活性氧在肿瘤坏死因子信号传导中的作用

肿瘤坏死因子α(TNF-α)是最强的细胞内信号诱导剂之一。它通过对半胱氨酸蛋白酶家族(caspases)、丝裂原激活的蛋白激酶(MAPK)、c-jun N端激酶(JNK)、核转录因子AP-1和NF-κB的活化,可诱导细胞凋亡、分化和基因转录。此外,TNF也可通过调节细胞的还原状态,特别是内源性活性氧的改变来介导细胞内的信号。综述了ROS在TNF所诱导细胞内信号传导的相关内容,以阐述ROS介导TNF在细胞内信号传导及调节细胞的存活和凋亡中的作用。     

Caspasess介导老年性痴呆发病机制的研究

Caspases是一类天冬氨酸特异性酶切半胱氨酸蛋白酶,是哺乳动物细胞凋亡的关键蛋白酶。随着研究的深入,发现caspases在神经退行性疾病的病理过程中起着很重要的作用。Caspases在这些疾病的病理过程中,不仅仅是起着凋亡的效应器作用,还能直接与老年性痴呆症、帕金森氏症、亨廷顿舞蹈病和脊椎小脑失调等疾病的致病蛋白质分子相互作用,参与致病机制。因此,文章重点综述caspases在老年性痴呆发病机制中的作用。     

葡萄籽提取物诱导前列腺癌PC-3细胞凋亡的实验研究

该文研究葡萄籽提取物(GSE)对前列腺癌PC-3细胞的生长抑制及凋亡作用,分析GSE诱导PC-3细胞凋亡的途径.采用四唑盐(MTT)比色法测定GSE对PC-3细胞的毒性作用;利用倒置显微镜、透射电镜观察GSE对PC-3细胞引起的形态学改变;碘化丙啶(PI)、吖啶橙(AO)/溴化乙锭(EB)双染法观察GSE诱导PC-3细胞凋亡的作用;Western blot探索细胞凋亡途径.结果表明,MTT法测定GSE能显著抑制PC-3细胞生长.倒置显微镜、透射电镜下可见细胞形态改变.PI染色显示,GSE将PC-3细胞周期阻滞在G0/G1期;GSE诱导PC-3细胞凋亡,AO/EB染色,荧光显微镜下可明显看到凋亡细胞、死亡细胞.Western blot显示GSE通过caspase途径诱导细胞凋亡.从细胞和亚细胞水平研究证实了GSE对人雄激素非依赖性PCaPC-3细胞具有毒性作用,GSE可通过线粒体途径诱导PC-3细胞凋亡和坏死,抑制细胞生长.     

骨髓增生异常综合征的细胞凋亡分子机制(综述)

骨髓增生异常综合征的基本病理特征是骨髓的无效造血。骨髓造血细胞的凋亡增加是导致无效造血的主要原因之一。目前认为 ,骨髓基质细胞分泌的细胞因子如肿瘤坏死因子 (TNF -α)、转化生长因子 β(TGF - β) ,Fas受体 /配体的高表达 ,凋亡调节基因的异常 ,如促凋亡基因 (Bax、Bad)与抗凋亡基因 (Bcl- 2、Bcl-X)的比值增高 ,及半胱氨酸 /天冬氨酸特异的蛋白酶 (caspases)的过度激活等 ,与病人的骨髓造血细胞凋亡增加有关。     

PPAR-α激动剂非诺贝特抑制前列腺癌PC-3细胞生长和诱导凋亡的作用研究

探讨过氧化物酶体增殖因子激活受体α(PPAR-α)激动剂非诺贝特对前列腺癌PC-3细胞生长的抑制和诱导凋亡的作用。分别用0、25、50、75、100μmol/L浓度的非诺贝特作用于PC-3细胞,经过24 h、48 h、72 h后,分别采用MTT、Western blot检测不同组别PC-3细胞的生长或凋亡情况。MTT结果显示:非诺贝特对PC-3细胞的增值存在显著的抑制影响,并且抑制效果和所加入的溶液浓度存在正相关性;流式细胞结果表明:非诺贝特经过24 h、48 h及72 h处理后,测定得到的凋亡细胞总百分比分别为8.41%±2.05%、16.77%±3.16%和23.65%±4.43%,较对照组4.65%±1.12%明显升高。Western blot的检测结果显示:50μmol/L非诺贝特处理PC-3细胞,48 h后其凋亡因子Caspase 3和AIF的表达均有明显的增加。据此,非诺贝特能够显著激活前列腺癌PC-3细胞凋亡因子Caspase 3和AIF的表达,实现诱导细胞凋亡,从而表现出对PC-3细胞增殖的抑制作用。     

