异位（过）表达PGRN抑制IL-1β引起的髓核细胞损伤

炎症因子IL-1β是引起髓核细胞功能异常的关键因素之一。颗粒体蛋白原（progranulin,PGRN）是一种多功能生长因子,在组织修复、炎症反应等过程中发挥重要作用,但其在髓核细胞中的作用尚不清楚。本研究以IL-1β诱导的髓核细胞炎性损伤模型为研究对象,探讨PGRN对IL 1β诱导的髓核细胞损伤的保护作用及其机制。基因转染结合MTT方法证明,与IL-1β处理的细胞比较,过表达PGRN可逆转IL-1β引起的原代培养的髓核细胞生长抑制,促进细胞增殖。TUNEL技术和流式细胞分析显示,PGRN抑制IL-1β诱导的髓核细胞凋亡。Western印迹和RT-qPCR方法揭示,与IL-1β处理的细胞相比,过表达PGRN显著上调聚蛋白聚糖（aggrecan）和II型胶原（collagen type II）的蛋白质表达,但下调基质金属蛋白酶-13（MMP-13）、分解素-金属蛋白酶ADAMTS-5的表达,同时抑制IL-1β诱导的炎性因子IL-6、IL-8和TNF-α的表达,说明PGRN可缓解IL-1β引起的炎性反应,并减少细胞外基质（ECM）相关蛋白质的降解。此外,过表达PGRN还可降低p65、p-IkB a和β-catenin的蛋白质表达水平,提示PGRN可抑制IL-1β下游TNF-α介导的NF-κB信号途径及β-catenin途径。总之,上述结果提示,过表达PGRN可通过抑制IL-1β诱导的炎性反应、髓核细胞凋亡及基质代谢紊乱,缓解IL-1β诱导的髓核细胞损伤;PGRN的这种抗炎、抗基质降解作用可能与PGRN参与调控NF-κB和β-catenin信号途径有关。     

抑制LRP16基因的表达显著下调TNF-α介导的NF-κB转录活性

LRP16是1个雌激素(E2)通过其受体α(ERα)诱导表达的靶基因.研究表 明,LRP16可以作为多种核受体（包括AR、ERα）的转录共激活因子.采用荧光素酶报 告检测显示,抑制LRP16基因表达显著削弱了TNF-α(10 ng/mL)介导的NF-κB转录活性;采用免疫荧光和Western印迹法研究抑制LRP16对NF-κB/p65亚基核转位的影响,结果显示,抑制LRP16表达并不能参与影响p65亚基核转位．上述结果提示,LRP16可能以核激活因子角色参与了NF-κB介导的信号途径．RT-PCR实验检测抑制LRP16基因表达对TNF-α诱导NF-κB靶基因调控作用,检测的靶基因包括IκB、A20、IL-8、 FLIP、XIAP.结果表明,在这些靶基因中只有XIAP、cIAP2产生了明显的下调趋势. 因此,LRP16是NF-κB的1个共激活因子,通过调控NF-κB与靶基因的结合能力,从而增强了NF-κB的转录活性.     

当归挥发油对哮喘BALB/c小鼠的平喘作用及对Th17免疫活性的影响

目的：探讨当归挥发油对哮喘BALB/c小鼠的平喘作用及对IL-17A、RORγt等Th17细胞活性因子的影响。方法：取雌性小鼠随机分为7组（n=12）：正常对照组、模型对照组、地塞米松组、当归挥发油高中低剂量组及当归油中+地米组,通过注射卵清白蛋白（OVA）致敏、吸入OVA激发的方法来复制哮喘动物模型,在当归挥发油的防治下,考察当归挥发油对哮喘行为学、肺功能、肺组织病理学、血清IL-17A及肺组织RORγt表达的影响。结果：60、120、240 mg/kg当归挥发油可维持哮喘小鼠体重的正常增长,改善哮喘行为学、肺功能及肺组织病理学变化,抑制IL-17A与RORγt的过度表达（P<0.05, 0.01）。同时,当归挥发油与地塞米松配伍后对维持体重的正常增长、缓解IL-17A与RORγt的过度表达具有一定的协同作用。结论：当归挥发油具有明显的防治哮喘作用,抑制IL-17、RORγt过度表达而抑制Th17细胞免疫活性是其作用机制之一;而当归挥发油与糖皮质激素配伍后有一定的协同作用。     

