活性氧在七氟烷致人口腔鳞状细胞癌凋亡中的作用研究

目的研究七氟烷在人口腔鳞状癌细胞中的抗癌作用及其潜在的分子机制。方法 MTT比色法检测七氟烷对人口腔鳞状癌细胞的增殖抑制作用;Annexin V-FITC/PI双染法和流式细胞术检测七氟烷对人口腔鳞状癌细胞的诱导凋亡作用、活性氧水平及加入活性氧清除剂N-乙酰半胱氨酸(NAC)后的细胞凋亡情况;利用Western blot实验对凋亡相关蛋白和信号通路相关蛋白进行检测。结果 MTT实验表明七氟烷能够抑制多种人口腔鳞状癌细胞的增殖,并且通过降低线粒体膜电位,调节人口腔鳞状癌细胞中Bcl-2家族、caspase-3和PARP蛋白的表达,最终诱导细胞凋亡。七氟烷还促进了人口腔鳞状癌细胞中活性氧的积累,并上调JNK、p53蛋白的磷酸化表达水平,激活JNK/p53信号通路。然而,在加入NAC之后,不但细胞凋亡情况被抑制,JNK/p53信号通路的激活也被显著逆转。结论七氟烷通过调节活性氧水平来激活人口腔鳞状癌细胞中的JNK/p53信号通路从而诱导细胞凋亡,因此七氟烷有成为治疗人口腔鳞状癌的备选药物的潜力。     

JNK/SAPK信号传递途径与细胞应激反应

c-JunNH_2-末端激酶(JNK)又称为应激活化蛋白激酶(SAPK),是有丝分裂原活化蛋白激酶(MAPK)家族成员之一。大量研究证实,JNK/SAPK信号传递途径在细胞应激反应中起重要作用。JNK/SAPK信号传递途径的激活促进细胞凋亡发生,其机制与诱导FasL表达、调控凋亡相关基因差异表达和改变细胞内Ca~(2 )环境与激活caspases家族有关;在某些情况下,JNK/SAPK信号传递途径的激活并不导致细胞凋亡,相反还可能与细胞增殖有关。JNK/SAPK信号传递途径在细胞应激反应中的作用表现出复杂性和多样性。     

JNK信号通路

c-Jun氨基末端激酶(c-Jun N-terminal kinase, JNK)家族是促分裂原活化蛋白激酶超家族的成员之一,JNK信号通路可被细胞因子、生长因子、应激等多种因素激活,在细胞增殖与分化、细胞凋亡、应激反应等多种细胞调控方面起着至关重要的作用,许多实验证实JNK与许多疾病有密切联系.本文综述JNK信号通路的基本构成、调节方式及在疾病中的作用.     

JNK信号转导通路及其在应激中的作用

JNK是细胞内重要的应激调节蛋白,环境应激和细胞因子均能导致JNK通路的激活.JNK通路除了与细胞生长周期阻滞及细胞凋亡有关外,近年来的研究还发现了JNK通路的其他作用.     

Comparative analysis of the role of JNK signaling pathway in cell proliferation and apoptosis of rat liver regeneration and cirrhosis

目的 从基因转录水平了解JNK信号通路在大鼠肝再生(LR)和肝硬化(LC)中的作用异同. 方法 采用2/3部分肝切除手术制备大鼠肝再生模型,以腹腔注射CCl4中性菜籽油溶液法建立大鼠肝硬化模型,采用大鼠Genome 230 2.0芯片检测不同时间点再生肝和肝硬化组织中JNK信号通路的基因表达谱,用生物信息学和系统生物学等方法分析基因表达谱预示的增殖和凋亡活动. 结果 JNK信号通路涉及302个基因和42条途径,大鼠Genome 230 2.0芯片含上述基因中的240个基因,其中,79个基因发生有意义表达变化,涉及LR的52个基因,LC的5个基因,有22个基因与两者相关.在大鼠LR启动阶段,途径1和16促进细胞增殖及途径22 ～ 33促进细胞凋亡作用强于对照;在进展阶段,途径1～17、34和35促进细胞增殖及途径22 ～33促进细胞凋亡作用强于对照,途径37～41抑制细胞凋亡作用弱于对照;在终止阶段,途径37、39、41和42诱导细胞凋亡作用弱于对照,同时,尚未发现途径18 ～ 21和36参与大鼠LR.而在LC发生中,JNK信号通路中的这些途径与对照相比均无显著差异.结论 JNK信号通路的37条途径调控大鼠肝再生的细胞增殖和凋亡,对大鼠肝硬化的调控作用则不显著.     