TNFα基因在小鼠成纤维细胞L929中的调控研究

质粒pEGFP-1是一种不含启动子序列的哺乳动物细胞表达载体，将系列缺失的TNFα基因启动子5‘UTR（untranslated region）和3‘UTR插入pEGFP-1中，构建了一组重组质粒，转染L929细胞后，发现当报告基因EGFP的上游含有包括5‘UTR在内的TNFα基因启动子序列时，重组质粒在L929细胞中均能表达EGFP，但是，当EGFP基因3‘端同时含有TNFα基因的3‘UTR时，重组质粒转染L929细胞后，均不能表达EGFP，因此，在L929细胞中，TNFα基因的3‘UTR对基因表达具有抑制作用，进一步研究发现，在L929细胞中，TNFα基因的3‘UTR对基因表达的抑制作用需要其5‘UTR的共同参与，而且，RT－PCR实验表明，LPS在其他细胞中对TNFα基因起诱导作用的机制在L929细胞中不存在。     

线粒体与细胞凋亡

线粒体与细胞凋亡关系密切。线粒体在凋亡信号诱导下 ,跨膜电位下降 ,通透性转变 ,位于线粒体内膜外侧的细胞色素C释放进入胞质 ,参与激活caspase - 3。caspase- 3是哺乳动物细胞凋亡的关键蛋白酶     

香蕉皮提取物体外诱导肿瘤细胞凋亡的实验研究

目的:观察香蕉皮提取物体外作用于Hela细胞和PC-3M细胞所致的凋亡作用.方法:从香蕉皮中提取一种粉末状多糖成分,使其作用于宫颈癌 Hela细胞和前列腺癌 PC-3M 细胞株,观察其作用效果.结果:香蕉皮提取物0.5mg/mL、1.0 mg/mL、2.0 mg/mL、4.0 mg/mL浓度,24h诱导Hela细胞凋亡率分别为62.48%、57.91%、52.76%、50.46%,诱导PC-3M的凋亡率分别为60.93%、61.76%、59.33%、63.95%.而在48h 诱导 Hela 的凋亡率为94.44%、93.95%、93.89%、93.91%,诱导 PC-3M 细胞凋亡率分别为93.96%、93.66%、93.97%、94.07%,与对照组比较 P < 0.01,有显著意义.结论:香蕉皮提取物在体外可明显有效地诱导Hela细胞、PC-3M细胞的细胞凋亡.     

整合素αvβ3对正常和癌变的人滋养层细胞分泌基质金属蛋白酶的影响

胚胎植入过程中,滋养层细胞对母体子宫内膜的侵润过程与肿瘤的迁移非常类似,显著区别在于前者是受严格调控的有节制侵润.利用体外培养的人早孕胚胎细胞滋养层细胞和人绒毛膜上皮癌细胞模型,通过RT-PCR、明胶酶谱和免疫细胞化学分析等手段,对比研究了整合素αvβ3对正常和肿瘤细胞产生基质金属蛋白酶的调节.结果表明,正常细胞滋养层细胞和绒毛膜上皮癌细胞均可产生整合素αvβ3;整合素αvβ3抗体能够促进正常滋养层细胞产生基质金属蛋白酶的能力,但抑制绒毛膜上皮癌细胞中基质金属蛋白酶的分泌.这表明整合素能够调节基质金属蛋白酶表达,同时也表明正常和肿瘤化的滋养层细胞中基质金属蛋白酶的表达存在截然不同的调节机制.     

血管内皮细胞凋亡过程中几种癌基因表达的研究

为了研究细胞凋亡的分子调控机制 ,用光学显微技术、DNA凝胶电泳和Northernblot方法 ,研究了去除生长因子 (FGF和血清 )和蛇毒诱导的两个血管内皮细胞凋亡系统中 p53、c H ras、c myc和bcl 2基因的表达 .发现去除生长因子诱导的细胞凋亡过程中 ,p53基因表达显著增加 ,c H ras和c myc基因表达无变化 ;蛇毒诱导细胞凋亡过程中 ,p53基因表达显著增加 ,c H ras和c myc基因表达无变化 .在正常生长和凋亡细胞中均未检测到bcl 2基因的明显表达 .实验结果表明 :p53基因参与上述两种细胞凋亡诱导系统的分子调控 ;c H ras基因只参与去除生长因子诱导的细胞凋亡过程 ,而不参与蛇毒诱导的细胞凋亡过程 ;这两种细胞凋亡诱导系统均与c myc基因表达无关 ;未见bcl 2基因明显参与血管内皮细胞的凋亡过程 .     

