Induction of vanadium accumulation and nuclear sequestration causing cell suicide in human Chang liver cells

Very little is known about the modulation of vanadium accumulation in cells, although this ultratrace element has long been seen as an essential nutrient in lower life forms, but not necessarily in humans where factors modulating cellular uptake of vanadium seem unclear. Using nuclear microscopy, which is capable of the direct evaluation of free and bound (total) elemental concentrations of single cells we show here that an NH₄Cl acidification prepulse causes distinctive accumulation of vanadium (free and bound) in human Chang liver cells, concentrating particularly in the nucleus. Vanadium loaded with acidification but leaked away with realkalinization, suggests proton-dependent loading. Vanadyl(4), the oxidative state of intracellular vanadium ions, is known to be a potent source of hydroxyl free radicals (OH^.). The high oxidative state of nuclei after induction of vanadyl(4) loading was shown by the redox indicator methylene blue, suggesting direct oxidative damage to nuclear DNA. Flow cytometric evaluation of cell cycle phase-specific DNA composition showed degradation of both 2N and 4N DNA phases in G₁, S and G₂/M cell cycle profiles to a solitary 1N DNA peak, in a dose-dependent manner, effective from micromolar vanadyl(4) levels. This trend was reproduced with microccocal nuclease digestion in a time response, supporting the notion of DNA fragmentation effects. Several other approaches confirmed fragmentation occurring in virtually all cells after 4 mM V(4) loading. Ultrastructural profiles showed various stages of autophagic autodigestion and well defined plasma membrane outlines, consistent with programmed cell death but not with necrotic cell death. Direct intranuclear oxidative damage seemed associated with the induction of mass suicide in these human Chang liver cells following vanadium loading and nuclear sequestration.     

甜菜碱通过钙通道升高鼠脾淋巴细胞内[Ca^2＋]i研究

研究甜菜碱对小鼠脾淋巴细胞内钙离子浓度的变化及相关钙通道的研究．应用激光共聚焦显微镜（LSCM）测小鼠脾淋巴细胞内钙浓度的变化，应用不同钙通道阻滞剂研究甜菜碱影响细胞内钙浓度变化途径．对终浓度4mmol／L甜菜碱作用淋巴细胞不同时间的细胞内钙离子浓度值分析表明：甜菜碱可以使淋巴细胞内Ca^2＋浓度升高，6h后效果最明显；对加入不同阻滞剂细胞内钙离子浓度变化分析表明：钙通道及蛋白阻滞剂硝苯地平、地尔硫卓、咪贝地尔、金雀异黄素对甜菜碱升高淋巴细胞内钙离子浓度没有影响；维拉帕米、新霉素、肝素、普鲁卡因能阻断甜菜碱对淋巴细胞内钙离子浓度的升高作用．由此可知：细胞内钙离子浓度升高主要通过以下途径：在G蛋白介导下通过影响L-型电压门控钙通道的仅。亚单位而引起外钙内流；通过影响胞内钙库的ILR钙通道和RyR钙通道而引起内钙外排．其共同引起胞质钙离子浓度增加．     

Regulation of the arachidonic acid-stimulated respiratory burst in neutrophils by intracellular and extracellular calcium

The respiratory burst is an important physiological function of the neutrophils in killing the bacteria invading in human body. We used chemiluminescence method to measure the exogenous arachidonic acid-stimulated respiratory burst, and measured the cytosolic free calcium concentration in neutrophils by the fluorescence method. It was found that, on one hand, the arachidonic acid-stimulated respiratory burst was enhanced by elevating the cytosolic free calcium concentration in neutrophils with a potent endomembrane Ca2+-ATPase inhibitor, Thapsgargin; on the other hand, chelating the intracellular or extracellular calcium by EGTA or BAPTA inhibited the respiratory burst. Results showed that calcium plays an important regulatory role in the signaling pathway involved in the exogenous arachidonic acid-stimulated respiratory burst of neutrophils.     

