胸腔积液CD14~+细胞表达白介素-16的研究

目的:探讨胸腔积液中CD14~+细胞能否合成和分泌白介素-16(IL-16)。方法:采用流式细胞术(Flow cytometry,FCM)检测细胞内细胞因子,即在胸腔积液中单个细胞水平上进行细胞膜表面抗原的染色和细胞内IL-16的测定;磁珠分选(Magneticcell sorting,MACS)法从胸腔积液中分离出CD14~+细胞,然后进行细胞培养并收集培养上清液,采用酶联免疫吸附法(Enzyme linked immunosorbent assay,ELISA)测定IL-16的浓度。结果:胸腔积液中CD14~+细胞内IL-16显阳性的细胞百分率为(16.28±13.63)%(n=35);磁珠分选出胸腔积液CD14~+细胞,其培养上清液中IL-16的浓度为(58.51±25.38)ng/L(n=5)。结论:在一定条件下,CD14~+细胞(单核/巨噬细胞)有部分能合成和分泌IL-16。     

Selection of mammalian cells based on their cell-cycle phase using dielectrophoresis

An effective, noninvasive means of selecting cells based on their phase within the cell cycle is an important capability for biological research. Current methods of producing synchronous cell populations, however, tend to disrupt the natural physiology of the cell or suffer from low synchronization yields. In this work, we report a microfluidic device that utilizes the dielectrophoresis phenomenon to synchronize cells by exploiting the relationship between the cell's volume and its phase in the cell cycle. The dielectrophoresis activated cell synchronizer (DACSync) device accepts an asynchronous mixture of cells at the inlet, fractionates the cell populations according to the cell-cycle phase (G₁/S and G₂/M), and elutes them through different outlets. The device is gentle and efficient; it utilizes electric fields that are 1–2 orders of magnitude below those used in electroporation and enriches asynchronous tumor cells in the G₁ phase to 96% in one round of sorting, in a continuous flow manner at a throughput of 2 × 10⁵ cells per hour per microchannel. This work illustrates the feasibility of using laminar flow and electrokinetic forces for the efficient, noninvasive separation of living cells.     

Enhanced self-renewal capability in hepatic stem/progenitor cells drives cancer initiation

BACKGROUND & AIMS: Transformed hematopoietic stem/progenitor cells with an enhanced or acquired self-renewal capability function as leukemic stem cells. In a variety of solid cancers, stem/progenitor cells could be also targets of carcinogenesis. However, it remains unclear whether disruption of stem cell function directly contributes to cancer initiation. We sought to elucidate the mechanisms of self-renewal in hepatic stem/progenitor cells and the relation between stem cell function and hepatocarcinogenesis. METHODS: Functional analyses of polycomb-group protein Bmi1 and Wnt/beta-catenin, the molecules that are responsible for the self-renewal capability of many types of stem cells, were conducted in c-Kit(-)CD29(+)CD49f(+/low)CD45(-)Ter-119(-) hepatic stem/progenitor cells using retrovirus- or lentivirus-mediated gene transfer. The tumorigenicity of these cells transduced with the indicated retroviruses was also assessed by transplantation into nonobese diabetic/severe combined immunodeficient mice. RESULTS: Forced expression of Bmi1 and constitutively active beta-catenin mutant similarly promoted the self-renewal of hepatic stem/progenitor cells. The transplantation of Bmi1- or beta-catenin-transduced cells clonally expanded from single hepatic stem/progenitor cells produced tumors, which exhibited the histologic features of combined hepatocellular and cholangiocarcinoma. CONCLUSIONS: These observations imply that the dysregulated self-renewal of hepatic stem/progenitor cells serves as an early event in hepatocarcinogenesis, and they highlight the important roles of Bmi1 and the Wnt/beta-catenin pathway in regulating the self-renewal of normal or cancer stem cells in liver.     