JNK信号通路调控大鼠再生肝8种细胞的增殖和凋亡

目的 从基因转录水平了解JNK信号通路在大鼠再生肝8种细胞中的作用。方法 用密度梯度离心和免疫磁珠等方法分离肝细胞（HC）、胆管上皮细胞（BEC）、卵圆细胞（OC）、肝星形细胞（HSC）、窦内皮细胞（SEC）、库普弗细胞（KC）、陷窝细胞（PC）、树突状细胞（DC）等8种肝脏细胞,用大鼠Genome 230 2.0芯片检测大鼠再生肝8种细胞的基因表达谱,用生物信息学和系统生物学等方法分析基因表达变化预示的JNK信号通路在调控大鼠再生肝8种细胞增殖、凋亡中的作用。用实时荧光定量PCR方法验证了芯片结果的可靠性。结果 JNK信号通路涉及240个基因和42条途径,其中,225个基因与大鼠肝再生相关。基因协同作用分析显示,在大鼠肝再生启动阶段,JNK信号通路启动HC和KC增殖,促进OC凋亡,启动部分PC和SEC增殖和促进部分PC和SEC凋亡;在进展阶段,JNK信号通路促进HC、BEC、KC和DC增殖,促进部分PC增殖、部分PC凋亡。在终止阶段,JNK信号通路促进HC、OC和PC凋亡,促进部分KC增殖、部分KC凋亡。结论 大鼠肝再生中JNK信号通路的42条途径和225个基因参与调控大鼠再生肝8种细胞的增殖和凋亡。     

应激活化的蛋白激酶在神经酰胺介导细胞凋亡中的作用

第二信使神经酰胺激活SAPK/JNK,活化的SAPK/JNK导致AP-1转录因子磷酸化,AP-1刺激自身表达,增强自身活性,介导细胞凋亡。神经酰胺及其代谢产物鞘氨醇和鞘氨醇-1磷酸酯分别激活JNK-p38路径和ERK路径,调节细胞增殖、分化或凋亡。     

MAPK信号转导在成骨细胞分化中的作用

MAPK信号转导途径是细胞外信号引起细胞核内反应的通道之一 ,通过三级酶促级联反应 ,最终磷酸化靶蛋白 ,激活转录因子 ,调节特定的基因表达。近年研究表明 ERK、P38和 JNK通路参与了成骨细胞分化增殖的信号转导过程。本文介绍了这三种 MAPK通路的基本概况 ,并着重阐述其在成骨细胞分化中的作用     

13-甲基十四烷酸诱导膀胱癌细胞株T24 凋亡的机制研究

目的： 研究13-甲基十四烷酸诱导膀胱癌T24细胞凋亡的作用,探讨其可能的机制。方法：将不同浓度的13-甲基十四烷酸作用于膀胱癌T24细胞,用MTT法检测细胞增殖, 流式细胞仪（FCM）检测细胞周期,TUNEL法检测细胞凋亡指数,蛋白免疫印迹法分析参与凋亡的蛋白。结果：药物处理2 h后p38、JNK磷酸化增强,AKT磷酸化减弱,8 h后线粒体中细胞色素C释放,FADD及磷酸化FADD没有明显变化,caspase酶底物 PARP、lamin B、RB在12 h出现明显的裂解片段,且药物诱导T24细胞凋亡的过程有时间浓度依赖效应。结论：在13-MTD诱导T24细胞凋亡的过程中p38MAPK和JNK/SAPK信号通路被激活,PI3K/AKT信号通路被抑制,进而激活线粒体凋亡途径,使线粒体内的细胞色素C释放到胞浆,激活caspase酶,裂解相应的凋亡底物lamin B、Rb、PARP,促进T24细胞凋亡。     

