The CHD3 remodeler PICKLE associates with genes enriched for trimethylation of histone H3 lysine 27

In Arabidopsis (Arabidopsis thaliana), the ATP-dependent chromatin remodeler PICKLE (PKL) determines expression of genes associated with developmental identity. PKL promotes the epigenetic mark trimethylation of histone H3 lysine 27 (H3K27me3) that facilitates repression of tissue-specific genes in plants. It has previously been proposed that PKL acts indirectly to promote H3K27me3 by promoting expression of the POLYCOMB REPRESSIVE COMPLEX2 complex that generates H3K27me3. We undertook expression and chromatin immunoprecipitation analyses to further characterize the contribution of PKL to gene expression and developmental identity. Our expression data support a critical and specific role for PKL in expression of H3K27me3-enriched loci but do not support a role for PKL in expression of POLYCOMB REPRESSIVE COMPLEX2. Moreover, our chromatin immunoprecipitation data reveal that PKL protein is present at the promoter region of multiple H3K27me3-enriched loci, indicating that PKL directly acts on these loci. In particular, we find that PKL is present at LEAFY COTYLEDON1 and LEAFY COTYLEDON2 during germination, which is when PKL acts to repress these master regulators of embryonic identity. Surprisingly, we also find that PKL is present at the promoters of actively transcribed genes that are ubiquitously expressed such as ACTIN7 and POLYUBIQUITIN10 that do not exhibit PKL-dependent expression. Taken together, our data contravene the previous model of PKL action and instead support a direct role for PKL in determining levels of H3K27me3 at repressed loci. Our data also raise the possibility that PKL facilitates a common chromatin remodeling process that is not restricted to H3K27me3-enriched regions.     

The polysaccharides from Ganoderma lucidum: Are they always inhibitors on human hepatocarcinoma cells?

The antitumor activity of intracellular polysaccharides from submerged fermentation of Ganoderma lucidum was investigated focusing on the inhibition on human liver cancer cells. The polysaccharides inhibited human hepatocarcinoma cell HepG2 during earlier phase with lower dosage but obviously became less functional in later phase regardless of the dosage applied. However, apoptosis of the drugged HepG2 cells appeared in later incubation phase with high dosage, and the apoptosis could be enhanced by supplemental dose of the intracellular polysaccharides. Nevertheless, the intracellular polysaccharides inhibited other human hepatocarcinoma cells such as BEL-7402 and Huh-7 but luckily stimulated human normal liver cell L02 only in a positive dose- and time-dependent manner; so did the sulfated extracellular polysaccharides when it inhibited HepG2 and L02 cells. However, the toxicity of sulfated extracellular polysaccharides to L02 cells can be eliminated by the intracellular polysaccharides.     

大鼠多囊卵巢颗粒细胞凋亡时MDA含量及SOD活性的变化

目的和方法以大鼠多囊卵巢（PCO）为动物模型,观察了PCO大鼠卵泡颗粒细胞凋亡的发生率,并检测了颗粒细胞凋亡时丙二醛（MDA）的含量及超氧化物歧化酶（SOD）活性的变化。结果①PCO颗粒细胞凋亡发生率明显高于正常（P<0．001）;②PCO颗粒细胞发生凋亡时,细胞内MDA含量增加而SOD活性降低。结论大鼠PCO颗粒细胞的凋亡可能与细胞内MDA含量增加和SOD活性降低有关。     

荧光定量RT-PCR检测血管内皮细胞PAI-1 mRNA方法的建立

为了建立用荧光定量PCR检测人微血管内皮细胞纤溶酶原激活剂抑制物-1（PAI-1）mRNA表达的方法。提取人微血管内皮细总RNA,经RT-PCR获得靶基因（PAI-1）及管家基因（β-actin）的PCR产物。纯化后,作为标准品梯度稀释,采用SYBR Green I定量PCR检测,建立标准曲线。方法学考核参数为特异性、线性范围、灵敏性和重复性。分析全反式维甲酸对内皮细胞表达PAI-1 mRNA的干预效果。这种定量方法特异性好,检测的灵敏度达10^3拷贝,线性范围为10^4-10^10拷贝,循环阈值与PCR体系中起始模板量的对数值之间有着良好的线性关系（r^2〉0.990）,批内变异≤4.93%,批间变异≤9.12%。全反式维甲酸能上调内皮细胞PAI-1 mRNA的表达,且呈剂量依赖性。因此,荧光定量RT-PCR检测血管内皮细胞PAI-1基因表达的方法有助于溶栓药物药理学的研究和新药筛选。     

