中国三种常见蒿属植物潜在地理分布及其主导气候因子

裂叶蒿(Artemisia tanacetifolia)、大籽蒿(Artemisia sieversiana)和艾(Artemisia argyi)是我国常见的蒿属(Artemisia)植物,其分布区域遍布全国.本文利用MaxEnt模型预测3种蒿属植物在当前气候条件以及未来两种气候情景下的潜在分布区.采用受试者工作特征...     

蒙古高原岩黄芪属植物的分支分类学研究

以蒙古高原岩黄芪属植物为对象, 应用徐克学的最大同步法, 探讨了蒙古高原岩黄芪属(豆科)植物的系统演化, 并根据分支分类结果对蒙古高原岩黄芪属进行了系统学处理。作者首次将蒙古高原岩黄芪属分为岩黄芪亚属、半灌木岩黄芪亚属(新拟)和无刺岩黄芪组、丛枝岩黄芪组、无茎岩黄芪组、半灌木岩黄芪组等4 个组。本文对蒙古高原岩黄芪组的划分符合苏联植物志(1945)中的观点。     

长爪沙鼠肝细胞体外分离培养体系的建立

目的建立长爪沙鼠原代肝细胞分离培养体系。方法以雄性长爪沙鼠为供体,采用组织消化法和Seglen两步灌流法分离肝细胞,以台盼蓝染色检测细胞得率和活率,过碘酸-希夫氏反应(PAS)鉴定肝细胞,倒置显微镜观察肝细胞形态变化,并使用含有多种细胞因子的培养基维持培养。结果组织消化法和Seglen两步灌流法平均每只长爪沙鼠可分别获得肝细胞(1.33±0.34)×107个、(3.97±1.15)×107个,细胞活率分别为(29.4±6.05)%、(80.3±4.56)%,这两种方法在细胞得率及活率方面存在显著差异。肝细胞内因有大量的糖原颗粒,经PAS染色后被染成红色。结果表明肝细胞在贴壁后72 h内,肝细胞形态发生显著变化。结论采用胶原酶经肝门静脉灌流分离肝细胞是一种高效获得肝细胞的方法。各种细胞因子有利于维持肝细胞在体外的生长分化,长爪沙鼠原代肝细胞分离培养体系的建立将为肝脏相关疾病研究和防治药物的开发提供技术支持。     

构建基于CRISPR/Cas9技术敲除ALOX5的质粒

旨在利用CRISPR/Cas9技术构建敲除花生四烯5-脂氧合酶基因(Arachidonate 5-lipoxygenase gene,ALOX5)的重组质粒。设计合成3对靶向敲除ALOX5第六外显子的sgRNA,将其分别插入到CRISPR/Cas9质粒骨架pX458载体中,转化感受态大肠杆菌DH5α后挑取克隆,通过测序评估重组质粒是否构建成功。将构建好的重组质粒转染293T细胞,在荧光显微镜下观察转染效果,挑取转染成功的细胞,用试剂盒提取转染细胞基因组DNA,PCR扩增含敲除位点的DNA片段,用测序技术获得核苷酸序列,用DNAStar软件分析转染细胞中ALOX5基因敲除情况。测序结果表明2对双链sgRNA寡核苷酸已插入质粒,且序列正确,靶向ALOX5基因的重组质粒pX458-sgRNAs-ALOX5构建成功。其在293T细胞中的转染效率约为50%,用一代测序法未检测到sgRNAs的切割效果。初步表明利用CRISPR/Cas9技术成功构建靶向ALOX5基因的重组质粒pX458-sgRNAs-ALOX5。     

杨树溃疡病生防菌株N6-34抗菌活性物质的分离纯化与结构鉴定

【背景】杨树溃疡病是一种主要由葡萄座腔菌引起的杨树枝干病害,危害严重。前期从杨树中分离到一株内生拮抗细菌N6-34,研究表明该菌株拮抗效果好,对多种植物病原菌均有较强的拮抗作用。【目的】对拮抗细菌N6-34产生的抗菌活性物质进行分离纯化,并鉴定了活性物质组分的结构。【方法】通过硫酸铵盐析、甲醇抽提、分子筛、高效液相色谱等方法分离纯化N6-34菌株的抗菌活性物质,并对其进行结构鉴定。【结果】N6-34菌株发酵液经多步分离纯化,共获得14个组分,其中有13个组分具有抗菌活性,经一级质谱分析,获得了13种抗菌活性组分的分子量;经二级质谱分析,将13种抗菌活性物质鉴定为Fengycin A或Fengycin B的同系物或同分异构体。【结论】从N6-34菌株发酵液中分离获得了13种抗菌成分,为杨树溃疡病的生物防治提供了理论依据。     

