Activity and QSAR study of baogongteng A and its derivatives as muscarinic agonists

Baogongteng A (BGT-A), a naturally occurring tropane muscarinic agonist isolated from Chinese medicinal plant, exhibits a bioactive effect different from those of many tropane alkaloids that are muscarinic antagonists. A series of racemic derivatives of BGT-A was synthesized to study the structure-activity relationships (SAR). To explore further the SAR in this series and to ultimately design muscarinic agonists for drug development, a Comparative Molecular Field Analysis (CoMFA) was performed. The values of the leave-one-out cross-validated correlation coefficient q2 and the conventional correlation coefficient r2 for the model are 0.613 and 0.965, respectively. The regression analysis of the data indicated that the steric effect of N-substituted group on tropane of analyzed compounds critically affected the agonistic activity to muscarinic receptors.     

Heterogeneity in premature senescence by oxidative stress correlates with differential DNA damage during the cell cycle

The development of cellular senescence both by replication and by oxidative stress is not homogenous in cultured primary human fibroblasts. To investigate whether this is due to the heterogeneity in the susceptibility of DNA in different phases of the cell cycle, we subjected synchronised cells to oxidative stress and examined the extent of DNA damage and its long-term effects on the induction of cellular senescence. Here, we first show marked heterogeneity in DNA damage as detected by markers of double strand breaks caused by oxidative stress in an asynchronous human fibroblast culture. Cell cycle synchronization followed by oxidative stress demonstrated that DNA in S-phase is most susceptible to oxidative stress whereas DNA in the quiescent phase is most resistant. DNA repair is an ongoing process after sensing DNA damage; reparable DNA damage is repaired even in cells that contain persistent DNA damage. The extent of persistent DNA damage is tightly correlated with permanent cessation of DNA replication and SA-beta-gal activity. Oxidative stress encountered by cells in S-phase resulted in more persistent DNA damage, more permanent cell cycle arrest and the induction of premature senescence.     

Structural Evidence for α-Synuclein Fibrils Using in Situ Atomic Force Microscopy

Human α-synuclein is a presynaptic terminal protein and can form insoluble fibrils that are believed to play an important role in the pathogenesis of several neurodegenerative diseases such as Parkinson‘s disease, dementia with Lewy bodies and Lewy body variant of Alzheimer‘s disease. In this paper, in situ atomic force microscopy has been used to study the structural properties of α-synuclein fibrils in solution using two different atomic force microscopy imaging modes: tapping mode and contact mode. In the in situ contact mode atomic force microscopy experiments α-synuclein fibrils quickly broke into fragments, and a similar phenomenon was found using tapping mode atomic force microscopy in which α-synuclein fibrils were incubated with guanidine hydrochloride (0.6 M). The α-synuclein fibrils kept their original filamentous topography for over 1h in the in situ tapping mode atomic force microscopy experiments. The present results provide indirect evidence on how 13-sheets assemble into α-synuclein fibrils on a nanometer scale.     

1型糖尿病葡萄糖代谢与胰岛素信号传导途径异常导致大鼠海马Alzheimer样tau蛋白过度磷酸化修饰

Tau蛋白过度磷酸化是Alzheimer病(Alzheimer disease, AD)的一个重要病理特征.采用 I 型糖尿病大鼠模型,研究胰岛素信号传导途径及葡萄糖代谢失调对tau蛋白过度磷酸化的形成机制进行探讨.以同龄Wistar大鼠做对照（CTL）,胰腺大部分切除造低胰岛素组（PX）,STZ较大剂量一次性注射造1型糖尿病模型即低胰岛素高血糖组（T1DM）.葡萄糖氧化酶法检测血浆血糖,放免法检测血浆胰岛素,蛋白质印迹分析海马内总tau蛋白及tau蛋白上部分位点（Ser199、Thr212、Ser214、Ser396及Ser422）的磷酸化及神经细胞膜上葡萄糖转运子3（Glucose transport 3,GLUT3）水平.γ-32P-ATP和特异性底物肽检测海马内胰岛素信号传导系统中的关键酶糖原合成酶激酶-3β（Glycogen synthase kinase-3β, GSK-3β）活性.发现3组大鼠海马回总tau蛋白水平无显著差异,但以高血糖、低胰岛素血症为特征的T1DM组在tau蛋白Ser199、Thr212、Ser214、Ser396及Ser422位点上,呈现过度磷酸化状态,以低胰岛素血症为特征而血糖正常的PX组在位点Ser199、Thr212及Ser396上磷酸化程度比CTL组显著上升, 在位点Ser214及 Ser422上的磷酸化程度的改变不显著;T1DM及PX组大鼠海马 GSK-3β活性显著高于CTL组, 而GLUT3水平在T1DM和PX组均降低, 尤以T1DM组降低更显著.研究结果显示,胰岛素水平低下可能通过激活GSK-3β和下调细胞内葡萄糖代谢的双重作用引起脑内tau蛋白过度磷酸化.     

昆虫先天性免疫信号通路研究进展

昆虫体内形成了强大的免疫防御系统,其被各种微生物攻击时能依靠病原相关分子模式识别蛋白对感染进行区分和激活体内信号通路诱导如抗菌肽之类的效应分子.昆虫体内控制先天性免疫的信号通路分别是:Toll通路、IMD通路和JAS/STAT通路,这3条通路在信号传递过程中存在协作,并且,这些通路与脊椎动物体内某些通路存在惊人相似、在免疫调控通路方面存在共同的进化起源.这揭示了先天性免疫在动物体内存在的普遍性和机体抵御病原感染的重要性.     