当归对阴虚哮喘小鼠气道黏液高分泌及TNF-α/NF-κB信号通路的影响*

目的: 探讨当归对阴虚哮喘小鼠气道黏液高分泌及TNF-α/NF-κB信号通路的影响。方法: 取KM小鼠随机分为空白对照组、模型对照组、氨溴索组、当归低、中、高剂量(2、4、8 g/kg)组(n=12),采用卵蛋白与甲状腺片复制阴虚哮喘模型,观测当归对小鼠哮喘症状、IgE、TNF-α以及肺组织Muc5ac与NF-κB表达的影响。结果: 2、4、8 g/kg当归能明显缓解阴虚哮喘小鼠的哮喘症状,降低血清IgE与BALF中TNF-α水平,抑制肺组织Muc5ac与NF-κB的过度表达。结论: 当归具有明显的平喘作用,抑制TNF-α/NF-κB信号通路而缓解气道黏液高分泌是其平喘的作用机制之一。     

流感病毒感染巨噬细胞对缺氧诱导因子-1α诱导炎症反应通路的机制研究

为探索缺氧诱导因子(hypoxia inducible factor,HIF)-1α诱导甲型流感病毒毒株感染小鼠巨噬细胞引起炎症反应的具体机制,本研究以甲型H1N1流感病毒(简称H1N1)株A/PR/8感染小鼠巨噬细胞RAW264.7后,在显微镜下观察其在感染后的表型变化,分别在不同时间段收集样本,通过聚合酶链反应(polymerase chain reaction,PCR)检测HIF-1α、干扰素(interferon,IFN)-γ、白细胞介素(interleukin,IL)-6、肿瘤坏死因子(tumor necrosis factor,TNF)-α和M蛋白(M protein,MBP)mRNA的变化,通过蛋白质印迹法(Western blot, WB)检测HIF-1α、核转录因子-κB(nuclear factor-κB,NF-κB)、促分裂原活化的蛋白激酶(mitogenactivated protein kinase,MAPK)、蛋白激酶B(protein kinase B,Akt)以及M蛋白的变化。随后,加入抑制剂2-MeOE-2(10 nmol/L)进行抑制试验,采用PCR和WB检测HIF-1α表达被抑制后上述炎症因子mRNA表达水平及炎症蛋白通路的变化。结果显示,H1N1PR8感染小鼠巨噬细胞RAW264.7后,H1N1PR8复制率在24 h达到峰值,HIF-1α mRNA在感染6 h后开始升高,12 h迅速上升,24 h达到峰值。IFN-γ、IL-6、TNF-α mRNA变化趋势基本与HIF-1α一致,但在感染12 h并未进入快速上升期。HIF-1α蛋白在感染后6 h表达明显增多,24 h达到峰值,与mRNA变化水平基本一致。NF-κB通路蛋白在感染12 h后明显增多,48 h开始减少。加入抑制剂2-MeOE-2后,培养感染细胞24 h,抑制剂组IL-6、TNF-α mRNA水平较对照组显著下降(P<0.05),抑制剂组NF-κB通路蛋白较对照组表达下降。本研究结果表明,小鼠巨噬细胞被H1N1感染后,HIF-1α可能通过激活NF-κB通路促进IL-6、TNF-α等炎症因子的分泌参与炎症反应。     