Induction of Autophagy Is an Early Response to Gefitinib and a Potential Therapeutic Target in Breast Cancer

Gefitinib (Iressa^®, ZD1839) is a small molecule inhibitor of the epidermal growth factor receptor (EGFR) tyrosine kinase. We report on an early cellular response to gefitinib that involves induction of functional autophagic flux in phenotypically diverse breast cancer cells that were sensitive (BT474 and SKBR3) or insensitive (MCF7-GFPLC3 and JIMT-1) to gefitinib. Our data show that elevation of autophagy in gefitinib-treated breast cancer cells correlated with downregulation of AKT and ERK1/2 signaling early in the course of treatment. Inhibition of autophagosome formation by BECLIN-1 or ATG7 siRNA in combination with gefitinib reduced the abundance of autophagic organelles and sensitized SKBR3 but not MCF7-GFPLC3 cells to cell death. However, inhibition of the late stage of gefitinib-induced autophagy with hydroxychloroquine (HCQ) or bafilomycin A1 significantly increased (p<0.05) cell death in gefitinib-sensitive SKBR3 and BT474 cells, as well as in gefitinib-insensitive JIMT-1 and MCF7-GFPLC3 cells, relative to the effects observed with the respective single agents. Treatment with the combination of gefitinib and HCQ was more effective (p<0.05) in delaying tumor growth than either monotherapy (p>0.05), when compared to vehicle-treated controls. Our results also show that elevated autophagosome content following short-term treatment with gefitinib is a reversible response that ceases upon removal of the drug. In aggregate, these data demonstrate that elevated autophagic flux is an early response to gefitinib and that targeting EGFR and autophagy should be considered when developing new therapeutic strategies for EGFR expressing breast cancers.     

A mitochondrial-associated link between an effector caspase and autophagic flux

It has become evident that caspases function in nonapoptotic cellular processes in addition to the canonical role for caspases in apoptotic cell death. We recently demonstrated that the Drosophila effector caspase Dcp-1 localizes to the mitochondria and positively regulates starvation-induced autophagic flux during mid-oogenesis. Loss of Dcp-1 leads to elongation of the mitochondrial network, increased levels of the adenine nucleotide translocase sesB, increased ATP levels, and a reduction in autophagy. We found that sesB is a negative regulator of autophagic flux, and Dcp-1 interacts with sesB in a nonproteolytic manner to regulate its stability, uncovering a novel mechanism of mitochondrial associated, caspase-mediated regulation of autophagy in vivo.     

Study of tetraphenylporphyrins as modifiers of insulin amyloid aggregation

Amyloid fibrils are rigid β‐pleated protein aggregates that are connected with series of harmful diseases and at the same time are promising as base for novel nanomaterials. Thus, design of compounds able to inhibit or redirect those aggregates formation is important both for the biomedical aims and for nanotechnology applications. Here, we studied the effect of tetraphenylporphyrins (metal free, their Cu and Pd complexes, and those functionalized by carboxy and amino groups on periphery) on insulin amyloid self‐assembling. The strongest impact on insulin aggregation was demonstrated by a metal‐free porphyrin bearing four carboxy groups. This compound strongly suppresses insulin aggregation (about 88% reduction in amyloid‐sensitive probe emission) inducing formation of fibrils with the length close to this of free insulin (1.7 ± 0.6 μm as compared with 1.4 ± 0.4 μm, respectively) with an essentially reduced tendency to lateral aggregation. Contrarily, the presence of tetraphenylporphyrin containing four amino groups only slightly affects fibrils' morphology and makes weaker impact on insulin aggregation yield (about 44% reduction). This is explained by the ability of aromatic carboxy groups of 5,10,15,20‐(tetra‐4‐carboxyphenyl)porphyrin to interact with complementary protein‐binding groups and thus stabilize the supramolecular complex. For 5,10,15,20‐(tetra‐4‐aminophenyl)porphyrin, full protonation takes place in acidic medium of protein aggregation reaction; this results in the high positive charge of TPPN4 (equal or close to +6) and hence higher contribution of coulombic repulsion to interaction of TPPN4 with insulin. One more possible mechanism of the lower inhibition effect of TPPN4 as compared with TPPC4 could be the more restricted possibility of the former as compared with the latter to form H bonds with insulin groups. It was also shown that metal‐free, Pd‐containing, and Cu‐containing tetraphenylporphyrins without peripheral substituents make almost the same impact on the protein self‐assembling. We suppose this to be due to coordination saturation of these metal atoms.