Nitric oxide and acid induce double-strand DNA breaks in Barrett's esophagus carcinogenesis via distinct mechanisms

BACKGROUND & AIMS: The luminal microenvironment including acid and nitric oxide (NO) has been implicated in Barrett's esophagus carcinogenesis. We investigated the ability of acid and NO to induce DNA damage in esophageal cells. METHODS: Transformed and primary Barrett's esophagus and adenocarcinoma cells were exposed to either acid, (pH 3.5), +/- antioxidant or NO from a donor or generated by acidification of nitrite in the presence of ascorbate +/- NO scavenger. Phosphorylation of histone H2AX and the neutral comet assay were used to detect DNA double-strand breaks (DSBs). Intracellular levels of reactive oxygen species and NO were detected with fluorescent dyes. Mitochondrial viability was measured with a rhodamine dye. Long-term survival was assessed by clonogenic assay. RESULTS: Exposure to acid (pH 3.5) for > or =15 minutes induced DSBs in all cell lines (P < .05). There was a concomitant increase in intracellular reactive oxygen species in the absence of mitochondrial damage, and pretreatment with antioxidants inhibited DNA damage. Exposure to physiologic concentrations of NO produced from the NO donor or acidification of salivary nitrite induced DSBs in a dose- (>25 micromol/L) and cell-dependent manner (adenocarcinoma >Barrett's esophagus, P < .05). This occurred preferentially in S-phase cells consistent with stalled replication forks and was blocked with a NO scavenger. NO also induced DSBs in primary Barrett's esophagus cells treated ex vivo. Cells were able to survive when exposed to acid and NO. CONCLUSIONS: Both acid and NO have the potential to generate DSBs in the esophagus and via distinct mechanisms.     

Endosomal trafficking of AMPA-type glutamate receptors

Many different forms of synaptic plasticity have been shown to ultimately modulate the number of AMPA-type glutamate receptors at the synapse. This trafficking involves lateral movements between synaptic and extrasynaptic sites at the neuron surface, as well as vesicular transport between the plasma membrane and intracellular compartments. Several new studies have shed light on the location and regulation of AMPA-type receptor (AMPAR) endocytosis, their intracellular sorting to divergent pathways at the level of endosomes, and the mechanism and sites of receptor recycling. This review summarizes this recent data on the trafficking along the endocytic pathway, and follows the path of internalized AMPAR from endocytosis up to sites of recycling.     

Effects of brain-derived neurotrophic factor on induced differentiation of SH-SY5Y cells in vitro

BACKGROUND： Previous studies have demonstrated that brain-derived neurotrophic factor （BDNF） promotes neural differentiation. However, the mechanisms involved in cell cycle-related protein regulation, which highly correlates to neural proliferation and apoptosis, remain poorly understood. OBJECTIVE： To investigate the effects of various concentrations of BDNF on cycle-related protein mRNA expression in induce-differentiated SH-SY5Y cells in vitro prior to and following G2 phase, and to analyze the neuroprotective effects of BDNF. DESIGN, TIME AND SETTING： A comparison, observational study, based on cell biology, was performed at the Department of Biochemistry, Medical College of Tongji University, from March 2005 to October 2006. MATERIALS： SH-SY5Y cells were provided by Shanghai Institute of Cytology, Chinese Academy of Science; BDNF by Alomone Labs, Israel; all-trans retinoic acid （ATRA） by Sigma-Aldrich, USA. METHODS： SH-SY5Y cells were randomly divided into three groups： blank control [cells were treated in Insulin-Transferrin-Selenium （ITS） solution for 7 days], ATRA （cells were treated with ITS solution containing 10 μmol/L ATRA for 7 days）, and BDNF （cells were treated identical to the ATRA group for 5 days, and then respectively treated in ITS solution containing 1, 10, and 100 μg/L BDNF for 2 days）. The experiment was repeated three times for each group. MAIN OUTCOME MEASURES： mRNA expression levels of cyclin A1, B1, B2, cyclin-dependent kinase 1, and 5 were detected using quantitative real-time RT-PCR; percentage of cells in G1, S, and G2 phases were detected using fluorescence-activated cell sorting. RESULTS： mRNA expression levels of cyclin A1 in the high-dose BDNF group was significantly less than the ATRA group （P 〈 0.05）.mRNA expression levels of cyclin B1 was significantly less in the different BDNF concentration groups compared with the control and ATRA groups （P 〈 0.05 or P 〈 0.01）. mRNA expression levels of cyclin B2 and cyclin-dependent kinase 1 were significantly decreased in the high-dose BDNF group （P 〈 0.05 or P 〈 0.01）. Cyclin-dependent kinase 5 mRNA expression was significantly greater in the low-dose and moderate-dose BDNF groups compared with the ATRA group （P 〈 0.05）. The percentage of cells in G1 phase was significantly greater in the different BDNF concentration groups compared with the ATRA and control groups （P 〈 0.01）. Moreover, the percentage of cells in S phase was significantly less in the three BDNF groups compared with the ATRA group （P 〈 0.01）. However, the percentage of cells in S phase was significantly less in the low-dose and high-dose BDNF groups compared with the control group （P 〈 0.01）. CONCLUSION： BDNF enhanced the percentage of cells in G1 phase, but did not alter mRNA expression of cell cycle-related proteins prior to or following G2 phase. These results suggested that BDNF was not a risk factor for inducing apoptosis.     

