凋亡相关的斑点样蛋白ASC

与凋亡相关的斑点样蛋白ASC是一种含胱天蛋白酶募集域和热蛋白样结构域的蛋白质,它与NF-кB、胱天蛋白酶-1(caspase-1)和热蛋白(pyrin)等多种分子有关,在炎症、细胞凋亡和肿瘤等方面发挥重要作用。     

NF-κB信号转导途径与肿瘤抗凋亡关系的研究进展

细胞核因子κB(NF—κB)家族及其介导的细胞信号转导通路广泛调控着人类免疫和炎症反应中一系列基因的表达,同时也发现它对肿瘤的发生发展有着重要作用,特别是它可以调控一些细胞凋亡相关基因如TRAF家族、IAPs家族、Bcl-2家族及FLIP基因、p53基因、COX-2基因的转录表达,从而大大提高肿瘤细胞的抗凋亡能力。本文就近年来对NF—κB通路与肿瘤抗凋亡关系的研究进展作一综述。     

大肠杆菌感染与人巨噬细胞系U937 细胞凋亡的关系及NF-κB的变化

目的探讨大肠杆菌(E.coli)感染与人巨噬细胞系U937细胞凋亡的关系及核转录因子(nuclear factorkappa B,NF-κB)表达的变化。方法以Annexin V FITC/PI双染流式细胞仪检测及Hoechst 33258荧光染色观察为指标,研究E.coli感染对人巨噬细胞系U937细胞凋亡的诱导作用;用Western blot方法检测NF-ΚB的表达。结果Ho-echst 33258荧光染色结果表明当细胞与细菌浓度比较低时(1:10)可引起部分细胞凋亡,Annexin V FITC/PI双染流式细胞仪结果表明,当细胞与细菌浓度比为1:20,1:50及1:100时,细胞凋亡率与对照组相比明显增高,有显著性差异(P<0.001)。NF-κB的表达随着E.coli浓度的增加而逐渐降低。结论E.coli以剂量依赖的方式诱导U937细胞凋亡,在此过程中NF-κB的表达逐渐降低。     

凋亡抑制蛋白Survivin 研究进展

存活素(Survivin)是凋亡抑制蛋白家族成员之一,具有抑制细胞凋亡和调节细胞周期的双重功能,主要表达于胚胎和发育的胎儿组织中,高表达于大多数恶性肿瘤组织,而在终末分化成熟的正常成人组织中无表达或低表达.本文就Survivin的结构、作用机制、组织分布及其在肿瘤治疗中的研究进展作一综述.     

NF-κB与肿瘤细胞增殖及转移关系的研究进展

NF—κB是一类具有多向调节作用的转录调控因子家族,与肿瘤的发生、发展密切相关。本文综述了NF—κB结构、功能及调节的特性,以及在肿瘤细胞凋亡、增殖及转移中的作用,它有可能成为肿瘤治疗的新靶点。     

红花桑寄生总黄酮提取物诱导淋巴瘤细胞株CA46凋亡及其分子机制研究

研究了一种寄主为夹竹桃的红花桑寄生总黄酮提取物(Nispex)对人Burkitt淋巴瘤细胞株CA46的抗肿瘤作用,探讨了其抗肿瘤作用的分子机制.应用MTT法研究Nispex对CA46细胞增殖的抑制效果,细胞集落培养法观察Nispex对CA46中增殖细胞群的影响.采用AO/EB荧光染色、TUNEL分析、DNA凝胶电泳分析以及AnnexinV流式细胞术检测细胞凋亡,Western blot检测Nispex对CA46细胞中NF-κBp65、Bcl-2、Bax、Caspase-3和PARP等蛋白表达的影响.结果表明,Nispex显著抑制CA46细胞增殖和诱导CA46细胞凋亡,作用48 h的IC50值为1.72μg/mL,细胞凋亡率与药物浓度正相关.Nispex能有效上调CA46细胞Bax、Caspase-3蛋白表达,下调NF-κBp65、Bcl-2、PARP蛋白表达.Nispex诱导CA46细胞凋亡可能是通过对NF-κB信号通路的抑制来实现的.     

NF-κB与持久炎症及肿瘤发生关系

NF-κB是一种序列特异性转录因子.早先的研究证明其主要功能是参与炎症反应和天然免疫应答.最近的研究发现,某一部位的持久炎症反应将导致NF-κB信号通路组成性持续激活,导致NF-κB靶基因的异常表达,这些基因的异常表达往往与肿瘤的发生、转移、组织浸润以及肿瘤细胞的抗凋亡作用相关.因此,将NF-κB作为靶向分子,抑制其活性已成为肿瘤防治的研究热点和新的思路.     

