Intracellular localisation of Mycobacterium marinum in mast cells

AIM:To study the bacteriocidal or bacteriostatic role of mast cells during infection with Mycobacterium.METHODS:Mycobacterium marinum(M.marinum)(BAA-535/M strain)was investigated for its ability to grow at a temperature relevant to the mammalian host.Primary mast cells were differentiated from bone marrows of mice,a human mast cell line(HMC-1)and a human monocytic cell line(Mono Mac6)were maintained in culture.Mice were stimulated by intraperitoneal injection of heat-killed M.marinum to study cytochemically the degranulation of peritoneal mast cells.HMC-1 cells were stimulated with M.marinum to analyse m RNA expression for inflammatory reactant genes,while HMC-1 and primary mouse mast cells were infected with M.marinum to establish in parallel cell viability(lactate dehydrogenase release and cell counts)and viable mycobacterial counts.Flow cytometry was used to assess intracellular presence of fluorescein isothiocyanate labelled M.marinum after trypan blue quenching and to measure the extent of infection-induced apoptosis or necrosis in HMC-1.A GFP expressing recombinant M.marinum strain was used to assess intracellular location by fluorescence microscopy.Light microscopy of osmium tetroxide and Gram Twortstained sections of 0.5 μm and transmission electron microscopy were undertaken as sensitive methods.RESULTS:Since its isolation,M.marinum has adapted to grow at 37 ℃.This study found that M.marinum infects HMC-1 cells and primary murine mast cells,where they survive,replicate,and cause dose dependent cell damage over the analysis period of up to 120 h.Amikacin was an effective aminoglycoside antibiotic to eliminate extracellular or membrane attached M.marinum in order to adequately quantify the intracellular bacterial loads.In vivo,intraperitoneal injection of heat-killed M.marinum led to the release of mast cell granules in mice.HMC-1 cells stimulated with M.marinum showed a biphasic pattern of increased mR NA expression for LL-37 and COX-2/TNF-a during 24 h of stimulation.In HMC-1,M.marinum localised to the cytoplasm whereas in primary mast cells,M.marinum were found in vacuoles.CONCLUSION:The effector role of mast cells in infection with M.marinum can be studied in vitro and in vivo.

Bone marrow mesenchymal stem cell transplantation retards the natural senescence of rat hearts

Aim Aging is an independent risk factor for heart disease, however the effective intervention has not been found so far. Bone marrow mesenchymal stem cells （BMSCs） have been shown to offer a wide variety of cel- lular functions including the protective effects on damaged hearts. Here we investigated the antiaging properties of BMSCs and the underlying mechanism in a cellular model of cardiomyocyte senescence and a rat model of aging hearts. Methods In vitro study, neonatal rat ventricular cells （NRVCs） and BMSCs were cocultured in the same dish with a semipermeable membrane to separate the two populations. In vivo, the BMSCs were injected into the rat hearts to observe their antiaging effects. The expression of β-galactosidase and aging-related proteins, and the lev- els of oxidative stress were determined in vivo and in vitro. The heart function was measured by the High-Resolution Imaging System. Results Monocultured NRVCs displayed the senescence-associated phenotypes, characterized by an increase in the number of β-galaetosidase-positive cells and decreases in the degradation and disappearance of cellular organelles in a time-dependent manner. The levels of reactive oxygen species and malondialdehyde were el- evated, whereas the activities of antioxidant enzymes superoxide dismutase and glutathione peroxidase were de- creased, along with upregulation of p53, p21cipl/wafl and p16INK4a in the aging eardiomyoeytes. These deleterious alterations were abrogated in aging NRVCs cocultured with BMSCs. Qualitatively, the same senescent phenotypes were consistently observed in aging rat hearts. Notably, BMSC transplantation significantly prevented these detri- mental alterations and improved the impaired cardiac function in the aging rats. Conclusions BMSCs possess strong antiseneseence action on the aging NRVCs and hearts and can improve cardiac function after transplantation in aging rats. The present study, therefore, provides an alternative approach for the treatment of heart failure in the elderly population.