右美托咪啶对肺缺血/再灌注诱发小鼠肾脏超微结构改变的影响

目的：评价右美托咪啶对小鼠肺缺血/再灌注诱发肾脏损伤的影响。方法：雄性健康SPF级C57BL/6J小鼠50只,体重20 g~24 g,8~10周龄,采用随机数字表法,将其分为5组（n=10）：假手术组（sham组）、肺缺血/再灌注损伤组（I/R组）、肺缺血/再灌注+生理盐水组（NS组）、右美托咪啶组（Dex组）、右美托咪啶+阿替美唑（Atip）（DA组）。采用小鼠在体左侧肺门夹闭30 min再灌注180 min方法制备肺缺血/再灌注损伤（I/R）模型。Dex组在肺门阻断前30 min腹腔注射右美托咪啶20 μg/kg,NS组为用同Dex组等体积的生理盐水替代Dex,DA组腹腔注射右美托咪啶（20 μg/kg）+阿替美唑（250 μg/kg）,其余处理同I/R组。再灌注结束后静脉取血ELISA法检测血浆中IL-1β和TNF-α浓度;取双肾组织,透射电镜下观察肾组织病理学结果。结果：与对照组相比,其余组血浆IL-1β和TNF-α浓度明显升高,肾组织病理学损伤明显加重;与I/R、NS、DA组相比,Dex组IL-1β和TNF-α浓度明显下降,差异有统计学意义（P<0.05）,且肾组织超微结构损伤有所减轻。结论：右美托咪啶预先给药可减轻小鼠肺缺血/再灌注诱发肾脏损伤,其机制可能与抑制炎性反应有关。     

马齿苋多糖对溃疡性结肠炎大鼠肠组织IL-6/STAT3及NF-κB的影响

目的：采用TNBS （2,4,6-三硝基苯磺酸）复制溃疡性结肠炎大鼠模型,探索马齿苋多糖对溃疡性结肠炎大鼠肠组织IL6/STAT3及NF-κB的影响,明确IL-6/STAT3信号通路与慢性炎症性肠病发病的关系,为慢性溃疡性结肠炎的治疗寻找新靶点。方法：将40只SD大鼠随机分为对照组、模型组、美沙拉嗪组和马齿苋组（n=10）。采用TNBS诱导复制结肠炎模型,造模成功后第3天开始灌胃给药：美沙拉嗪组剂量为每次10 mg/kg,每日1次,连续3周;马齿苋组给予马齿苋多糖,每次10 ml/kg,每日1次,连续3周;模型组和对照组大鼠给予等体积生理盐水灌胃,每日1次,连续3周。收集大鼠结肠内容物称重,干燥后再次称重,取结肠组织作病理切片。采用ELISA试剂盒检测血清IL-6、IL-1β、TNF-α和核转录因子-kappa B （NF-κB）含量;免疫组化染色法测定结肠髓过氧化物酶（MPO）;RT-PCR法检测信号转导和转录激活因子（STAT3）、IL-6的mRNA。结果：与模型组、美沙拉嗪组比较,马齿苋组大鼠排便状态明显改善,肠粘膜水肿减轻;血清IL-6、sIL-6Rα、gp130,肠组织MPO、NF-κB含量均降低（P<0.01）。与模型组比较,马齿苋组STAT3、IL-6mRNA的表达水平明显降低（P<0.01）。与对照组比较,上述指标无显著性差异（P>0.05）。结论：马齿苋多糖通过降低大鼠血清IL-6、sIL-6Rα、gp130含量及肠组织MPO、NF-κB水平,减轻sIL-6Rα与IL-6形成复合物所致的炎症反应;经IL-6/STAT3信号通路下调大鼠肠组织STAT3和IL-6的mRNA水平,从而抑制炎症的发生。     