鸡减蛋综合征病毒(EDSV)基因组右末端结构的分析

从中国发病鸡中分离的鸡减蛋综合征病毒（ＥｇｇＤｒｏｐＳｙｎｄｒｏｍＶｉｒｕｓ,ＥＤＳＶ）弱毒株（ＡＡ－２）,其基因组全长约为３３ｋｂ。用限制性内切酶ＨｉｎｄⅢ水解ＥＤＳＶ全基因组,构建了以ｐＢｌｕｅｓｃｒｉｐｔⅡ（ＫＳ＋）为载体的右末端片段的克隆（约４．２ｋｂ）,对其进行了序列测定和结构分析。该片段全长４１８３个碱基对（ｂｐ）,位于基因组右末端８７．３ｍ．ｕ．－１００ｍ．ｕ．。结果显示,该片段与哺乳动物腺病毒右末端Ｅ４区结构不同,与禽Ⅰ型腺病毒代表株ＣＥＬＯ右末端片段亦无同源性。本文为深入了解ＥＤＳＶ基因组结构特点,ＥＤＳＶ与其他腺病毒基因结构与功能的进化关系和ＥＤＳＶ载体的构建奠定了分子生物学基础     

Preparation of Mesoporous Nano-Hydroxyapatite Using a Surfactant Template Method for Protein Delivery

Mesoporous nano-hydroxyapatite (n-HA) has gained more and more attention as drug storage and release hosts.The aim of this study is to observe the effect of the ratio of surfactant to the theoretical yield of HA on the mesoporous n-HA,then to reveal the effect of the mesoporous nanostrueture on protein delivery.The mesoporous n-HA was synthesized using the wet precipitation in the presence of cetyltrimethylammonium bromide (CTAB) at ambient temperature and normal atmospheric pressure.The morphology,size,crystalline phase,chemical composition and textural characteristics of the product were well characterized by X-ray Powder Diffraction (XRD),Fourier Transform Infrared Spectroscopy (FTIR),Scanning Electron Microscopy (SEM),Transmission Electron Microscopy (TEM),Dynamic Light Scattering (DLS) and N2 adsorption/desorption,respectively.The protein adsorption/release studies were also carried out by using Bovine Serum Albumin (BSA) as a model protein.The results reveal that the mesoporous n-HA synthesized with CTAB exhibits high pure phase,low crystallinity and the typical characteristics of the mesostructure.The BSA loading increases with the specific surface area and the pore volume of n-HA,and the release rates of BSA are different due to their different pore sizes and pore structures,n-HA synthesized with 0.5％ CTAB has the highest BSA loading and the slowest release rate because of its highest surface area and smaller pore size.These mesoporous n-HA materials demonstrate a potential application in the field of protein delivery due to their bioaetive,biocompatible and mesoporous properties.     

重寄生真菌盾壳霉pH信号通路中Pal相关基因的功能鉴定

【目的】研究pH信号通路(Pal)在重寄生真菌盾壳霉与寄主核盘菌互作过程中的作用。【方法】从盾壳霉全基因组信息中分析获得了6个Pal相关基因CmpalA、CmpalB、CmpalC、CmpalF、CmpalH和CmpalI的全编码序列和氨基酸序列,通过PEG介导的原生质转化技术获得了CmpalA、CmpalB、CmpalC、CmpalF和CmpalH等5个基因的敲除突变体,分析这些敲除突变体与野生型在菌落培养性状、重寄生能力、降解草酸能力、产生抗真菌物质能力等方面的差异。【结果】与野生型相比,在pH 6–8的条件下,5个Pal相关基因敲除突变体的菌丝生长受到显著抑制,这说明缺失Pal相关基因使盾壳霉对高pH值环境更加敏感。菌核重寄生试验发现5个Pal相关基因敲除突变体的重寄生能力均显著低于野生型。qRT-PCR试验结果表明,敲除Pal相关基因之后导致重寄生相关酶基因Cmch1、Cmg1和Cmsp1的表达量显著降低,而且pH信号通路下游的CmpacC基因的表达量也显著降低。Pal相关基因敲除突变体在pH 6条件下对草酸盐的降解能力显著高于野生型,同时这5个突变体在pH 8条件下产生抗真菌物质能力也显著高于野生型。【结论】pH信号通路相关基因的缺失影响盾壳霉对环境pH的响应。pH信号通路在盾壳霉与核盘菌互作中发挥重要作用,不仅影响盾壳霉的重寄生作用,而且还影响盾壳霉的草酸降解作用和抗真菌作用。     

