姜黄素抑制NRLP-3表达对抗肿瘤坏死因子α诱导的人血管内皮细胞炎症

背景：前期研究表明,姜黄素对细胞氧化应激炎症有重要作用。目的：拟进一步阐明姜黄素在血管内皮细胞病理性炎症反应中的生物学作用及机制。方法：以人血管内皮细胞为细胞模型,应用肿瘤坏死因子α（10μg/L）作为刺激因子,姜黄素（0,50,100μmol/L）孵育24 h作为干预治疗组。应用HRP标记的BSA检测内皮细胞通透性,应用罗丹明标记的鬼笔碱染色检测F-actin变化,酶联免疫吸附实验检测细胞分泌白细胞介素1β的变化,进一步通过免疫荧光染色检测细胞内核转录因子κB蛋白表达和转位;应用Western blot方法检测炎症小体相关基因NRLP-3和caspase-1的表达。结果与结论：HRP-BSA检测提示,姜黄素可呈剂量依赖性对抗刺激引起的血管内皮细胞通透性增加。同时,抑制F-actin应力纤维的形成。ELISA检测发现,姜黄素治疗后,可呈剂量依赖性抑制肿瘤坏死因子α刺激引起的血管内皮细胞白细胞介素1β分泌。同时,免疫荧光分析证实,肿瘤坏死因子α刺激后,对照组血管内皮细胞中核转录因子κB表达持续增高且发生明显的入核转位现象,而100μg/L姜黄素干预组细胞中炎症递质转录因子核κB表达定位无明显改变。Western blotting结果证实,炎症小体调控基因NRLP3和caspase-1表达在姜黄素干预组受到明显抑制。结果表明姜黄素可通过减少核转录因子κB的表达对抗肿瘤坏死因子α刺激引起的炎症小体活化和白细胞介素1β分泌,对抗细胞病理性炎症损伤发生。     

脊髓型颈椎病患者颈椎间盘白细胞介素6和肿瘤坏死因子α水平与日本骨科学会颈髓功能评分的相关性

目的:检测脊髓型颈椎病患者颈椎间盘组织中白细胞介素6、肿瘤坏死因子α的水平,分析其与日本骨科协会颈髓功能评分(JOA)的相关性,探讨免疫反应在脊髓型颈椎病发病机制中的作用。方法:病例来源于2005-11-10/2006-06-30在安徽医科大学第一附属医院骨科住院患者,27例脊髓型颈椎病患者为脊髓型颈椎病组,另外16例无颈椎病史的颈椎外伤患者为对照组。两组均经颈椎前路手术中切取的颈椎间盘组织,应用放射免疫分析法测定两组椎间盘组织中白细胞介素6和肿瘤坏死因子α的水平。脊髓型颈椎病患者组依据JOA评分分成重、中度组(0～12分,20例)和轻度组(13～17分,7例)。结果:43例全部进入结果分析。①脊髓型颈椎病组颈椎间盘组织内白细胞介素6、肿瘤坏死因子α的水平均明显高于对照组[白细胞介素6:(34.521±18.592),(15.041±6.562)ng/L;肿瘤坏死因子α:(6.071±1.912),(3.143±0.630)pmol/L;P均<0.01]。②脊髓型颈椎病患者重、中度组椎间盘中肿瘤坏死因子α水平明显高于轻度组(P<0.01)。结论:①脊髓型颈椎病患者的颈椎间盘组织内白细胞介素6、肿瘤坏死因子α的分泌增多。②脊髓型颈椎病患者颈椎间盘中肿瘤坏死因子α的水平与脊髓功能有相关性,提示脊髓型颈椎病患者临床症状的轻重与免疫反应之间存在着一定的关系。     