神经酰胺:细胞凋亡信号转导的第二信使分子

神经酰胺是细胞凋亡信号调控中的一个第二信使分子,许多应激刺激能激活神经鞘磷脂循环产生神经酰胺诱导多种细胞体系发生凋亡。神经酰胺介导细胞凋亡的机理尚未完全明了,可能是通过多个下游靶分子包括CAPK、CAPP、PKC、SAPK/JNK、CPP32、Bcl-2以及Ras→Racl JNK/p38-K→GADD153信号传导链等作用于不同的信号转导途径而诱导细胞凋亡的。     

Continuous antagonism by Dkk1 counter activates canonical Wnt signaling and promotes cardiomyocyte differentiation of embryonic stem cells

Embryonic stem (ES) cells give rise to mesodermal progenitors that differentiate to hematopoietic and cardiovascular cells. The wnt signaling pathway plays multiple roles in cardiovascular development through a network of intracellular effectors. To monitor global changes in wnt signaling during ES cell differentiation, we generated independent ES cell lines carrying the luciferase gene under promoters that uniquely respond to specific wnt pathway branches. Our results show that successive, mutually exclusive waves of noncanonical and canonical wnt signaling precede mesoderm differentiation. Blocking the initial noncanonical JNK/AP-1 signaling with SP60125 aborts cardiovascular differentiation and promotes hematopoiesis, whereas interference with the subsequent peak of canonical wnt signaling using Dkk1 has the opposite effect. Dkk1 blockade triggers counter mechanisms that lead to delayed and extended activation of canonical wnt signaling and mesoderm differentiation that appear to favor the cardiomyocytic lineage at the expense of hematopoietic cells. The cardiomyocytic yield can be further enhanced by overexpression of Wnt11 leading to approximately 95-fold enrichment in contracting cells. Our results suggest that the initial noncanonical wnt signaling is necessary for subsequent activation of canonical signaling and that the latter operates under a regulatory loop which responds to suppression with hyperactivation of compensatory mechanisms. This model provides new insights on wnt signaling during ES cell differentiation and points to a method to induce cardiomyocytic differentiation without precise timing of wnt signaling manipulation. Taking into account the heterogeneity of pluripotent cells, these findings might present an advantage to enhance the cardiogenic potential of stem cells.     

Differential activation and regulation of mitogen-activated protein kinases through the antigen receptor and CD40 in human B cells.

In human B cells, antigen receptor ligation and CD40 ligation are known to activate the extracellular-regulated kinases (ERK) and c-Jun N-terminal kinase (JNK) pathways, which in turn regulate many important B cell functions. We previously reported that antigen receptor ligation activated the ERK pathway whereas CD40 ligation activated the JNK/stress-activated protein kinase (SAPK) pathway. Here, we demonstrate that another SAPK, p38/Hog1, is activated by both antigen receptor ligation or CD40 ligation in a human B-lymphoblastoid cell line and tonsillar B cells. Wortmannin, an inhibitor of phosphatidylinositol 3-kinase, partially inhibited ERK2 and p38 activation triggered through the B cell receptor whereas activation of JNK1 and p38 through CD40 was not affected. PD98059, a specific inhibitor of mitogen-activated extracellular response kinase kinase (MEK), significantly inhibited ERK2 activation and partially inhibited p38 activation triggered by anti-IgM antibody treatment, but did not affect CD40-dependent signaling events. In addition, anti-IgM antibody-induced signaling pathways were shown to be PKC-dependent in contrast to the CD40-induced signaling pathways. Thus, the B cell receptor and CD40 recruit the ERK, JNK and p38 pathways by using different upstream effectors.     

c-Jun NH2-terminal kinase-mediated signaling is essential for Pseudomonas aeruginosa ExoS-induced apoptosis