用近红外光谱法实时监测蛹虫草发酵中胞内多糖的质量浓度

应用近红外光谱(NIR)结合偏最小二乘法(PLS)建立一种实时监测蛹虫草发酵中胞内多糖质量浓度的新方法.对39个批次的蛹虫草在3个不同条件的5L发酵罐中进行蛹虫草深层发酵,发酵过程中间隔一定时间取样,采集样品的近红外光谱,并按常规方法测定样品中胞内多糖质量浓度,再采用PLS法建立样品的近红外光谱与胞内多糖质量浓度间的模型,所建模型经过选择最适光谱预处理方法和最适隐变量数进行优化,其留一交互验证预测值与化学测定参考值间的相关系数R=0.8750,交互验证均方根误差RMSECV=0.3052.采用最优PLS模型对样品中胞内多糖质量浓度进行预测,校正集预测均方根误差RMSEC=0.1670,预测集预测均方根误差RMSEP=0.3650,表明模型的稳健性和预测性能较好。     

δ-Protocadherins: unique structures and functions

δ-Protocadherins constitute a group of cadherins characterized by several conserved motifs in their cytoplasmic domains. We present a phylogenetic analysis that further divides this group into δ1-protocadherins (comprising protocadherin-1, −7, −9 and −11 or -X/Y) and δ2-protocadherins (comprising protocadherin-8, −10, −17, −18 and −19). The δ-protocadherin genes, which are located on different chromosomes in man and mouse, have a similar gene structure. They are expressed as multiple splice forms, differing mostly in their cytoplasmic domains. Some δ-protocadherins were reported to mediate weak cell-cell adhesion in vitro and cell sorting in vivo. In addition, individual δ-protocadherins might play important roles in signaling pathways, as they bind to proteins such as TAF1/Set, protein phosphatase-1α and the Frizzled 7 receptor. The spatiotemporally restricted expression of δ-protocadherins in different tissues and species and the results of their functional analysis, mainly in Xenopus, suggest that they play multiple, tightly regulated roles in vertebrate development. Received 18 July 2005; received after revision 26 August 2005; accepted 2 September 2005     

Cellular microbiology of intracellular <Emphasis Type="Italic">Salmonella enterica</Emphasis>: functions of the type III secretion system encoded by <Emphasis Type="Italic">Salmonella</Emphasis> pathogenicity island 2

The facultative intracellular pathogen Salmonella enterica resides in a special membrane compartment of the host cell and modifies its host to achieve intracellular survival and proliferation. The type III secretion system encoded by Salmonella pathogenicity island 2 (SPI2) has a central role in the interference of intracellular Salmonella with host cell functions. SPI2 function affects antimicrobial defense mechanisms of the host, intracellular transport processes, integrity and function of the cytoskeleton and host cell death. These modifications are mediated by translocation of a large number of effector proteins by the SPI2 system. In this review, we summarize recent work on the cellular phenotypes related to SPI2 function and contribution of SPI2 effector proteins to these manipulations. These studies reveal a complex set of pathogenic interferences between intracellular Salmonella and its host cells.Received 11 June 2004; received after revision 8 July 2004; accepted 12 July 2004     

Effect of Aβ1-40 on membrane permeability and intracellular free Ca2+ of nerve cells

The effects of soluble and fibrillar Aβ1-40 on membrane permeability and intracellular free Ca^2 of nerve cells were investigated by the laser confocal microscopy. Results indicate that: i) Effects of soluble and fibrillar Aβ1-40 on cell membrane permeability are both concentration-dependent. Soluble Aβ1-40 increases membrane permeability only at concentration of 3μmol/L, while the toxic effect of fibrillar Aβ1-40 is much stronger, its evident effect begins from 1μmol/L. When its concentration rose to 3μmol/L, not only the membrane permeability increased, but also the nuclear membrane broke seriously, ii) Both soluble and fibrillar Aβ1-40 at high concentrations increased the intracellular free Ca^2 , and the increased amplitudes are concentration-dependent. However, the fibrillar one induces the increase of intracellular Ca^2 much quicker and synchronously.These results indicate that some correlation exists between the neurotoxicity of high concentration soluble and fibrillar Aβ1-40 and the change of physico-chemical properties and intracellular Ca ion imbalance.     

基于表达谱分析筛选肾母细胞瘤关键基因

为探究肾母细胞瘤(Nephroblastoma)发生的关键基因,并筛选出潜在的治疗靶点及生物标志,采用由GEO数据库获取的基因芯片GSE11151和GSE53224,经归一化处理后,通过GO和KEGG分析筛选出差异基因,并通过构建蛋白互作网络获得其中的关键基因.总计获得差异基因404个,其中上调基因385个,下调基因19个.PMCH、CCR5、CCR7、RGS1和KNG1作为关键基因,涉及趋化因子信号通路、G蛋白偶联信号通路,参与肿瘤微环境的形成.这5个关键基因在肾母细胞瘤的发生中有着重要作用,并可能作为潜在治疗靶点及生物标志.