全自动药品单剂量分包机的差错分析及对策

目的：分析评价全自动药品单剂量分包机的常见差错,并制定对策。方法：随机抽取调整前后两个时期（调整前：2007年8月15日-2007年10月15日,调整后：2008年1月1日-2月28日）同一病区的摆药医嘱,并统计分析调整前后的差错情况。结果：全自动药品单剂量分包机的差错率由调整前的4.81‰下降到调整后的1.66‰,通过差错调整措施,差错率显著降低（P〈0.05）。结论：全自动药品单剂量分包机具有很高的实用价值,虽然在使用过程中会出现一定的差错,但通过制定对策,差错率可显著降低。     

杭州市生活垃圾分类节点的选择

对源头分类、收集过程二次分类、转运站分类和处置前分类的4个环节的生活垃圾分类实施方式进行比选.选择了源头分类作为杭州市开展生活垃圾分类处理的最优途径.     

实现IHE PIX核心匹配算法的关键技术

患者身份的唯一性识别与整合是实现区域医疗信息共享的基础。IHEPIX是国际上公认的解决患者身份唯一性问题的技术框架,着重介绍了在“医联工程”中实现IHE PiX匹配算法的三种关键技术：身份信息框架、多级停用词划分、关键信息项排序。该匹配算法在项目中的成功运用证实了这些关键技术的有效性。     

Functional role of Alix in HIV-1 replication

Retroviral Gag proteins encode small peptide motifs known as late domains that promote the release of virions from infected cells by interacting directly with host cell factors. Three types of retroviral late domains, with core sequences P(T/S)AP, YPX_nL, and PPPY, have been identified. HIV-1 encodes a primary P(T/S)AP-type late domain and an apparently secondary late domain sequence of the YPX_nL type. The P(T/S)AP and YPX_nL motifs interact with the endosomal sorting factors Tsg101 and Alix, respectively. Although biochemical and structural studies support a direct binding between HIV-1 p6 and Alix, the physiological role of Alix in HIV-1 biology remains undefined. To elucidate the function of the p6–Alix interaction in HIV-1 replication, we introduced a series of mutations in the p6 Alix binding site and evaluated the effects on virus particle production and virus replication in a range of cell types, including physiologically relevant primary T cells and macrophages. We also examined the effects of the Alix binding site mutations on virion morphogenesis and single-cycle virus infectivity. We determined that the p6–Alix interaction plays an important role in HIV-1 replication and observed a particularly severe impact of Alix binding site mutations when they were combined with mutational inactivation of the Tsg101 binding site.