脑性瘫痪儿疾病静止期血清白细胞介素6、肿瘤坏死因子α水平与健康儿童的比较

目的:检测脑性瘫痪(脑瘫)儿疾病静止期血清白细胞介素6、肿瘤坏死因子α水平,并与健康儿童相比较。方法:对象来源于2001-01/2002-12吉林大学第一医院的28例脑瘫儿(脑瘫组)及健康体检儿童15例(对照组),采取静脉血应用酶联免疫吸附试验法测定其血清白细胞介素6、肿瘤坏死因子α水平,并进行两组间的比较。结果:按意向处理分析,①脑瘫组和对照组血清白细胞介素6、肿瘤坏死因子α检测的阳性率基本相似61%(17/28),54%(15/28);47%(7/15),53%(8/15)。②脑瘫组血清白细胞介素6的含量明显高于对照组(41.16±10.52)ng/L,(30.05±2.52)ng/L,t=3.93,P<0.05;肿瘤坏死因子α含量明显高于对照组(37.37±11.84)ng/L,(21.98±3.75)ng/L,t=4.28,P<0.05。结论:脑瘫儿及正常儿童血清中均存在白细胞介素6和肿瘤坏死因子α,在疾病静止期测得白细胞介素6和肿瘤坏死因子α水平显著增高。脑瘫儿确实存在免疫调节机制紊乱,其细胞因子在脑瘫发病机制中的可能作用尚需进一步探讨。     

阿仑膦酸钠对髋关节假体周围界膜组织分泌肿瘤坏死因子α的影响

背景:目前肿瘤坏死因子α的研究有望成为防治假体周围骨丢失的新的切入点,但阿仑膦酸钠对界膜分泌肿瘤坏死因子α的影响及作用机制尚不清楚.目的:观察阿仑膦酸钠对髋关节假体周围界膜组织分泌肿瘤坏死因子α的影响.设计、时间及地点:随机分组设计,对比观察,实验于2006-02/03在广州中医药大学附属骨科医院骨科实验室完成.材料和对象:界膜组织(15 g)取自广州中医药大学第一附属医院(患者知情并同意);雄性新西兰白兔6只用于制备阿仑膦酸钠含药血清,阿仑膦酸钠为石家庄制药厂产品.方法:无菌条件下从人工髋关节假体周围股骨分离界膜组织(15 g),并将其放入RPMI培养液中培养,将界膜组织剪成约250 mg的碎块,计30块,随机分为3组:空白对照组、低浓度阿仑膦酸钠组(100 g/L)、高浓度阿仑膦酸钠组(200 g/L),每组10个样本.低浓度阿仑膦酸钠组和高浓度阿仑膦酸钠组分别加入1.8 mL和1.6 mL,空白对照组加入1.8 mL.然后在低浓度阿仑膦酸钠组、高浓度阿仑膦酸钠组的培养孔中分别加入入0.2 mL和0.4 mL的阿仑膦酸钠含药血清,空白对照组加入0.2 mL空白血清.添加血清后在37℃、含体积分数为5%CO2的饱和湿度条件下培养72 h.主要观察指标:酶联法检测各组界膜组织中肿瘤坏死因子α的表达.结果:低浓度阿仑膦酸钠组和高浓度阿仑膦酸钠组界膜组织分泌肿瘤坏死因子α的量分别为(3.93±0.03),(3.92±0.04)pg/g,与空白对照组(4.04±0.13)pg/g相比,差异有显著性意义(P<0.01).结论:阿仑膦酸钠能显著抑制髋关节假体周围界膜组织分泌肿瘤坏死因子α.     