氧化应激对磷酸三钙磨损颗粒诱导的假体周围骨溶解的影响及其机制

目的：探讨氧化应激对磷酸三钙（TCP）磨损颗粒诱导的假体周围骨溶解的影响及其作用机制。方法：36只雄性ICR小鼠随机分为3组（n=12）：假手术（Sham）组、TCP磨损颗粒（TCP）组和N-乙酰-L-半胱氨酸（NAC）组。将TCP磨损颗粒30 mg包埋于小鼠颅骨顶部构建假体周围骨溶解模型,于术后第2天颅顶骨膜局部注射NAC（1.0 mg/kg）,隔日1次,持续2周后处死动物采血、取颅骨。抗酒石酸酸性磷酸酶（TRAP）染色观察小鼠颅骨假体周围骨溶解情况;ELISA和化学比色法检测血清中肿瘤坏死因子-α（TNF-α）、白介素-1β（IL-1β）、白介素-6（IL-6）和总抗氧化能力（T-AOC）含量及超氧化物歧化酶（SOD）活性;Western blot检测假体周围骨组织中内质网应激标志蛋白葡萄糖调节蛋白78（GRP78）、蛋白激酶R样内质网激酶（PERK）、磷酸化PERK（p-PERK）、真核细胞翻译起始因子2α（eIF2α）和磷酸化eIF2α（p-eIF2α）的表达。结果：与Sham组比较,TCP组小鼠血清TNF-α、IL-1β和IL-6水平及假体周围骨溶解面积显著增加（P<0.05）,T-AOC含量和SOD活性明显降低（P<0.05）,GRP78蛋白质表达、p-PERK/PERK和p-eIF2α/eIF2α值显著升高。与TCP组比较,NAC组小鼠血清TNF-α、IL-1β和IL-6水平及骨溶解面积明显减少（P<0.05）,血清T-AOC含量和SOD活性明显增加（P<0.05）,GRP78蛋白质表达、p-PERK/PERK和p-eIF2α/eIF2α值明显降低。结论：抑制氧化应激可阻止TCP磨损颗粒诱导的假体周围骨溶解,其机制可能与PERK/eIF2α通路的失活有关。     

SR 59230A对心衰大鼠心脏MicroRNAs表达的影响

目的：观察β3肾上腺素受体（β3-AR）对心衰大鼠心脏MicroRNAs表达的影响及可能的作用机制。方法：大鼠冠脉左前降支结扎造成心衰模型,假手术大鼠只穿线不结扎。造模成功大鼠再随机分为：心衰组（CHF control group）和心衰+SR 59230A组（CHF+SR group）;假手术大鼠也随机分为假手术组（Sham group）和假手术+SR 59230A组（Sham+SR group）。Sham+SR组和CHF+SR组每天两次腹腔注射SR （85 mmol/L,1 ml）,连续注射7周。结果：①miScript miRNA PCR Arrays显示,在体阻断β3-AR后,假手术组与心衰组有18种MicroRNAs共同表达下调;经文献比对,与NF-κB相关的MicroRNAs有6种,分别为miR-125b-5p,miR-143-3p,miR-145-5p,miR-26a-5p,miR-30a-5p和miR-320-5p。②大鼠心脏组织切片观察到NF-κB在心衰大鼠心肌细胞核与细胞质中均有分布,而p53在心肌细胞质分布较多,NF-κB和p53表达明显高于假手术组（P<0.05）。阻断β3-AR后,心衰组心脏NF-κB和p53表达显著减少（P<0.05）,而假手术组NF-κB和p53表达略增加（P<0.05）。③Western blot结果发现心衰大鼠NF-κB p65表达高于假手术组（P<0.05）,给予β3-AR阻断剂后,心衰组心脏NF-κB p65和p53-Phospho-Serine 15表达均下降（P<0.05）,而假手术组心脏阻断β3-AR后,NF-κB、p53和p53-Phospho-Serine 15表达均增加（P<0.05）。结论：阻断β3肾上腺素受体有利于缓解心衰大鼠心脏的损伤;β3-AR可引起MicroRNAs表达变化且与NF-κB信号通路有关。     

