降低PARP酶活性对培养细胞SCE及微核效应的影响

聚腺苷二磷酸核糖基化作用是细胞内的一种重要的核蛋白转译后加工修饰，它参与细胞内很多重要的生物事件．催化该反应的酶是聚腺苷二磷酸核糖合酶（ＰＡＲＰ），底物为ＮＡＤ．本文使用ＰＡＲＰ酶的抑制剂苯甲酰胺处理培养细胞，研究了降低ＰＡＲＰ酶活性对培养细胞姐妹染色单体交换及微核效应的影响，为全面评价ＰＡＲＰ酶抑制剂在细胞内的功效打下基础．     

降低PARP酶活性对培养细胞SCE及微核效应的影响

聚腺苷二磷酸核糖基化作用是细胞内的一种重要的核蛋白转译后加工修饰，它参与细胞内很多重要的生物事件。催化该反应的酶是聚腺苷二磷酸核糖合酶（ＰＡＲＰ），底物为ＮＡＤ。本文使用ＰＡＲＰ酶的抑制剂苯甲酰胺处理培养细胞，研究了降低ＰＡＲＰ酶活性对培养细胞姐妹染色单体交换及微核效应的影响，为全面评价ＰＡＲＰ酶抑制剂在细胞内的功效打下基础。     

环腺苷二磷酸核糖（cADPR）类似物的合成与诱导钙释放活性的研究

Ca²⁺信号传导通路是生物体内重要的胞内信号传导途径之一。局部钙信号主要来源于细胞内钙库释放,而这些钙信号受到各种第二信使的控制和Ca²⁺通道蛋白的调节。环腺苷二磷酸核糖（cADPR）作为烟酰胺腺嘌呤二核苷酸（NAD⁺）的代谢物,发现于1987年,是一种信号传导分子,它广泛存在于各种生物系统中,通过介导兰诺定( RyR) 受体调节钙动员活性。研究cADPR以及具有不同生物活性的类似物之间的构效关系是探究分子内钙释放机制的主要手段,另外,一些结构新颖的拮抗剂和激动剂可以作为研究细胞系统复杂机制的研究工具。作者概括性地介绍了cADPR结构类似物——N¹-乙氧基甲基-环肌苷-5'-二磷酸核糖（cIDPRE）和N¹-[(磷酰基-O-乙氧基)-甲基-N⁹-[(磷酰基-O-乙氧基)-甲基-次黄嘌呤-环磷酸焦酯（cIDPDE）的合成与性质。这两种类似物cIDPDE和cIDPRE可作为研究完整细胞钙信号系统的膜透性激动剂。     

PARP1调节肝癌细胞的生长和迁移

为了探究聚腺苷二磷酸-核糖聚合酶1(PARP1)在肝细胞癌发生发展中的功能,通过免疫组化检测PARP1在肝癌组织中的表达水平;通过CRISPR/CAS9技术构建PARP1缺陷的肝癌细胞株,并采用CCK8实验、克隆形成法和细胞迁移实验来探究PARP1缺陷对肝癌细胞的影响.免疫组化结果显示PARP1在肝癌细胞中高表达,PARP1缺陷抑制肝癌细胞生长和迁移.     

PARP介导氧化应激与糖尿病视网膜病变的研究进展

糖尿病视网膜病变(DR)因其发病率高、病情严重而倍受人们关注,氧化应激能通过各种途径损伤糖尿病的视网膜.聚腺苷二磷酸核糖聚合酶(PARP)是目前国外研究较热的一种酶,并有较多实验证据表明PARP能介导氧化应激并参与糖尿病视网膜疾病的发生.该文就PARP在DR发病中的作用做一综述.     