Caspase-8 regulates TNF-α-induced epithelial necroptosis and terminal ileitis

Dysfunction of the intestinal epithelium is believed to result in the excessive translocation of commensal bacteria into the bowel wall that drives chronic mucosal inflammation in Crohn's disease, an incurable inflammatory bowel disease in humans characterized by inflammation of the terminal ileum. In healthy individuals, the intestinal epithelium maintains a physical barrier, established by the tight contact of cells. Moreover, specialized epithelial cells such as Paneth cells and goblet cells provide innate immune defence functions by secreting mucus and antimicrobial peptides, which hamper access and survival of bacteria adjacent to the epithelium. Epithelial cell death is a hallmark of intestinal inflammation and has been discussed as a possible pathogenic mechanism driving Crohn's disease in humans. However, the regulation of epithelial cell death and its role in intestinal homeostasis remain poorly understood. Here we demonstrate a critical role for caspase-8 in regulating necroptosis of intestinal epithelial cells (IECs) and terminal ileitis. Mice with a conditional deletion of caspase-8 in the intestinal epithelium (Casp8(ΔIEC)) spontaneously developed inflammatory lesions in the terminal ileum and were highly susceptible to colitis. Casp8(ΔIEC) mice lacked Paneth cells and showed reduced numbers of goblet cells, indicating dysregulated antimicrobial immune cell functions of the intestinal epithelium. Casp8(ΔIEC) mice showed increased cell death in the Paneth cell area of small intestinal crypts. Epithelial cell death was induced by tumour necrosis factor (TNF)-α, was associated with increased expression of receptor-interacting protein 3 (Rip3; also known as Ripk3) and could be inhibited on blockade of necroptosis. Lastly, we identified high levels of RIP3 in human Paneth cells and increased necroptosis in the terminal ileum of patients with Crohn's disease, suggesting a potential role of necroptosis in the pathogenesis of this disease. Together, our data demonstrate a critical function of caspase-8 in regulating intestinal homeostasis and in protecting IECs from TNF-α-induced necroptotic cell death.     

凋亡抑制蛋白XIAP研究进展

X连锁凋亡抑制蛋白(XIAP)是哺乳动物中具有抑制细胞凋亡作用的蛋白,是IAPs家族的一员.XIAP通过杆状病毒IAP重复序列(BIR)直接与起始以及效应caspases结合,抑制了细胞凋亡的线粒体途径,也可以通过NF-κB途径抑制细胞表面受体介导的凋亡.XIAP具有不同于Bcl-2的作用机制,是IAPs家族中最具有抑制活性的一个.XIAP的作用受到线粒体释放的蛋白Smac的拮抗,以及受到自身具有泛素连接酶E3活性的RING指结构域的调节.阐述XIAP抑制caspase以及Smac等拮抗XIAP的机理对于治疗肿瘤以及过度凋亡疾病具有重要的意义.     

Treatment of PC-3 cells with ultrasound combined with microbubbles induces distinct alterations in the expression of Bcl-2 and Bax

The objective of the present study was to investigate whether ultrasound combined with microbubbles induces apoptotic cell death in androgen-independent prostate cancer cells and to identify the probable mechanism. We used ultrasound in continuous wave mode with a frequency of 21 kHz and a spatial-average temporal-average intensity of 46 mW/cm². Ultrasound combined with microbubbles (200 μL SonoVue?) was used to treat androgen-independent human prostate cancer PC-3 cells for 30 s. PC-3 cells were divided into three groups: the control group, the ultrasound group and the ultrasound combined with microbubbles group. Immediately after treatment, trypan blue exclusion was used to assess cell viability. Cell apoptosis at 24 h after treatment was measured using transmission electron microscopy and flow cytometry. Western blotting was used to evaluate the expression of the apoptosis-related proteins, Bcl-2 and Bax. Ultrasound combined with microbubbles had a minimal effect on the viability of PC-3 cells and induced minimal levels of cell lysis. The level of apoptosis in PC-3 cells induced by this modality was significantly higher than in controls (12.77 ± 0.31% vs. 2.56 ± 0.22%, P<0.01). Treatment with ultrasound combined with microbubbles increased the expression of Bax, a pro-apoptotic protein, and decreased the expression of Bcl-2, an anti-apoptotic protein. It was concluded that ultrasound combined with microbubbles induces apoptotic cell death in human prostate cancer PC-3 cells through down-regulation of Bcl-2 and up-regulation of Bax.     