肠道病原菌III型分泌系统效应蛋白调控宿主细胞NF-κB和MAPK信号通路的研究进展

病原细菌感染对人类健康构成了严重的威胁,一类具有III型分泌系统(Type III secretion system,T3SS)的肠道致病细菌可以通过T3SS将效应蛋白“注射”到宿主细胞中,模拟和操纵宿主细胞的多种信号转导通路,包括细胞凋亡、细胞自噬和炎症反应等,从而有效地逃逸宿主的防御,增强感染性和致病性。本文综述了肠道病原菌T3SS效应蛋白在调控宿主炎症反应中NF-κB和MAPK通路的最新研究进展。     

核因子 -kB 在动脉粥样硬化中的研究进展

动脉粥样硬化(atherosclerosis,AS)与心脑血管疾病及周围血管疾病等多种疾病的发病关系密切,而AS的慢性炎症性特征已被广泛认可,从早期内皮功能紊乱到最终的斑块破裂,炎性进程伴随于AS形成的各个阶段。核因子-κB(NF-κB)作为调控炎症信号的重要转录因子,通过调节细胞因子、趋化因子、粘附分子、免疫受体、氧化应激相关酶等一系列特异基因的表达而参与AS的病理过程,并在细胞凋亡和细胞增殖等方面发挥着重要的作用,是AS形成及防治相关研究中的重要靶点。本文通过概述近年来关于NF-κB通路的信号传导在AS形成中的作用,回顾本通路相关药物的研究进展,旨在探讨NF-κB在动脉粥样硬化研究中所面临的问题,为以NF-κB为靶点的抗AS药物研究提供理论参考。     

NF—κB与TNF诱导的细胞凋亡

不同类型的细胞对于ＴＮＦ－α诱志的细胞凋亡的敏感性可以有很大差别,但是多数的细胞在同时给予蛋白质合成抑制处理时会对ＴＮＦ－α变得非常敏感。有学者认为有一个基因或一组基因在ＴＮＦ受体活化后被诱导而传导细胞凋亡信号。近期的研究表明,一种被ＴＮＦ激活的ＮＦ－κＢ转录因子在这一过程中起部分的作用。     

1H, 15N and 13C backbone and side chain chemical shifts of human ASC (apoptosis-associated speck-like protein containing a CARD domain)

The backbone and side chain resonance assignments of human ACS (∼22 KD), apoptosis-associated speck-like protein containing a caspase recruitment domain and a pyrin domain, have been determined by triple-resonance NMR techniques.     

Cloning of a novel human caspase-9 splice variant containing only the CARD domain

Caspase-9 plays a key role in the intrinsic apoptotic pathway and currently two splice variants (caspase-9-alpha and -beta) have been identified. The present study cloned and characterized a novel caspase-9 splice variant, hereby designated Casp9-gamma. Casp9-gamma is generated from an additional alternative 3' splice site in the fourth exon of caspase-9, resulting in a 58-nucleotide fragment insertion compared with the full-length caspase-9-alpha. The fragment introduces an in-frame stop codon, and the resulting open reading frame (ORF) is preterminated. The Casp9-gamma comprises the deduced 154 amino acid residues containing only the caspase recruitment domain (CARD) and does not contain the large and small subunits. The Casp9-gamma does not promote apoptosis when overexpressed in mammalian cells. Moreover, it inhibits the cleavage of procaspase-3 mediated by proapoptotic member Bax or apoptosis inductor staurosporine. Therefore, Casp9-gamma may function as an endogenous apoptotic inhibitor by interfering with the CARD-CARD interaction between Apaf-1 (apoptotic protease activating factor-1) and procaspase-9. In addition, Casp9-gamma does not enhance NF-kappaB activation in transfected 293T cells, conflicting with previous evidence that the isolated CARD of caspase-9 activates NF-kappaB in ND7 cells. This suggests that the procaspase-9-mediated NF-kappaB activation in response to cellular stresses is cell type-specific through an unidentified mechanism.     

Baohuoside I Inhibits the Proliferation of Hepatocellular Carcinoma Cells via Apoptosis Signaling and NF-kB Pathway

Baohuoside I is a flavonoid isolated from Epimedium koreanum Nakai and has many pharmacological activities. However, its role in liver cancer remains unclear. This study aimed to investigate the inhibitory effect of Baohuoside I on the Human Hepatocellular Carcinoma (HCC) cell lines QGY7703, and underlying mechanisms. QGY7703 cells were used as the model to assess the function of Baohuoside I in vitro. The effects of Baohuoside I on QGY7703 cells’ growth, proliferation, and invasiveness were confirmed by CCK-8, lactate dehydrogenase release, and invasion assays. Cell apoptosis was analyzed by flow cytometry, and the levels of cleaved Caspase-3, Bax, and Bcl-2 were quantified by western blot. Western blot analysis, nuclear translocation of NF-κB, and Q-PCR were used to measure the expression of affected molecules. In QGY7703 cells, Baohuoside I induced the expression of molecules related to NF-κB pathway. The toxicity of Baohuoside I on QGY7703 cells was also confirmed in vivo, in a tumor model. Baohuoside I had a significant toxic effect on QGY7703 cells from a concentration of 10 μM. This compound significantly inhibited the proliferation of QGY7703 cells by inducing apoptosis and downregulating NF-κB signaling pathway. Thus, Baohuoside I is a novel candidate drug and opens new possibilities of clinical strategies for HCC treatment.     