复方扁咽宁对急性咽炎大鼠疗效及其机制

目的:观察复方扁咽宁对急性咽炎大鼠的治疗作用及其作用机理,为其临床应用提供依据。方法:采用氨水直接喷雾法复制大鼠急性咽炎模型,将其随机分为模型组、复方扁咽宁高、中、低剂量组,另取正常大鼠作为对照,每组10只,高、中、低剂量组剂量(g/kg)分别按28.8、14.4、7.2(生药量计)灌胃药物水提液,正常组与模型组给予等量蒸馏水,观察并记录各组大鼠的症状表现;于末次给药24 h后,腹主动脉采血,ELISA法检测血清中IL-1β及TNF-α的含量,采用HE染色法观察各组大鼠肺组织及肺支气管组织形态,透射电镜法观察气管纤毛形态。结果:模型组急性咽炎表现明显:咽部红肿、充血,气管及肺组织出现病理改变,血清中IL-1β及TNF-α的含量显著升高(与正常组比较,P＜0.05),提示造模成功;与模型组相比,复方扁咽宁低、中、高剂量组急性咽炎的症状如咽部红肿、充血等现象得到明显缓解,肺组织及肺支气管组织形态得到明显的改善,血清中IL-1β及TNF-α的含量显著下降,中剂量组和高剂量组疗效更显著。结论:复方扁咽宁可加速急性咽炎的康复进程,改善支气管及肺的组织形态,其作用机制可能与抑制血清IL-1β和 TNF-α的释放等有关。     

Low-dose naltrexone rescues inflammation and insulin resistance associated with hyperinsulinemia

The incidence of diabetes, obesity, and metabolic diseases has reached an epidemic status worldwide. Insulin resistance is a common link in the development of these conditions, and hyperinsulinemia is a central hallmark of peripheral insulin resistance. However, how hyperinsulinemia leads to systemic insulin resistance is less clear. We now provide evidence that hyperinsulinemia promotes the release of soluble pro-inflammatory mediators from macrophages that lead to systemic insulin resistance. Our observations suggest that hyperinsulinemia induces sirtuin1 (SIRT1) repression and stimulates NF-κB p65 nuclear translocation and transactivation of NF-κB to promote the extracellular release of pro-inflammatory mediators. We further showed that low-dose naltrexone (LDN) abrogates hyperinsulinemia-mediated SIRT1 repression and prevents NF-κB p65 nuclear translocation. This, in turn, attenuates the hyperinsulinemia-induced release of pro-inflammatory cytokines and reinstates insulin sensitivity both in in vitro and in vivo diet-induced hyperinsulinemic mouse model. Notably, our data indicate that Sirt1 knockdown or inhibition blunts the anti-inflammatory properties of LDN in vitro. Using numerous complementary in silico and in vitro experimental approaches, we demonstrated that LDN can bind to SIRT1 and increase its deacetylase activity. Together, these data support a critical role of SIRT1 in inflammation and insulin resistance in hyperinsulinemia. LDN improves hyperinsulinemia-induced insulin resistance by reorienting macrophages toward anti-inflammation. Thus, LDN treatment may provide a novel therapeutic approach against hyperinsulinemia-associated insulin resistance.     

B4GALT3促进肝癌细胞增殖和侵袭

β-1,4-半乳糖基转移酶III（β-1,4-galactosyltransferase III,B4GALT3）在肿瘤的作用正受到关注,但其在肝癌中的表达模式及其作用有待阐明。基于TCGA肿瘤组织数据库和GTEx正常组织数据库进行的生物信息学分析,发现相比于人正常肝组织,B4GALT3在人肝癌组织中的表达显著上调。实时荧光定量PCR结果发现肝癌细胞中B4GALT3的mRNA和Western 印迹检测蛋白质表达水平显著上调。其中肝癌细胞SMMC7721中B4GALT3的mRNA表达水平是正常肝细胞L-02的9.85倍。对TCGA数据库进行分析发现,B4GALT3表达水平与肝癌患者的生存率呈负相关。在内源性高表达B4GALT3的SMMC7721肝癌细胞中,干扰B4GALT3表达,可显著抑制该细胞的增殖能力和侵袭能力。干扰B4GALT3表达能显著上调SMMC7721细胞中p27和E-cadherin的蛋白质表达水平,干扰B4GALT3表达后SMMC7721细胞中,p27和E-cadherin的mRNA水平较对照组上调6.15倍和7.83倍。总之,B4GALT3在肝癌中表达上调,且促进肝癌细胞的增殖和侵袭。     