高迁移率族蛋白B1对磷酸钙诱导巨噬细胞释放炎症因子的协同作用

背景：研究表明,巨噬细胞及其炎症反应参与了肾结石的发生发展。前期实验发现结石晶体可刺激巨噬细胞释放高迁移率族蛋白B1。目的：观察高迁移率族蛋白B1对磷酸钙诱导巨噬细胞释放白细胞介素1β、白细胞介素6、肿瘤坏死因子α、单核细胞趋化因子1的协同作用。方法：实验分两部分：①将成功诱导为巨噬细胞的U937细胞分为空白组、100 mg/L磷酸钙组、100μg/L高迁移率族蛋白B1组、100 mg/L磷酸钙＋100μg/L高迁移率族蛋白B1组,干预1,2,4 h后收集细胞上清液。②将已成功诱导为巨噬细胞的U937细胞分为100 mg/L磷酸钙组、磷酸钙＋10μg/L高迁移率族蛋白B1组、磷酸钙＋50μg/L高迁移率族蛋白B1组、磷酸钙＋100μg/L高迁移率族蛋白B1组,干预4 h后收集细胞上清液。Elisa法检测白细胞介素1β、白细胞介素6、肿瘤坏死因子α、单核细胞趋化因子1水平。结果与结论：ELISA结果显示,磷酸钙组,100μg/L高迁移率族蛋白B1组上清液白细胞介素1β、白细胞介素6、肿瘤坏死因子α、单核细胞趋化因子1质量浓度均高于空白组,磷酸钙＋100μg/L高迁移率族蛋白B1组上清液上述因子质量浓度均显著高于其他3组（P〈0.05）,且呈时间依赖性。不同质量浓度高迁移率族蛋白B1＋磷酸钙组细胞上清液白细胞介素1β、白细胞介素6、肿瘤坏死因子α、单核细胞趋化因子1水平均显著高于磷酸钙组（P〈0.05）,且呈浓度依赖性。结果表明,磷酸钙及高迁移率族蛋白B1均可诱导巨噬细胞释放白细胞介素1β、白细胞介素6、肿瘤坏死因子α、单核细胞趋化因子1;高迁移率族蛋白B1可协同磷酸钙诱导巨噬细胞释放白细胞介素1β、白细胞介素6、肿瘤坏死因子α、单核细胞趋化因子1。     

DAR疗法对SAP大鼠血液TNF-α和IL一6的影响

目的 研究DAR疗法治疗重症急性胰腺炎(SAP)的机制.方法 实验动物为20只SD大鼠.用5%牛黄胆酸钠逆行胰胆管注射法制备SAP模型,造模成功后,随机分为DAR组和对照组,每组10只.DAR组用地塞米松(0.2 mg/kg)、山莨菪碱(0.2 mg/kg)腹腔注射,用生大黄(200 mg/kg)灌胃;对照组用等量生理盐水替代地塞米松、山莨菪碱和大黄,用法同DAR组.两组大鼠均于6 h后活杀.所有动物抽腹主动脉血检测肿瘤坏死因子-α和白细胞介素-6的含量.结果 DAR组腹主动脉血肿瘤坏死因子-α和白细胞介素-6的含量均显著低于对照组.结论 DAR疗法可能通过降低血液肿瘤坏死因子-α和白细胞介素-6的含量,预防和治疗急性胰腺炎导致的全身炎症反应综合征,进一步预防和治疗多器官功能障碍综合征.这可能是DAR疗法治疗SAP的部分机制.     

DAR疗法对SAP大鼠血液TNF-α和IL-6的影响

目的研究DAR疗法治疗重症急性胰腺炎(SAP)的机制。方法实验动物为20只SD大鼠。用5%牛黄胆酸钠逆行胰胆管注射法制备SAP模型,造模成功后,随机分为DAR组和对照组,每组10只。DAR组用地塞米松(0.2mg/kg)、山莨菪碱(0.2mg/kg)腹腔注射,用生大黄(200mg/kg)灌胃;对照组用等量生理盐水替代地塞米松、山莨菪碱和大黄,用法同DAR组。两组大鼠均于6h后活杀。所有动物抽腹主动脉血检测肿瘤坏死因子-α和白细胞介素-6的含量。结果DAR组腹主动脉血肿瘤坏死因子-α和白细胞介素-6的含量均显著低于对照组。结论DAR疗法可能通过降低血液肿瘤坏死因子-α和白细胞介素-6的含量,预防和治疗急性胰腺炎导致的全身炎症反应综合征,进一步预防和治疗多器官功能障碍综合征。这可能是DAR疗法治疗SAP的部分机制。     