Novel Biomarkers for Non-functioning Invasive Pituitary Adenomas were Identified by Using Analysis of microRNAs Expression Profile

The microRNAs (miRNAs) are involved in multiple pathological processes among various types of tumors. However, the functions of miRNAs in benign brain tumors are largely unexplored. In order to explore the pathogenesis of the invasiveness in non-functional pituitary adenoma (NFPA), the miRNAs expression profile was analyzed between invasive and non-invasive non-functional pituitary adenoma by miRNAs microarray. Six most significant differentially expressed miRNAs were identified including four upregulated miRNAs hsa-miR-181b-5p, hsa-miR-181d, hsa-miR-191-3p, and hsa-miR-598 and two downregulated miRNAs hsa-miR-3676-5p and hsa-miR-383. The functions and corresponding signaling pathways of differentially expressed miRNAs were investigated by bioinformatics techniques, including Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis. The result of GO analysis indicates regulation of voltage-gated potassium channel activity, positive regulation of sodium ion transport, positive regulation of GTPase activity, negative regulation of Notch signaling pathway, etc. KEGG pathway reveals a series of biological processes, including prolactin signaling pathway, endocrine and other factor-regulated calcium reabsorption, fatty acid metabolism, neuroactive ligand-receptor interaction, etc. The miRNAs hsa-miR-181a-5p was verified by quantitative real-time PCR, and the expression level was in accordance with the microarray result. Our result can provide the evidence on featured miRNAs which play a prominent role in pituitary adenoma as effective biomarkers and therapeutic targets in the future.     

中华蟾蜍消化道6种酶的组织化学定位

目的研究中华蟾蜍消化道酸性磷酸酶(ACP)、碱性磷酸酶(ALP)、腺苷三磷酸酶(ATPase)、非特异性酯酶(NSE)、过氧化物酶(POX)和琥珀酸脱氢酶(SDH)等6种酶的分布。方法在消化道的8个部位取材,采用冰冻切片技术、石蜡切片技术、酶的组织化学方法和光密度定量分析。结果 ACP主要分布于胃贲门中贲门腺部,十二指肠和回肠中酶反应呈弱阳性。ALP主要分布于食管、十二指肠至回肠的粘膜上皮,十二指肠酶活性最高。ATPase在消化道各部位均有分布,胃中胃腺部和回肠粘膜上皮酶活性显著较高(P0.05)。NSE和POX在整个消化道粘膜上皮和粘膜固有层均有分布,胃各部位酶活性显著较低(P0.05)。SDH除在食管和直肠酶活性显著较低外,其它部位均有大量分布,十二指肠和回肠酶活性显著较高(P0.05)。结论中华蟾蜍消化道粘膜6种酶的分布同其它动物有相似之处,也有其自身特点。6种酶在消化道中的分布与消化道各部位的生理机能密切相关。     

瘦素诱导体外培养大鼠脂肪间充质干细胞凋亡

为观察瘦素诱导体外培养大鼠脂肪间充质干细胞凋亡的作用, 采用胶原酶消化法分离培养大鼠附睾脂肪垫间充质干细胞, 第3代细胞用于实验。细胞免疫荧光化学方法鉴定CD105、Vimentin表达阳性率约80%以上, 10-6 mol/L的瘦素作用细胞48 h、72 h后激光共聚焦显微镜观察分别可见早期及中晚期特征表现; 0 mol/L、10-8 mol/L、10-7 mol/L、10-6 mol/L瘦素分别作用于细胞48 h后, 应用AnnexinⅤ/PI双染色法流式细胞仪检测早期凋亡率分别为2.50%±0.72%、6.78%±1.99%、11.99%±1.58%、17.93%±4.82% (P<0.05); 随着瘦素浓度的增加和作用时间的延长, Caspase-3的活性逐渐增高, 至48 h时达到高峰。说明瘦素可以直接诱导脂肪间充质干细胞凋亡, 从数量上减少脂肪组织的含量。