FDP改善运动疲劳红细胞能量代谢

探索1,6-二磷酸果糖（FDP）在分子水平对蹦床运动员疲劳后红细胞（RBC）能量代谢的影响规律,为运动性疲劳的恢复提供实验研究依据。FDP800mmol／L和0mmol／L分别孵育疲劳后红细胞,分别用细胞化学方法、比色法、高效液相色谱法检测细胞色素氧化酶活力,2,3-二磷酸甘油酸（2,3-DPG）水平,三磷酸腺苷（ATP）、二磷酸腺苷（ADP）、一磷酸腺苷（AMP）含量并计算能荷。FDP处理后红细胞ATP、能荷、细胞色素氧化酶活力和2,3-DPG水平显著增加,ADP和AMP含量显著下降。FDP能够通过增加细胞色素氧化酶活力,提高2,3-DPG水平来改善疲劳红细胞能量供应,缓解运动性疲劳。     

胶质细胞在吗啡耐受中的作用

随着对神经系统研究的不断深入，胶质细胞越来越受到人们的重视。本文就胶质细胞在吗啡中的作用作一综述，主要包括胶质细胞的激活，细胞内的信号转导及分泌的细胞因子三个方面。     

细胞代谢调控网络

代谢是细胞及机体获取营养物质及能量的重要生命过程,其稳态平衡是机体应对内外环境变化进行正常生命活动的根本保障.代谢稳态平衡的破坏会导致各种疾病.细胞代谢稳态的维持有赖于机体和细胞之间及细胞内部多层次的调节网络.本文总结了内分泌系统部分激素调控细胞代谢及细胞内各代谢通路之间相互调节共同维持代谢稳态的机制,重点阐述了单磷酸腺苷激活的蛋白激酶(adenosine monophosphate-activated protein kinase, AMPK)及雷帕霉素机制性靶蛋白(mechanistic target of rapamycin, mTOR)信号通路对合成代谢及分解代谢的综合调控.     

神经病理性疼痛的小胶质细胞机制

小胶质细胞是中枢神经系统内重要的免疫细胞,起递呈抗原、吞噬病原体、分泌多种细胞因子和神经修复的作用。神经损伤后,小胶质细胞会被激活,可释放大量的细胞因子、炎性介质,激活补体,引起神经炎症和神经免疫反应,导致各种神经功能紊乱,引起痛觉过敏和异常痛敏。本文从小胶质细胞的激活、以及激活途径的关键分子TLR4来探讨神经病理性疼痛的发病机制。     

Exfoliazone通过诱导细胞凋亡抑制HepG2细胞体外增殖

探讨Exfoliazone对肝癌细胞HepG2增殖及凋亡的影响.采用细胞培养技术,用不同浓度的药物在一定的时间内处理细胞株,应用MTT比色法检测Exfoliazone对HepG2细胞体外生长的抑制作用;4′,6-二脒基-2-苯基吲哚(DAPI)荧光染料染色,显示凋亡细胞形态;用Western-Blotting检测聚腺苷二磷酸核糖聚合酶(PARP)蛋白的表达;Annexin V/PI双染检测Exfoliazone对HepG2细胞凋亡的影响.Exfoliazone可抑制HepG2细胞的生长,并呈现剂量和时间的抑制,在药物处理24、48和72 h后,半数细胞抑制浓度(IC50)分别为35、16 和8 μmol/L.进一步研究表明,Exfoliazone可诱导PARP切割,细胞出现核质浓集和凋亡小体等典型的凋亡形态学特征,Annexin V/PI染色证实Exfoliazone能诱导HepG2细胞凋亡,并且以早期凋亡为主.Exfoliazone可能通过诱导HepG2细胞凋亡抑制癌细胞的生长.     

Mesenchymal stem cells within tumour stroma promote breast cancer metastasis

Mesenchymal stem cells have been recently described to localize to breast carcinomas, where they integrate into the tumour-associated stroma. However, the involvement of mesenchymal stem cells (or their derivatives) in tumour pathophysiology has not been addressed. Here, we demonstrate that bone-marrow-derived human mesenchymal stem cells, when mixed with otherwise weakly metastatic human breast carcinoma cells, cause the cancer cells to increase their metastatic potency greatly when this cell mixture is introduced into a subcutaneous site and allowed to form a tumour xenograft. The breast cancer cells stimulate de novo secretion of the chemokine CCL5 (also called RANTES) from mesenchymal stem cells, which then acts in a paracrine fashion on the cancer cells to enhance their motility, invasion and metastasis. This enhanced metastatic ability is reversible and is dependent on CCL5 signalling through the chemokine receptor CCR5. Collectively, these data demonstrate that the tumour microenvironment facilitates metastatic spread by eliciting reversible changes in the phenotype of cancer cells.     