川芎嗪可影响肿瘤坏死因子α刺激肝星状细胞结缔组织生长因子、核因子κB及相关基因产物的表达

背景：川芎嗪延缓肝纤维化形成的机制尚不清楚。目的：观察川芎嗪对肿瘤坏死因子α刺激的肝星状细胞增殖及结缔组织生长因子、核因子κB及其相关基因产物白细胞介素6表达的影响。方法：体外培养HSC-T6细胞株,取对数生长期的细胞用于实验。实验分为4组：对照组仅加入细胞;TNF-α组加入10μg/L TNF-α;川芎嗪干预组先加入终浓度为10μg/L的TNF-α作用30min后,分别加入川芎嗪50,100,200,400,600,1000mg/L;PDTC组先加入终浓度为10μg/L的TNF-α作用30min后,再加入终浓度18μmol/L的核因子κB阻断剂PDTC。结果与结论：MTT结果显示100,200,400,600,1000mg/L的川芎嗪均能抑制HSC-T6增殖,且呈剂量依赖性。免疫细胞化学染色及Western blot检测发现10μg/L的肿瘤坏死因子α刺激后,HSC-T6细胞结缔组织生长因子、核因子κB及白细胞介素6的表达明显增多（P〈0.01或P〈0.05）,200,400,600mg/L的川芎嗪及18μmol/L的PDTC均可明显降低肿瘤坏死因子α刺激后HSC-T6细胞结缔组织生长因子、核因子κB及白细胞介素6的表达（P〈0.01）,且随着川芎嗪质量浓度的增加,抑制作用增强,PDTC的抑制作用最明显。相关性分析结果显示HSC-T6细胞结缔组织生长因子和核因子κB的表达呈正相关（r=0.980,P〈0.01）。说明川芎嗪可以抑制肝星状细胞结缔组织生长因子、核因子κB及白细胞介素6的表达,抑制肝星状细胞的增殖。     

醒脑静抑制重组人肿瘤坏死因子介导的人脐静脉内皮细胞增殖

背景：前期研究将传统名方＂安宫牛黄丸＂经提取精制而成的新型水溶性静脉注射液醒脑静注射液应用于临床治疗冠心病并取得很好的疗效,但其具体机制未完全明确。目的：观察醒脑静对重组人肿瘤坏死因子介导的人脐静脉内皮细胞增殖的影响,探讨醒脑静抑制血管内皮细胞损伤增生的应用价值。方法：取3~5代人脐静脉内皮细胞,在培养基中添加10μg/L重组人肿瘤坏死因子α诱导增殖,醒脑静组加入不同浓度的醒脑静干预,阳性对照组添加氟伐他汀,并设立单纯细胞培养的空白对照组。结果与结论：倒置相差显微镜下可见不同浓度的醒脑静均可使人脐静脉内皮细胞呈现胞浆收缩,贴壁细胞减少,坏死细胞增多。醒脑静组的细胞增殖率低于肿瘤坏死因子α组（P〈0.05）,且呈剂量及作用时间依赖关系。证实醒脑静能有效抑制重组人肿瘤坏死因子α介导的人脐静脉内皮细胞增殖。     

微弧氧化镁合金对巨噬细胞肿瘤坏死因子和白细胞介素6的影响

背景:微弧氧化处理可提高镁合金的抗腐蚀性能,延缓其降解速率.目的:观察微弧氧化处理镁合金对小鼠巨噬细胞肿瘤坏死因子α和白细胞介素6的影响.方法:在小鼠RAW264.7细胞中分别加入镁合金浸提液(对照组)、微弧氧化处理镁合金浸提液(实验组)及RPMI-1640(空白对照组),1 h后加入脂多糖,作用24 h后,收集细胞上清液,检测肿瘤坏死因子α和白细胞介素6水平,MTT法测定脂多糖刺激前后的RAW264.7细胞活性.结果与结论:①脂多糖刺激前的RAW264.7细胞活性:对照组>实验组>空白对照组(P均<0.05).②脂多糖刺激后的 RAW264.7细胞活性:实验组与对照组高于刺激前,但差异无显著性意义;空白对照组明显高于刺激前(P <0.01).3组间RAW264.7细胞活性差异无显著性意义(P >0.05).③脂多糖刺激后RAW264.7细胞白细胞介素6与肿瘤坏死因子α的表达量:对照组白细胞介素6表达量明显高于实验组和空白对照组(P <0.05);3组肿瘤坏死因子α表达量两两之间比较差异无显著性意义(P >0.05).表明微弧氧化处理镁合金对小鼠巨噬细胞活性无明显影响,同时降低了炎性反应.     