Suppression of CED-3-independent apoptosis by mitochondrial betaNAC in Caenorhabditis elegans

To ensure cell survival, it is essential that the ubiquitous pro-apoptotic machinery is kept quiescent. As death is irreversible, cells must continually integrate developmental information with regulatory inputs to control the switch between repressing and activating apoptosis. Inappropriate activation or suppression of apoptosis can lead to degenerative pathologies or tumorigenesis, respectively. Here we report that Caenorhabditis elegans inhibitor of cell death-1 (ICD-1) is necessary and sufficient to prevent apoptosis. Loss of ICD-1 leads to inappropriate apoptosis in developing and differentiated cells in various tissues. Although this apoptosis requires CED-4, it occurs independently of CED-3--the caspase essential for developmental apoptosis--showing that these core pro-apoptotic proteins have separable roles. Overexpressing ICD-1 inhibits the apoptosis of cells that are normally programmed to die. ICD-1 is the beta-subunit of the nascent polypeptide-associated complex (betaNAC) and contains a putative caspase-cleavage site and caspase recruitment domain. It localizes primarily to mitochondria, underscoring the role of mitochondria in coordinating apoptosis. Human betaNAC is a caspase substrate that is rapidly eliminated in dying cells, suggesting that ICD-1 apoptosis-suppressing activity may be inactivated by caspases.     

Phytophthora elicitor PB90 induced apoptosis in suspension cultures of tobacco

The protein elicitor PB90 secreted by Phytophthora boehmeriae is an efficient elicitor inducing the hypersensitive response and systemic acquired resistance in tobacco plants. Here, we observed cell death in suspension-cultured cells of Nicotiana tabacum BY-2 with PB90 treatment using Trypan blue staining method. And this cell death could be suppressed by cycloheximide, an inhibitor of proteins synthesis, which implies that PB90-induced celldeath was an active cell death process requiring new protein synthesis. DAPI staining revealed that PB90 induce rapid chromatin condensation, margination, apoptotic bodies‘ formation and DNA laddering, further TUNEL assay also observed the specific breakage of 3 ‘-OH ends. All of the above common morphological characteristics indicated that PB90 induced apoptosis in suspension cultures of tobacco, suggesting that hypersensitive response induced by PB90 is an apoptotic process.     

Endonuclease G is an apoptotic DNase when released from mitochondria.

Nucleosomal fragmentation of DNA is a hallmark of apoptosis (programmed cell death), and results from the activation of nucleases in cells undergoing apoptosis. One such nuclease, DNA fragmentation factor (DFF, a caspase-activated deoxyribonuclease (CAD) and its inhibitor (ICAD)), is capable of inducing DNA fragmentation and chromatin condensation after cleavage by caspase-3 (refs 2,3,4). However, although transgenic mice lacking DFF45 or its caspase cleavage site have significantly reduced DNA fragmentation, these mice still show residual DNA fragmentation and are phenotypically normal. Here we report the identification and characterization of another nuclease that is specifically activated by apoptotic stimuli and is able to induce nucleosomal fragmentation of DNA in fibroblast cells from embryonic mice lacking DFF. This nuclease is endonuclease G (endoG), a mitochondrion-specific nuclease that translocates to the nucleus during apoptosis. Once released from mitochondria, endoG cleaves chromatin DNA into nucleosomal fragments independently of caspases. Therefore, endoG represents a caspase-independent apoptotic pathway initiated from the mitochondria.     

Structural and biochemical basis of apoptotic activation by Smac/DIABLO

Apoptosis (programmed cell death), an essential process in the development and homeostasis of metazoans, is carried out by caspases. The mitochondrial protein Smac/DIABLO performs a critical function in apoptosis by eliminating the inhibitory effect of IAPs (inhibitor of apoptosis proteins) on caspases. Here we show that Smac/DIABLO promotes not only the proteolytic activation of procaspase-3 but also the enzymatic activity of mature caspase-3, both of which depend upon its ability to interact physically with IAPs. The crystal structure of Smac/DIABLO at 2.2 A resolution reveals that it homodimerizes through an extensive hydrophobic interface. Missense mutations inactivating this dimeric interface significantly compromise the function of Smac/DIABLO. As in the Drosophila proteins Reaper, Grim and Hid, the amino-terminal amino acids of Smac/DIABLO are indispensable for its function, and a seven-residue peptide derived from the amino terminus promotes procaspase-3 activation in vitro. These results establish an evolutionarily conserved structural and biochemical basis for the activation of apoptosis by Smac/DIABLO.     

TNF和OA诱发人神经母细胞瘤SK细胞程序死亡

肿瘤坏死因子（ＴＮＦ）和Ｏｋａｄａｉｃａｃｉｄ（ＯＡ）能诱发人神经母细胞瘤ＳＫ细胞死亡，死亡细胞缩小变圆，细胞质凝聚，ＤＮＡ断裂形成以２００ｂＰ左右为单位的梯形分布，且上述过程可由蛋白质合成抑制剂亚胺环己酮（ＣＨＸ）抑制，显示ＴＮＦ和ＯＡ诱发的ＳＫ细胞死亡为细胞程序死亡．