Abscisic acid controls calcium-dependent egress and development in Toxoplasma gondii

Calcium controls a number of critical events, including motility, secretion, cell invasion and egress by apicomplexan parasites. Compared to animal and plant cells, the molecular mechanisms that govern calcium signalling in parasites are poorly understood. Here we show that the production of the phytohormone abscisic acid (ABA) controls calcium signalling within the apicomplexan parasite Toxoplasma gondii, an opportunistic human pathogen. In plants, ABA controls a number of important events, including environmental stress responses, embryo development and seed dormancy. ABA induces production of the second-messenger cyclic ADP ribose (cADPR), which controls release of intracellular calcium stores in plants. cADPR also controls intracellular calcium release in the protozoan parasite T. gondii; however, previous studies have not revealed the molecular basis of this pathway. We found that addition of exogenous ABA induced formation of cADPR in T. gondii, stimulated calcium-dependent protein secretion, and induced parasite egress from the infected host cell in a density-dependent manner. Production of endogenous ABA within the parasite was confirmed by purification (using high-performance liquid chromatography) and analysis (by gas chromatography-mass spectrometry). Selective disruption of ABA synthesis by the inhibitor fluridone delayed egress and induced development of the slow-growing, dormant cyst stage of the parasite. Thus, ABA-mediated calcium signalling controls the decision between lytic and chronic stage growth, a developmental switch that is central in pathogenesis and transmission. The pathway for ABA production was probably acquired with an algal endosymbiont that was retained as a non-photosynthetic plastid known as the apicoplast. The plant-like nature of this pathway may be exploited therapeutically, as shown by the ability of a specific inhibitor of ABA synthesis to prevent toxoplasmosis in the mouse model.     

酵母双杂交筛选与脱落酸受体相互作用的蛋白质及其重转酵母的验证

脱落酸（abscisic acid,ABA）是一种重要的植物激素,它能使植物适应逆境胁迫,从而调节植物的生长发育状况.现今ABA受体已被公认为RCARs/PYR/PYLs家族蛋白.本实验通过拟南芥为模式植物,利用酵母双杂交系统共转化方法从拟南芥cDNA文库中筛选与RCAR3相互作用的蛋白质,获得多个阳性克隆,从中选取一个与生长素调节相关的基因NIT1的全长cDNA重新转化酵母验证,发现与RCAR3仍然有相互作用,排除了假阳性的可能.从而为下一步研究该基因与RCAR3的相互关系做好了准备.     

蓝光调节气孔运动的信号途径及其与脱落酸相互关系研究

气孔运动与植物水分代谢密切相关.保卫细胞可有效感知和整合多种环境信号,通过控制离子进出调节其膨压,影响气孔开与闭.诸多研究表明,蓝光信号诱导气孔开放和逆境信号脱落酸(ABA)促进气孔关闭构成了气孔运动的两大研究领域.该文就保卫细胞中蓝光信号传递及与ABA信号交叉控制气孔开闭的研究进展进行综述,以了解气孔对蓝光和ABA反应的最新进展,为发展耐旱与提高作物水分利用效率生物技术的改进提供理论支持.     

Functional analysis of the ABA-responsive protein family in ABA and stress signal transduction in Arabidopsis

The phytohormone abscisic acid (ABA) plays an important role in plant growth and development, for example in seed dormancy and germination, as well as in plant responses to environmental stresses, such as drought and high salinity. Previous studies have shown that ABA regulates the expression of genes with an ABA-responsive element (ABRE) and their corresponding physiological responses. Bioinformatics analysis identified a GRAM domain-containing gene family that has a multiple ABRE cis-element, which was termed the ABA-responsive protein (ABR) family. To analyze the function of the ABR family, we identified homozygous T-DNA insertion mutants and constructed abr1, 2, 3 double mutants and triple mutant. The abr1, abr2 and abr3 single mutants showed a normal phenotype; however, the germination of seeds of the double mutants and triple mutant were insensitive to ABA, NaCl, mannitol and glucose. ABR1-GFP was distributed as a punctate structure in the cytosol and may be localized in the endomembrane system. The ABR2-GFP and ABR3-GFP proteins localized in the cytoplasm. In addition, ABR1, ABR2 and ABR3 were expressed in various tissues, and could be induced by several abiotic stresses, especially by ABA. The expressions of these genes were significantly suppressed in aba2, abi1 and abi2 null mutants. These results suggested that the ABR family may act downstream of ABI1 and ABI2 in the ABA signal transduction process in plants.     