A transforming growth factor beta-like immunosuppressive factor in immunoglobulin G-binding factor

Immunoglobulin G-binding factors (IgG-BF), which are produced by cells of the immune system, inhibit antibody production. In this paper, we show that transforming growth factor-beta (TGF-beta) suppresses secondary in vitro anti-sheep red blood cell responses of mouse splenocytes and lipopolysaccharide- or anti-IgM-stimulated mouse B cell responses in a way similar to, and with the same kinetics as, rodent IgG-BF. Moreover, the immunosuppressive activity of IgG-BF was totally neutralized by polyclonal and monoclonal anti-TGF-beta antibodies and it eluted with TGF-beta by gel exclusion chromatography, suggesting that a TGF-beta-like immunosuppressive factor is present in IgG-BF. We also show that TGF-beta behaves as an IgG-BF since it binds to insolubilized IgG, but not to insolubilized F(ab')2 or bovine serum albumin. Altogether, the data support the concept of a biological role for TGF-beta in the IgG-mediated negative feedback of antibody responses.     

Measuring tissue factor (factor III) activity in plasma

This is a method for measuring tissue factor (TF, Factor III, tissue thromboplastin) activity in plasma by using a chromogenic substrate. As pretreatment, the euglobulin fraction of plasma was prepared by removing endogenous inhibitors and heated at 60 degrees C for 3 min to remove fibrinogen. This allowed us to measure the low TF activity in plasma that could not otherwise be measured. Neither phospholipids nor coagulation factors VII, IX, X, or Xa in the samples interfere. Within-run and day-to-day reproducibility were both good. The mean value obtained by this method for normal persons was 1.02 (SD 0.91) arbitrary units/L. A markedly high plasma TF activity of 20 arb. units/L or more was observed in patients with some types of disseminated intravascular coagulation.     

Role of human factor VIII in factor X activation.

The cofactor function of human Factor VIII in Factor X activation was investigated by an initial-rate assay of 3H-Factor X activation in the presence of human factor IXa, Ca2+, and either phospholipid or fresh washed human platelets. Purified Factor VIII that has not been activated by thrombin or Factor Xa supports Factor X activation after a lag of several minutes. A specific inhibitor of Factor Xa, which had no inhibitory activity against Factor IXa, markedly prolonged this lag, whereas specific thrombin inhibitors did not prolong the lag. These data support the conclusion that unactivated Factor VIII has no ability to support Factor X activation in a purified system until it is activated by Factor Xa feedback during the lag period. When Factor VIII was optimally preactivated by thrombin, the lag was completely abolished, regardless of the order of addition of the other reactants or the phospholipid source. These data indicate that there is no slow, time-dependent ordering of the reactants at the phospholipid or activated platelet surface if Factor VIII has been preactivated. Unactivated platelets did not support Factor X activation by Factors IXa and VIII. The effect of activated Factor VIII on the kinetics of bovine Factor X activation was primarily to increase the Vmax (54-fold), whereas with human Factor X, Factor VIII both increased the Vmax 56-fold and decreased the Km sixfold to 0.14 microM, similar to the plasma concentration of Factor X. Therefore, a change in the plasma factor X concentration would be expected to have a major effect on the rate of Factor X activation in vivo.     