4-1BBL通过激活Wnt/β-catenin信号通路促进人胃癌细胞的增殖和迁移

4-1BB和4-1BB配体(4-1BBL),又被称为CD137和CD137配体,分别属于肿瘤坏死因子(TNF)受体和配体家族的成员。4-1BBL 与4-1BB相互作用可以激活T细胞免疫应答。因此,4-1BBL一直在抗肿瘤免疫应答中发挥经典的免疫共刺激分子作用。近期研究发现,4-1BBL在肿瘤细胞中另有其他的生物学功能,但4-1BBL在胃癌进展过程中的功能尚不明确。本文探讨了4-1BBL在人胃癌细胞中的生物学功能和分子作用机制。首先,通过检索TCGA和Kaplan Meier plotter数据库发现,4-1BBL在胃癌组织中的表达显著高于癌旁组织（P<0.001）,且4-1BBL的高表达与胃癌的不良预后正相关（P<0.05）。细胞生物学的结果显示,敲除4-1BBL明显抑制胃癌细胞的增殖（P<0.05）、侵袭和迁移（P<0.05）,促进胃癌细胞的凋亡（P<0.05）;另外,蛋白质免疫印迹结果表明,敲除4-1BBL可使β-联蛋白、c-Myc和细胞周期蛋白D1(cyclin D1)的蛋白质表达水平下降,抑制Wnt/β-catenin信号通路。相反,过表达4-1BBL则显著促进胃癌细胞增殖（P<0.05）、侵袭和迁移（P<0.05）,减少胃癌细胞的凋亡（P<0.05）;且过表达4-1BBL促进β-联蛋白(β-catenin)、c-Myc和细胞周期蛋白D1的蛋白质表达,激活Wnt/β-catenin信号通路。综上所述,4-1BBL可通过激活Wnt/β-catenin信号通路促进人胃癌细胞的增殖和迁移。     

上调基因4与癌症

上调基因4（up-regulated gene-4,URG4）是受乙肝病毒X蛋白（hepatitis B virus X protein,HBx）激活的下游基因之一,最初在转染HBx的HepG2细胞中被克隆,因其上调细胞增殖效应而得名。与未转染的细胞相比,URG4在HBx转染后的HepG2细胞中表达明显增加,证实了URG4与乙肝病毒相关的肝细胞癌发生相关。近年来,人们发现URG4不仅在肝癌细胞中高表达,还与胃癌、骨肉瘤、非小细胞肺癌、神经母细胞瘤等多种癌症相关。本文结合近年来对URG4的研究成果,包括URG4基因及其蛋白质的结构与功能,URG4在癌症发生发展中的作用,以及在癌症早期诊断和预后中的意义进行综述,可望为后续深入地开展URG4与癌症的研究提供参考。     

The critical role of lipopolysaccharide in the upregulation of aquaporin 4 in glial cells treated with Shiga toxin

Background

In 2011, there was an outbreak of Shiga toxin-producing Escherichia coli (STEC) infections in Japan. Approximately 62 % of patients with hemolytic-uremic syndrome also showed symptoms of encephalopathy. To determine the mechanisms of onset for encephalopathy during STEC infections, we conducted an in vitro study with glial cell lines and primary glial cells.