Roles of a sustained activation of NCED3 and the synergistic regulation of ABA biosynthesis and catabolism in ABA signal production in Arabidopsis

ABA, acting as a stress signal, plays crucial roles in plant resistance to water stress. Because ABA signal production is based on ABA biosynthesis, the regulation of NCED, a key enzyme in the ABA biosynthesis pathway, is normally thought of as the sole factor controlling ABA signal production. Here we demonstrate that ABA catabolism in combination with a synergistic regulation of ABA biosynthesis plays a crucial role in governing ABA signal production. Water stress induced a significant accumulation of ABA, which exhibited different patterns in detached and attached leaves. ABA catabolism followed a temporal trend of exponential decay for both basic and stress ABA, and there was little difference in the catabolic half-lives of basic ABA and stress ABA. Thus, the absolute rate of ABA catabolism, i.e. the amount of ABA catabolized per unit time, increases with increased ABA accumulation. From the dynamic processes of ABA biosynthesis and catabolism, it can be inferred that stress ABA accumulation may be governed by a synergistic regulation of all the steps in the ABA biosynthesis pathway. Moreover, to maintain an elevated level of stress ABA sustained activation of NCED3 should be required. This inference was supported by further findings that the genes encoding major enzymes in the ABA biosynthesis pathway, e.g. NCED3, AAO3 and ABA3 were all activated by water stress, and with ABA accumulation progressing, the expressions of NCED3, AAO3 and ABA3 remained activated. Data on ABA catabolism and gene expression jointly indicate that ABA signal production is controlled by a sustained activation of NCED3 and the synergistic regulation of ABA biosynthesis and catabolism.     

拟南芥ABA受体与UGT71B6的相互作用研究

为进一步分析ABA尿苷二磷酸葡萄糖基转移酶UGT71B6参与植物抗逆反应的具体机制,本研究通过体外酵母双杂交实验与GST-Pull down实验、体内双分子荧光互补实验(BIFC)分析了ABA受体RCARs与UGT71B6之间的相互作用关系.实验结果显示ABA受体RCAR12、RCAR13与UGT71B6,在体内、体外均存在直接的相互作用.研究表明UGT71B6的生理功能与ABA受体之间存在重要联系, UGT71B6可能通过ABA受体的介导参与了植物逆境应答反应.实验结果也为初步探究ABA受体是否参与ABA稳态调节过程提供一定参考.     

2种非生物胁迫和7种激素对水稻OsAQP基因表达的影响

OsAQP是从cDNA文库中分离的一种新的水稻水通道蛋白基因,本实验室前期利用半定量RT-PCR技术发现该基因的表达与非生物胁迫、植物激素有关.本文采用qRT-PCR方法,检测了干旱、高盐胁迫以及ABA等7种植物激素对OsAQP在幼苗期水稻根和叶中表达的影响,结果显示干旱、高盐以及7种植物激素MeJA,SA,6-BA,GA,IAA,ABA,BR均能不同程度诱导OsAQP基因上调表达,在根中的诱导上调水平显著高于叶,提示该基因功能涉及生长发育、抗逆应答等多种生物学过程,且与水稻的根的关系更为密切.     

梅花鹿鹿茸软骨细胞和间充质干细胞永生化细胞株的构建及鉴定

分离梅花鹿鹿茸软骨细胞和间充质干细胞,使用SV40 LT抗原慢病毒载体建立永生化软骨细胞和间充质干细胞系并鉴定。将含有SV40 LT基因片段的慢病毒载体,转染人胚肾细胞293T获得包装后的病毒粒子,感染鹿茸软骨细胞及间充质干细胞,连续传代培养,通过形态观察、细胞增殖、real-time PCR、甲苯胺蓝染色法和流式细胞术等方法检测SV40 LT抗原表达以及细胞性质。实验所建立的永生化鹿茸软骨细胞系与间充质干细胞系能够稳定传代并具有较强的体外增值活性。RT-PCR检测到SV40T抗原的表达,通过甲苯胺蓝染色法和流式细胞术鉴定所得细胞系具有原代细胞的基本性质。成功获得永生化的软骨细胞与间充质干细胞系,为后续鹿茸生长及发育研究奠定了基础。