, there is strong evidence that GSE could potentially protect LECs from the damaging effects of oxidative stress.     

Moringa oleifera leaf flavonoids protect bovine mammary epithelial cells from hydrogen peroxide-induced oxidative stress in vitro

Bovine mammary epithelial cells (BMECs) of high-producing dairy cows are subject to constant oxidative stress as a result of high metabolic rate and physiological adaptation to intensive farming. Moringa (Moringa oleifera) leaf has been proposed to have the antioxidant potential in scavenging free radicals due to the presence of flavonoids. In this study, we investigated the cytoprotective effects of moringa leaf flavonoids in alleviating oxidative stress in BMECs in vitro. Oxidative stress was established by exposing isolated BMECs to H₂O₂ for 2 hr. Doses of moringa leaf flavonoids were evaluated by treating BMECs for 12 hr. The optimal concentrations of H₂O₂ and moringa leaf flavonoids were 500 μmol/L and 1.0 mg/ml, respectively. The results showed that moringa leaf flavonoids increased the activities of superoxide dismutase, catalase, and glutathione peroxidase; and reduced malondialdehyde activity and intracellular accumulation of reactive oxygen species (ROS) in the H₂O₂-induced oxidative stress system. A Hoechst33258 staining assay revealed that moringa leaf flavonoids decreased the apoptosis rate in BMECs, while leaving membrane integrity and nucleolar morphology unchanged. We concluded that moringa leaf flavonoids have the antioxidant capacity to effectively reduce oxidative stress in BMECs.