Cell Evolutionary Algorithm: a New Optimization Method on Ground-State Energy of the Atomic

The purpose of this paper is to present a new general approach to solve ground-state energies of the double-electron systems in a uniform magnetic field, in which the basic element of evolution is the set in the solution space, rather than the point. The paper defines the Cell Evolutionary Algorithm, which implements such a view of the evolution mechanism. First, the optimal set in which the optimal solution may be obtained. Then this approach applies the embedded search method to get the optimal solution. We tested this approach on the atomic structure, and the results show that it can improve not only the efficiency but also the accuracy of the calculations as it relates to this specific problem.

连翘酯苷在体外对鸡IFN-α和JAK-STAT信号通路相关因子的影响(英文)

[目的]探讨连翘酯苷作用于鸡胚肾(CEK)细胞后,IFN-α的表达变化及JAK-STAT通路相关因子mRNA的表达变化,及连翘酯苷对IFN-α和JAK-STAT信号通路的影响。[方法]将连翘酯苷设为3个浓度(100、200和400μg/ml),采用荧光定量RT-PCR,检测IFN-α和JAK-STAT通路相关因子的mRNA表达变化;用western blotting检测IFN-α的蛋白表达情况。[结果]与正常组相比,连翘酯苷不仅显著提高了IFN-α的表达量(P<0.05);而且明显上调了STAT1、JAK1、IFNAR1、IFNAR2、IRF1、IRF7的表达。[结论]连翘酯苷能够在鸡胚肾细胞(chicken embryo kidney cells,CEKs)上诱导IFN-α的表达,且能够正向调节JAK-STAT信号通路传导。

玉米赤霉烯酮对猪睾丸间质细胞DNA损伤作用的彗星试验(英文)

[目的]观察玉米赤霉烯酮(ZEN)对猪睾丸间质细胞(Leydig cell)的DNA损伤效应。[方法]以体外培养的猪Leydigcell为材料,用四氮唑蓝比色分析法(MTT法)测定ZEN对离体培养的猪睾丸间质细胞的半数致死浓度,选用0(对照组)、1、5、10和20μmol/L浓度的ZEN体外作用于猪Leydigcell,通过彗星试验观察了ZEN对猪Leydig cell DNA的损伤效应。[结果]ZEN浓度为0、1、5、10和20μmol/L时,所造成的细胞拖尾率分别为16.67%、34.00%、40.67%、52.00%和64.67%,各染毒组细胞拖尾率与对照组比较,差异均有极显著统计学意义(P<0.01),且存在明显的剂量-效应关系;各剂量组对应的拖尾细胞尾长(Taillength)分别为57.60±4.78、57.75±6.25、78.97±5.83、100.50±6.94和146.83±12.31μm,尾部DNA含量(Tail DNA %)分别为21.29±2.25%、22.24±2.43%、31.21±6.27%、37.45±4.33%和60.68±9.83%,与对照组比较,5μmol/L及以上ZEN浓度染毒组的Tail length和Tail DNA %均有显著性差异(P<0.05),且随ZEN浓度增加呈上升趋势。[结论]ZEN对猪Leydig cell存在遗传毒性,可以损伤Leydig cell的DNA,且有明显的剂量-效应关系。

转基因植物中Bt Cry2Ab蛋白的抗原表位特征预测(英文)

[目的]通过生物信息学方法了解转基因作物中BtCry2Ab蛋白的二级结构、三级结构、B细胞抗原表位及T细胞抗原表位等结构特征,为今后的抗体设计奠定基础。[方法]以在NCBI数据库中获得Cry2Ab蛋白序列为材料,应用DNAStar中多种方法预测其B细胞表位,并通过在线预测软件NetMHCII2.2分析Cry2Ab蛋白与MHC-II类分子的结合能力从而预测其T细胞抗原表位。[结果]对Cry2Ab蛋白的B细胞抗原表位的预测表明,Cry2Ab蛋白的第208～215区域是潜在的B细胞抗原表位。对Cry2Ab蛋白与MHC-II类分子结合能力的分析表明,Cry2Ab蛋白的第177～185区域、299～307区域和255～263区域是潜在的T细胞抗原表位,暗示含有HLA-DRB10101和HLA-DRB10701等位基因的人群对Cry2Ab蛋白更敏感。[结论]该研究有助于深入了解Cry2Ab蛋白的生物学特征,为完善转基因食品过敏原性的评估方法提供新线索。