瓦山安息香中苯并呋喃促雌激素合成的机理研究

雌激素是人体内重要的激素之一，具有广泛的生理功能。雌激素缺乏与许多疾病相关，如卵巢功能低下，更年期综合征以及骨质疏松等；雌激素过剩也将导致某些疾病，如乳腺癌、卵巢癌、子宫内膜癌等。目前，如何降低肿瘤组织中的雌激素水平而达到治疗肿瘤的目的，已经得到广泛的研究，但促雌激素生成或调节卵巢功能药物或其相关研究则很少。 本实验室前期的研究发现，瓦山安息香属植物果实中的乙醇提取物具有促雌激素生成作用，通过活性追踪和结构鉴定，确认促E2 生成的主要成分为苯并呋喃类化合物。苯并呋喃类化合物的作用与芳香酶有关，但其确切的作用机理有待证实和深入研究。 为了探讨安息香苯并呋喃类化合物的促雌激素合成的作用机理，拟采用如下的实验方案： 1、细胞学方面，对小鼠3T3-L1 前脂肪细胞、人乳腺癌细胞MCF-7、MDA-MB-231 以及人卵巢癌细胞OVCAR-3、OVCAR-4、OVCAR-5、OVCAR-8、IGROV1 等细胞株，采用RT-PCR 和ELISA 方法研究芳香酶Aro基因的表达和雌二醇E2 的生成，芳香酶抑制剂Formestane 作为阳性对照，研究时效曲线和量效曲线，确定安息香苯并呋喃类化合物SP25 的有效浓度和作用时间。 2、RNAi 方面，设计合成了针对人芳香酶Aro基因的3 对RNAi 序列，转染入细胞，芳香酶促进剂Forskolin 和地塞米松、芳香酶抑制剂Formestane 作为阳性对照，采用实时定量PCR 技术，研究RNA 干扰后，安息香苯并呋喃类化合物SP25 对人芳香酶Aro基因表达水平瓦山安息香苯并呋喃促雌激素合成的机理研究的影响。 3、雌激素受体方面，设计一段ERE 的雌激素调控元件，构建重组荧光素酶报告基因载体，瞬时转染人乳腺癌细胞株MDA-MB-231，建立针对雌激素受体的报告基因筛选模型，观察安息香苯并呋喃类化合物SP25 对雌激素受体的选择性和亲和力，从受体水平考察安息香苯并呋喃类化合物SP25 促进雌激素生成的药理学机理。 实验结果显示： 1、分化后的小鼠3T3-L1 前脂肪细胞、人乳腺癌细胞MCF-7 、MDA-MB-231 以及人卵巢癌细胞OVCAR-3、OVCAR-4、OVCAR-8 等细胞株具有芳香酶基因的表达。睾酮向雌二醇的转化能够被芳香酶抑制剂Formestane 所阻断，其中OVCAR-3 最适合进行下一步的RNAi研究。 2、RNAi 实验结果显示，设计的3 对RNAi 序列中R2 的干扰效果最强，相应的阴性对照C2 与R2 的表达量相差118 倍（24 小时）和19 倍（48 小时），显示R2/C2 这组序列可用于进一步的RNAi 试验。以R2 干扰OVCAR-3 细胞株，药物作用24、48 小时后，芳香酶抑制剂Formestane 与R2 相对表达量相比分别为0.83 倍和0.04 倍；芳香酶促进剂Forskolin 与R2 相对表达量相比分别为3.61 和1.84 倍；芳香酶促进剂地塞米松与R2 相对表达量相比分别为5.76 倍和3.49倍；苯并呋喃类化合物SP25 与R2 相对表达量相比分别为8.13 倍和4.59 倍。实验证实安息香苯并呋喃类化合物SP25 能够促进因RNAi 而发生基因沉默的人芳香酶Aro表达水平的上调。 3、雌激素受体实验结果显示，构建成功重组pERE-pGL3-promoter 荧光素酶报告基因载体和基于报告基因系统的雌激素受体激动剂或拮抗剂的细胞筛选模型。实验结果表明安息香苯并呋喃类化合物SP25 与雌激素受体ERα和ERβ亲和力选择性之比约为3：1 ，SP25通过与雌激素受体ERα结合作用其受体，刺激芳香酶的表达。 本课题通过RNA 干扰、ELISA、荧光实时定量PCR、报告基因筛选模型等技术手段，从细胞水平、蛋白酶水平和基因表达水平、雌激素受体水平等方面系统地研究了从瓦山安息香属植物果实中提取的苯并呋喃SP25 促进促雌激素生成的机理研究。试验结果显示苯并呋喃类化合物SP25 促雌激素生成的主要作用机制是直接促进芳香酶基因表达水平，以及与雌激素受体a 结合，刺激芳香酶活性。 Estrogen is an important hormone that has versatile physiologicalfunctions. Lack of estrogen will lead to many diseases such as lower ovarianfunction, climacteric syndrome and osteoporosis. Excessive estrogen alsoinduces breast carcinoma, oophoroma and endometrial carcinoma and otherdiseases. To depress the estrogen level in tumor tissue to cure carcinomawas widely studied, but there is only few studies reported on the induction ofestrogen and on the regulation of ovary function. We found that the extracts from seeds of Styrax perkinsiae couldpromote the synthesis of estrogen. The active compounds benzofurans wereidentified. Effect of benzofurans may be