As an opportunistic bacterial pathogen, Pseudomonas aeruginosa mainly affects immunocompromised individuals as well as patients with cystic fibrosis. In a previous study, we showed that ExoS of P. aeruginosa, when injected into host cells through a type III secretion apparatus, functions as an effector molecule to trigger apoptosis in various tissue culture cells. Here, we show that injection of the ExoS into HeLa cells activates c-Jun NH(2)-terminal kinase (JNK) phosphorylation while shutting down ERK1/2 and p38 phosphorylation. Inhibiting JNK activation by expression of a dominant negative JNK1 or with a specific JNK inhibitor abolishes ExoS-triggered apoptosis, demonstrating the requirement for JNK-mediated signaling. Following JNK phosphorylation, cytochrome c is released into the cytosol, leading to the activation of caspase 9 and eventually caspase 3. Although c-Jun phosphorylation is also observed as a result of JNK activation, ongoing host protein synthesis is not essential for the apoptotic induction, suggesting that c-Jun- or other AP-1-driven activation of gene expression is dispensable in this process. Therefore, ExoS has opposing effects on different cellular pathways that regulate apoptosis: it shuts down host cell survival signal pathways by inhibiting ERK1/2 and p38 activation, and it activates proapoptotic pathways through activation of JNK1/2 leading ultimately to cytochrome c release and activation of caspases. These results highlight the modulation of host cell signaling by the type III secretion system during interaction between P. aeruginosa and host cells.     

紫草素对非酒精性脂肪性肝炎的影响及分子机制研究

目的 探究紫草素对非酒精性脂肪性肝炎(nonalcoholic steatohepatitis,NASH)是否具有改善作用及其作用机制.方法 给予C57BL/6小鼠胆碱和蛋氨酸缺乏(methionine choline deficient,MCD)饮食诱导NASH模型,检测小鼠体质量、血清转氨酶、血脂以及肝脏中脂质含量...     

Lipid rafts and oxidative stress-induced cell death

Reactive oxygen species (ROS) are generated in response to a number of physiologic or pathologic conditions. In addition to ROS produced extrinsically, a cell may produce ROS as a result of normal metabolism and signaling processes. When sufficient quantities of ROS are present within the cell, this oxidative stress may have profound effects on the cell, including the induction of cell death. Various signaling pathways are initiated in response to oxidative stress, through which the cell's demise is assured. Many of these signaling pathways involve cholesterol-enriched domains of the cell membrane known as lipid rafts. These lipid rafts are platforms for initiation or transduction of the signal and may modulate protein activity through a direct change in local membrane structure or by allowing protein-protein interactions to occur with higher affinity/specificity or both. Among the examples discussed in this review are death-receptor signaling, induction of membrane-associated tyrosine kinase activation, and activation of transient receptor protein (TRP) channels. Special attention also is given to the RIP1/TRAF2 pathway, which involves the downstream activation of the stress-activated protein kinase JNK. The activation of the JNK pathway plays a key role in the induction of cellular death in response to ROS.     

Tumor necrosis factor alpha pathways develops liver apoptosis in type 1 diabetes mellitus

We analyzed the contribution of TNF-α intracellular pathway in the development of apoptosis in the liver of streptozotocin-induced diabetic rats. In liver tissue, diabetes promoted a significant increase of TNF-α/TNF-R1, and led to the activation of caspase-8, of nuclear factor kappa B (NFκB), and JNK signaling pathways. The activation of NFκB led to an induction of iNOS and consequent increase in NO production. As a consequence of such changes a significant increase of caspase-3 activity and of apoptotic index were observed in the liver of diabetic animals. Importantly, the treatment in vivo of diabetic rats with etanercept (TNF-α blocking antibody) or aminoguanidine (selective iNOS inhibitor) significantly attenuated the induction of apoptosis by reduction of caspase-3 activity. Overall, we demonstrated that in the diabetes enhances TNF-α in the liver, which may be a fundamental key leading to apoptotic cell death, through activation of caspase-8, NFκB and JNK pathways.