Expression of apoptosis-associated speck-like protein containing a caspase recruitment domain, a pyrin N-terminal homology domain-containing protein, in normal human tissues.

Apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC) is a pyrin N-terminal homology domain (PYD)- and caspase recruitment domain (CARD)-containing a proapoptotic molecule. This molecule has also been identified as a target of methylation-induced silencing (TMS)-1. We cloned the ASC cDNA by immunoscreening using an anti-ASC monoclonal antibody. In this study, we determined the binding site of the anti-ASC monoclonal antibody on ASC and analyzed the expression of ASC in normal human tissues. ASC expression was observed in anterior horn cells of the spinal cord, trophoblasts of the placental villi, tubule epithelium of the kidney, seminiferous tubules and Leydig cells of the testis, hepatocytes and interlobular bile ducts of the liver, squamous epithelial cells of the tonsil and skin, hair follicle, sebaceous and eccrine glands of the skin, and peripheral blood leukocytes. In the colon, ASC was detected in mature epithelial cells facing the luminal side rather than immature cells located deeper in the crypts. These observations indicate that high levels of ASC are abundantly expressed in epithelial cells and leukocytes, which are involved in host defense against external pathogens and in well-differentiated cells, the proliferation of which is regulated.     

TRAF2 expression in differentiated muscle

Recent data involving traf2 knockout mice have suggested a necessity of the protein in viability of skeletal muscle tissue. traf2−/− mice are born with decreased muscle mass that is hypothesized to be due to the increased circulating tumor necrosis factor in these mice. We show that TRAF2 protein is present at high levels in terminally differentiated skeletal muscle in the developing mouse. In vitro differentiation of mouse myoblasts displays a dramatic increase in TRAF2 protein levels. Although basal NF-κB activity decreases during myogenesis, TNF-induced NF-κB activity is 10 times greater in myotubes compared with myoblasts, presumably because of the stockpiling of TRAF2 protein in these cells. This may represent a strong anti-apoptotic TRAF2-mediated response specifically tailored to myotubes. These data help explain why muscle integrity is at risk in traf2−/− mice. J. Cell. Biochem. 71:461–466, 1998. © 1998 Wiley-Liss, Inc.     

Caspase-1 protein induces apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC)-mediated necrosis independently of its catalytic activity

The adaptor protein, apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC), connects pathogen/danger sensors such as NLRP3 and NLRC4 with caspases and is involved in inflammation and cell death. We have found that ASC activation induced caspase-8-dependent apoptosis or CA-074Me (cathepsin B inhibitor)-inhibitable necrosis depending on the cell type. Unlike necroptosis, another necrotic cell death, ASC-mediated necrosis, was neither RIP3-dependent nor necrostatin-1-inhibitable. Although acetyl-YVAD-chloromethylketone (Ac-YVAD-CMK) (caspase-1 inhibitor) did not inhibit ASC-mediated necrosis, comprehensive gene expression analyses indicated that caspase-1 expression coincided with the necrosis type. Furthermore, caspase-1 knockdown converted necrosis-type cells to apoptosis-type cells, whereas exogenous expression of either wild-type or catalytically inactive caspase-1 did the opposite. Knockdown of caspase-1, but not Ac-YVAD-CMK, suppressed the monocyte necrosis induced by Staphylococcus and Pseudomonas infection. Thus, the catalytic activity of caspase-1 is dispensable for necrosis induction. Intriguingly, a short period of caspase-1 knockdown inhibited IL-1β production but not necrosis, although longer knockdown suppressed both responses. Possible explanations of this phenomenon are discussed.     

Biochemical and functional analysis of TIR domain containing protein from Brucella melitensis

Toll/interleukin-1 like receptors are evolutionarily conserved proteins in eukaryotes that play crucial role in pathogen recognition and innate immune responses. Brucella are facultative intracellular bacterial pathogens causing brucellosis in animal and human hosts. Brucella behave as a stealthy pathogen by evading the immune recognition or suppressing the TLR signaling cascades. Brucella encode a TIR domain containing protein, TcpB, which suppresses NF-κB activation as well as pro-inflammatory cytokine secretion mediated by TLR2 and TLR4 receptors. TcpB targets the TIRAP mediated pathway to suppress TLR signaling. With the objective of detailed characterization, we have over expressed and purified TcpB from Brucella melitensis in native condition. The purified protein exhibited lipid-binding properties and cell permeability. NF-κB inhibition property of endogenous TcpB has also been demonstrated. The data provide insight into the mechanism of action of TcpB in the intracellular niche of Brucella.