Results

Shiga toxin 2 (Stx-2) in combination with lipopolysaccharide (LPS), or LPS alone activates nuclear factor-κB (NF-κB) signaling in glial cells. Similarly, Stx-2 in combination with LPS, or LPS alone increases expression levels of aquaporin 4 (AQP4) in glial cells. It is possible that overexpression of AQP4 results in a rapid and increased influx of osmotic water across the plasma membrane into cells, thereby inducing cell swelling and cerebral edema.

Conclusions

We have showed that a combination of Stx-2 and LPS induced apoptosis of glial cells recently. Glial cells are indispensable for cerebral homeostasis; therefore, their dysfunction and death impairs cerebral homeostasis and results in encephalopathy. We postulate that the onset of encephalopathy in STEC infections occurs when Stx-2 attacks vascular endothelial cells of the blood–brain barrier, inducing their death. Stx-2 and LPS then attack the exposed glial cells that are no longer in contact with the endothelial cells. AQP4 is overexpressed in glial cells, resulting in their swelling and adversely affecting cerebral homeostasis. Once cerebral homeostasis is affected in such a way, encephalopathy is the likely result in STEC patients.

Electronic supplementary material

The online version of this article (doi:10.1186/s12929-015-0184-5) contains supplementary material, which is available to authorized users.     

Tumor Necrosis Factor (TNF) Receptor-associated Factor (TRAF)-interacting Protein (TRIP) Negatively Regulates the TRAF2 Ubiquitin-dependent Pathway by Suppressing the TRAF2-Sphingosine 1-Phosphate (S1P) Interaction

The signaling pathway downstream of TNF receptor (TNFR) is involved in the induction of a wide range of cellular processes, including cell proliferation, activation, differentiation, and apoptosis. TNFR-associated factor 2 (TRAF2) is a key adaptor molecule in TNFR signaling complexes that promotes downstream signaling cascades, such as nuclear factor-κB (NF-κB) and mitogen-activated protein kinase activation. TRAF-interacting protein (TRIP) is a known cellular binding partner of TRAF2 and inhibits TNF-induced NF-κB activation. Recent findings that TRIP plays a multifunctional role in antiviral response, cell proliferation, apoptosis, and embryonic development have increased our interest in exploring how TRIP can affect the TNFR-signaling pathway on a molecular level. In our current study, we demonstrated that TRIP is negatively involved in the TNF-induced inflammatory response through the down-regulation of proinflammatory cytokine production. Here, we demonstrated that the TRAF2-TRIP interaction inhibits Lys⁶³-linked TRAF2 ubiquitination by inhibiting TRAF2 E3 ubiquitin (Ub) ligase activity. The TRAF2-TRIP interaction inhibited the binding of sphingosine 1-phosphate, which is a cofactor of TRAF2 E3 Ub ligase, to the TRAF2 RING domain. Finally, we demonstrated that TRIP functions as a negative regulator of proinflammatory cytokine production by inhibiting TNF-induced NF-κB activation. These results indicate that TRIP is an important cellular regulator of the TNF-induced inflammatory response.     

Opposing roles for TRAF1 in the alternative versus classical NF-κB pathway in T cells

T cells lacking TRAF1 hyperproliferate in response to T cell receptor signaling but have impaired signaling downstream of specific TNFR family members such as 4-1BB. Here we resolve this paradox by showing that while TRAF1 is required for maximal activation of the classical NF-κB pathway downstream of 4-1BB in primary T cells, TRAF1 also restricts the constitutive activation of NIK in anti-CD3-activated T cells. Activation of the alternative NF-κB pathway is restricted in unstimulated cells by a cIAP1/2:TRAF2:TRAF3:NIK complex. Using knockdown of NIK by siRNA we show that in activated CD8 T cells TRAF1 is also involved in this process and that constitutive activation of the alternative NF-κB pathway is responsible for costimulation independent hyperproliferation and excess cytokine production in TRAF1-deficient CD8 T cells compared with WT CD8 T cells. The T cell costimulatory molecule 4-1BB critically regulates the survival of activated and memory CD8 T cells. We demonstrate that stimulation through 4-1BB induces cIAP1-dependent TRAF3 degradation and activation of the alternative NF-κB pathway. We also show that while both TRAF1 and cIAP1 have non-redundant roles in suppressing the alternative NF-κB pathway in T cells activated in the absence of costimulation, activation of the classical NF-κB pathway downstream of 4-1BB requires TRAF1, whereas cIAP1 plays a redundant role with cIAP2. Collectively these results demonstrate that TRAF1 plays a critical role in regulating T cell activation both through restricting the costimulation independent activation of NIK in activated T cells and by promoting the 4-1BB-induced classical NF-κB pathway.     