125I和65Zn在大蟾蜍体内的吸收与分布

利用同位素示踪法研究了不同引入途径的125I和65Zn在大蟾蜍体内的吸收与分布。无论采用注射还是灌喂的方法,蟾蜍对125I的残留动态差别不大,都可分为快清除期和平台期,0～2d为快清除期,3～7d为平台期,经过了快清除期之后,残留的125I约为起始量的8%,并维持到实验结束。注射了65Zn后,0～7d的吸收动态曲线有所起伏,但波动很小,其波动范围为102.4%～114.48%。表明大部分的65Zn仍留在体内,几乎没有排出体外。注射65Zn的转移并不明显。但灌喂65Zn的动态变化则大些,而且转移明显。其活度曲线出现阶段性下降,有两个下降期,第一个下降期为4h～3d,第二个下降期为5～7d。说明灌喂组对65Zn的转移比注射组的活跃,但在实验的第7d残留率仍在60%以上。     

人IFN-β启动子基因报告质粒的构建及SARS-CoV3a和7a蛋白对IFN-β诱生作用的初步研究

3a蛋白和7a蛋白是SARS-CoV的非结构蛋白,分别主要由SARS基因组中ORF 3a 和ORF 7a所编码。在体内和体外感染病毒的细胞中均发现了有3a蛋白的表达。首先将pGL3-Control载体进行了改构,除去了SV40启动子基因,获得了pGL3-Enhancer载体,将获得的IFN-β启动子基因连入载体中,构建了带有人IFN-β启动子基因的荧光素酶报告质粒IP-21,并且通过实验证明所构建的质粒在干扰素的诱导剂NDV的作用下能够表达荧光素酶活性,照度计检测荧光素酶活性增强,从而验证了所构建的重组质粒的有效性。为了观察SARS-CoV非结构蛋白3a和7a对干扰素诱生途径的影响,将IP-21瞬转入稳定表达有3a和7a蛋白的CHO细胞,通过荧光素酶的信号强弱对3a和7a的作用进行分析,结果表明SARS-CoV的3a和7a蛋白能够刺激荧光素酶的表达,即在体外的细胞实验中,3a和7a能有效地激活IFN-β的启动子部分。此结果对进一步研究3a和7a的功能以及对SARS-CoV的致病机制的进一步研究有一定意义。     

油菜单显性核雄性不育基因的分子标记

以陕西省杂交油菜研究中心选育的单显性核不育油菜分离群体为材料,利用集群分离法（BSA）对该油菜单显性核不育基因进行了RAPD分析。在随机选取的300个10碱基随机引物中,引物S243（5′CTATGCCGAC3′）在可育集团与不育集团间扩增出特异而可重复的1．5kb的多态性片段OPU-031500,而在细胞质雄性不育和其它核不育类型油菜中均未扩增出上述特异性片段,从而确证此RAPD标记OPU-031500。片段是与甘蓝型油菜单显性核不育基因连锁的。将该多态性片段克隆并测序,发现其序列与拟南芥的一段DNA序列高度同源。根据同源序列及测序结果设计两对特异引物（P1／P2和P3／P4）,引物P3／P4在可育系中可扩增到约1．5kb的单一特异片断,而在不育系中无带,从而将RAPD标记转化为稳定可靠的SCAR标记。     

灵香草的组织培养与快速繁殖

1植物名称灵香草（Lysimachia foenum-graecumHance）。2材料类别顶芽。3培养条件芽的生长及分化培养基：（1）MS;（2）MS＋6-BA 1.0 mg·L-（-1）（单位下同）＋NAA 0.5;（3）MS＋6-BA 2.0＋NAA 0.5;（4）MS＋6-BA 3.0＋NAA 1.0;（5）1/2MS;（6）1/2MS＋6-BA 1.0＋NAA 0.5;（7）1/2MS＋6-BA 2.0＋NAA 0.5;（8）1/2MS＋6-BA 3.0＋NAA 1.0。丛生芽诱导培养基：（9）MS＋6-BA 0.5＋NAA 0.2＋1g·L-（-1）活性碳;（10）MS＋6-BA 1.0＋NAA 0.2＋1 g·L-（-1）活性碳;（11）MS＋6-BA 2.0＋NAA 1.0＋1 g·L-（-1）活性碳;     

草酸氧化酶在水稻胚芽鞘衰老中的作用

草酸氧化酶（OxO）催化草酸氧化产生CO2和H2O2,其在植物发育及防御过程中可能具有重要作用。本文以水稻品种‘湘糯1号’（‘Xiangnuo 1’）为材料,对胚芽鞘中的OxO及其生理功能进行了研究。结果表明,胚芽鞘中的H2O2含量在其衰老时增加;OxO活性在浸种后96h时较低,之后也迅速增加,在240h达到最高;而可溶性蛋白、O2-·和草酸含量以及过氧化氢酶（CAT）活性则随着胚芽鞘的衰老迅速降低。由于H2O2能够诱导细胞死亡,推测OxO可能通过降解草酸产生H2O2参与胚芽鞘的衰老。