related to aromatase, but the mechanism was not clear. To reveal the mechanism of these benzofurans to promote estrogensynthesis, the following protocols were adopted: 1 Cytology: 3T3-L1 preadipocytes,human ovary carcinoma celllines OVCAR-3,OVCAR-4,OVCAR-5,OVCAR-8,IGROV1 andbreast carcinoma cell lines MCF-7 and MDA-MB-231 were usedto determine Aro gene expression and estrogen production withRT-PCR AND ELISA methods. Formestane, an aromataseinhibitor, was used as positive control. And dose-curve,time-curve and the effective concentration of SP25 were also studied. 2 Designed 3 pairs of RNAi for human aromatase gene, andtransfected into cell. Aromatase inducer Forskolin andDexamethasone, and aromatase inhibitor Formestane were usedas positive controls. We studied the change of Aro expressionlevel with SP25 by using real-time PCR after RNA interfering. 3 Estrogen Receptor: We constructed the recombined Luciferasereport vector and establish a screening system for estrogenagonist and antagon. With this system, we studied the affinity ofSP25 and estrogen receptor. Results: 1 Differentiated 3T3-L1 preadipocytes¡¢human ovary carcinomacell lines:OVCAR-3, OVCAR-4, OVCAR-8 and breast carcinomacell lines MCF-7, MDA-MB-231 had detected aromatase geneexpression.And OVCAR-3 is more suitable for further aromatasegene function research. 2 In RNAi assay, R2 has a strong interfering effcet in OVCAR-3 cellline, and ratio of C2 (the negative control) to R2 were 118 times(24 hours) and 19 times (48 hours). This means sucessful inRNA interfering. After R2 acted on OVCAR-3 cell line, the ratiosof formestane to R2 were 0.83 and 0.04 times, 5.76 and 3.49times (Dex), 3.61 and 1.84 times (forskolin) and 8.13 and 4.59times (sp25) after drug treated 24 or 48 hours respectively.These results indicated that SP25 can directly induce aromatasegene up-regulation. 3 We had constructed pERE-pGL3-promoter recombined vectorand the Luciferase report gene screening system. Luciferasereport gene assay showed that sp25 had a higher affinity with strogen receptor alpha than estrogen receptor beta, this indicated that SP25 can act on estrogen receptor and induce aromatase. Our results revealed that the mechanisms of benzofuran to promoteestrogen were the upregulation aromatase gene expression and promotion ofaromatase activity and have partially elective affinity with estrogen receptoralpha.