MicroRNA-146a and MicroRNA-146b Regulate Human Dendritic Cell Apoptosis and Cytokine Production by Targeting TRAF6 and IRAK1 Proteins

We have previously reported 27 differentially expressed microRNAs (miRNAs) during human monocyte differentiation into immature dendritic cells (imDCs) and mature DCs (mDCs). However, their roles in DC differentiation and function remain largely elusive. Here, we report that microRNA (miR)-146a and miR-146b modulate DC apoptosis and cytokine production. Expression of miR-146a and miR-146b was significantly increased upon monocyte differentiation into imDCs and mDCs. Silencing of miR-146a and/or miR-146b in imDCs and mDCs significantly prevented DC apoptosis, whereas overexpressing miR-146a and/or miR-146b increased DC apoptosis. miR-146a and miR-146b expression in imDCs and mDCs was inversely correlated with TRAF6 and IRAK1 expression. Furthermore, siRNA silencing of TRAF6 and/or IRAK1 in imDCs and mDCs enhanced DC apoptosis. By contrast, lentivirus overexpression of TRAF6 and/or IRAK1 promoted DC survival. Moreover, silencing of miR-146a and miR-146b expression had little effect on DC maturation but enhanced IL-12p70, IL-6, and TNF-α production as well as IFN-γ production by IL-12p70-mediated activation of natural killer cells, whereas miR-146a and miR-146b overexpression in mDCs reduced cytokine production. Silencing of miR-146a and miR-146b in DCs also down-regulated NF-κB inhibitor IκBα and increased Bcl-2 expression. Our results identify a new negative feedback mechanism involving the miR-146a/b-TRAF6/IRAK1-NF-κB axis in promoting DC apoptosis.     

神经节苷脂对脂多糖损伤大鼠嗜铬细胞瘤细胞的保护作用

目的：研究内源性神经节苷脂对大鼠嗜铬细胞瘤细胞株（PC12）脂多糖（LPS）损伤后的作用及机制。方法：培养PC12细胞,建立LPS损伤模型,采用MTT法检测不同浓度LPS对PC12细胞存活率的改变、葡萄糖神经酰胺合成酶抑制剂D（D-PDMP）耗竭内源性神经节苷脂时LPS对PC12细胞存活率的改变以及添加外源性神经节苷脂GM1后对PC12细胞存活率的保护作用;倒置显微镜和荧光显微镜观察细胞状态;RT-PCR检测NF-κB的相对表达含量。结果：LPS能导致PC12细胞形态学的改变及存活率下降,且随着LPS浓度的增加细胞存活率逐渐降低;神经节苷脂GM1能明显改善LPS所致的细胞形态学以及存活率的改变;工具药D-PDMP耗竭内源性神经节苷脂后,LPS对PC12细胞的损伤更严重,添加外源性神经节苷脂GM1,细胞形态学及存活率得到明显改善,细胞存活率上升;LPS损伤时细胞内NF-κB含量增加;D-PDMP预先耗竭内源性神经节苷脂时NF-κB含量增加更多;添加外源性神经节苷脂GM1后NF-κB含量降低。结论：内源性神经节苷脂对PC12细胞LPS损伤具有保护作用,可能是通过NF-κB信号通路来发挥作用的。