Human factor in anesthesiology

The authors analyze the role of "human factor" in anesthesiology as a combination of characteristics which can influence the efficiency of physician's work. Anesthesiologists should possess professional skills directly concerned with mental adaptation of personality responsible for decision making in stress or emergency. Examination of 37 anesthesiologists who had given their informed consent helped determine the psychodiagnostic criteria of professional skills and gave grounds for forming groups liable to maladaptation under high psychoemotional strain. Relationships between age, sex and professional activity were assessed. It is recommended to introduce a system of professional selection of candidates wishing to become an anesthesiologist, of individual approach to optimization of the work intensity for practicing anesthesiologists. Psychological relaxation for anesthesiologists is also essential.     

Identification of a macromolecular factor in the ileum which binds intrinsic factor and immunologic identification of intrinsic factor in ileal extracts

The precipitate which resulted when (57)CoB(12) bound to normal human gastric juice was subjected to a 15% concentration of Na(2)SO(4) contained virtually no radioactivity. However, after in vivo incubation of the gastric juice-(57)CoB(12) mixture in the distal ileum of the guinea pig, the dialyzed extract of the washed mucosa contained a fraction of (57)CoB(12) which was precipitated at 15% Na(2)SO(4). In addition, in vitro incubation of gastric juice-(57)CoB(12) with an extract of the ileal mucosa or brush border membranes also resulted in the formation of a 15% Na(2)SO(4)-insoluble fraction which contained (57)CoB(12). The formation of this (57)CoB(12)-containing insoluble fraction did not occur or was diminished by (a) addition of an excess of B(12)-free normal human gastric juice. (b) reducing the incubation pH to 2, (c) incubating the mixture at 4 degrees C, (d) pretreating the ileal extract at 56 degrees C for 30 min, (e) incubating the reaction in sodium EDTA but not calcium EDTA, (f) incubating gastric juice-(57)CoB(12) with an extract of jejunal mucosa. Sephadex gel filtration was used to demonstrate that the factor in the ileal extract which reacted with the gastric juice-(57)CoB(12) filtered through G-100 and G-200 columns in the excluded volume.When the ileal extract obtained after in vivo incubation with gastric juice-(57)CoB(12) was subjected to starch gel electrophoresis one peak of radioactivity remained at the origin and another moved anodally. Eluates of each peak reacted with anti-intrinsic factor antibody indicating that at least the immunologically reacting portion of the intrinsic factor molecule was present in two fractions with different electrophoretic mobility.These studies indicate that immunologically intact intrinsic factor can be extracted from the ileum after in vivo incubation with gastric juice-(57)CoB(12), and that a macromolecular factor is present in the distal ileal mucosa which binds intrinsic factor both in vitro and in vivo, changing its solubility and electrophoretic properties. It is suggested that this ileal binding factor is the previously postulated intestinal receptor for intrinsic factor.     

Tumor necrosis factor alpha-induced angiogenesis depends on in situ platelet-activating factor biosynthesis

Tumor necrosis factor (TNF) alpha, a potent inhibitor of endothelial cell growth in vitro, is angiogenic in vivo. Therefore, it was suggested that the angiogenic properties of this agent might be consequent to the production of secondary mediators. Since TNF-alpha stimulates the synthesis of platelet-activating factor (PAF) by monocytes and endothelial cells, we investigated the possible involvement of PAF in the angiogenic effect of TNF-alpha. Angiogenesis was studied in a murine model in which Matrigel was used as a vehicle for the delivery of mediators. In this model the angiogenesis induced by TNF-alpha was shown to be inhibited by WEB 2170, a specific PAF receptor antagonist. Moreover, in mice injected with TNF-alpha, PAF was detected within the Matrigel, 6 and 24 h after TNF-alpha injection. The synthesis of PAF within the Matrigel was concomitant with the early migration of endothelial cells and infiltration of monocytes. No infiltration of lymphocytes or polymorphonuclear leukocytes was observed. Synthetic PAF as well as PAF extracted and purified from mice challenged with TNF-alpha induced a rapid angiogenic response, inhibited by WEB 2170. These results suggest that the angiogenic effect of TNF-alpha is, at least in part, mediated by PAF synthesized from monocytes and/or endothelial cells infiltrating the Matrigel plug.