岷江干旱河谷典型阳坡土壤质量动态与评估

横断山区干旱河谷地区的土壤是制约植被恢复的一个关键要素，但干旱条件下土壤质量与动态演变状况并不清楚，本研究以岷江干旱河谷核心地段的土壤为对象，研究了阳坡褐土土壤的物理、化学以及生物学特征在海拔梯度（1800m、2000m、2200m）格局及其时间动态（2005-2007）变化，应用主成分分析综合评价了土壤质量及其时空差异性，明确了土壤质量的变化趋势。主要研究结论如下： 1) 海拔梯度上土壤物理性状的变化，2005 年和2007 年土壤物理性状综合质量随着海拔的升高均得到了优化，即海拔2200 m＞海拔2000 m＞海拔1800 m。 2) 海拔梯度上土壤化学性质的变化，土壤化学综合性质2005 年随海拔升高而趋于变得更优，而2007 年海拔2000 m 最优，海拔1800 m 则最差。 3) 海拔梯度上土壤生物学性质的变化，2005 年土壤生物学性质随海拔升高表现出趋于更好，2007 年海拔2000 m 最优而海拔1800 m 地段最差。 4) 从土壤物理、化学、生物学三方面出发，应用主成分分析，分别分析得出2005 年和2007 年不同海拔高度的土壤质量综合得分。根据综合得分得出土壤质量综合评价的排列顺序为：2005 海拔2200 m＞2007 年海拔2000 m＞2005 年海拔2000 m＞2007 年海拔1800 m＞2007 海拔2200 m＞2005 年海拔1800 m。2005年土壤综合质量随海拔升高而趋好，2007 年则以海拔2000 m 最优，海拔1800 m和2200 m 差异不大。 5）排除人为干扰后，干旱河谷土壤物理性状在海拔1800 m 略有恢复，海拔2000 m 变化不明显，而海拔2200 m 仍有退化趋势；土壤化学性质在海拔1800 m和2000 m 地段得到恢复，而海拔2200 m 处仍有退化；土壤生物学性质在海拔2000 m 地段呈恢复趋势，而1800 m 和2200 m 仍处于退化状态。综合质量分析表明，与2005 年相比，2007 年海拔1800 m 和2000 m 地段土壤质量趋于变优而海拔2200 m 地段仍有退化迹象。 Soil is a key factor that affect the restoration of vegetation in the Hengduan Mountains dry valley area. But the dynamics and quality of soil is not knowed in dry area. In this study, soil physiochemical and biological characteristics ranging from 1800~2200m above sea level from a typical south-facing slope at the Minjiang River dry valley area had been studied, and characteristics of changes in soil quality along altitudinal gradients and time scales were also discussed. The principal component analysis was used to assess the soil quality. The main results were as follows: 1) Changes in soil physical properties along altitudes. Soil physical properties obtained the optimization along with the elevation in 2005 and 2007. 2) Changes in soil chemical properties. It was summarized that soil chemical properties increased with elevation in 2005, but the soil of 2000 m was the best in 2007. 3) Changes in soil biological characteristics. Soil biological properties increased with elevation in 2005, but the soil of 2000 m was the best and 1800 m was the worst in 2007. 4) Change tendency of soil quality. With the soil physics, chemistry and biology characteristics, we analysised the change tendency of soil quality in altitudes. The result indicated that soil quality increased with altituded in 2005, and soil quality of 2000 m was the best in 2007. 5) In brief, the soil quality is by physics, chemistry as well as the biology synthesizes the influence the final outcome. And the soil quality's change was manifested by soil physics, chemistry and the biology characteristics. All the results indicated soil quality still degenerated at 2200 m in in the dry valley of Minjiang River. And soil quality of 1800 m and 2000 m resumed slight.

夏季降水对青海湖水量平衡的影响

IEECAS SKLLQG

陕北水蚀风蚀交错区两种生物结皮对土壤饱和导水率的影响

该文以陕北水蚀风蚀交错区普遍发育的地表和地上两种生物结皮为研究对象,分别以3种非生物结皮(无结皮、物理结皮、去除生物结皮)为对照,使用盘式入渗仪测定其饱和导水率。结果表明:与无结皮土壤相比,两种类型生物结皮均可极显著降低土壤饱和导水率;与去除生物结皮土壤相比,两种类型生物结皮对土壤饱和导水率的降低均不显著;与有物理结皮发育的土壤相比,地表生物结皮对土壤饱和导水率的降低不显著,而地上生物结皮对土壤饱和导水率的降低显著。一方面,两种生物结皮对土壤饱和导水率均有明显降低作用,预示生物结皮在降雨活动中可能会增加径流、降低入渗,阻碍研究区水分亏缺条件下的植被恢复和生态与环境建设。另一方面,与不同的对照相比,生物结皮对土壤饱和导水率的影响截然不同,该结论可在一定程度上解释当前有关生物结皮影响土壤水分入渗方面所存在的分歧。

密度对三种豆科牧草生产力和水分利用率的影响

研究播种密度对3种豆科牧草生产力和水分利用效率的影响,以探讨其在草地农业系统中的合理优化模式。结果表明:沙打旺(Astragelus adsurgens)在高密区第2年生产力最高(18321 kg.hm-2),第3年开始下滑;苜蓿(Medicago sativa)在中、高密区生产力的增幅和增量差别不显著,以第3年最高(22563和22108kg.hm-2);胡枝子(Lespedeza daurica)在中密区第3年最高(7856 kg.hm-2);加大密度使沙打旺提早进入生长盛期,第2年即达到高生产力和较高水分利用效率;苜蓿以中密区(20~30株/m2)效果最好;达乌里胡枝子的生产力和水分利用率偏低,在黄土塬区草地农业中没有优势。

区域可持续发展中生态足迹分析

以辽宁省2004～2008年统计数据为基础,应用生态足迹分析方法,计算了辽宁省2003～2007年的生态足迹和生态承载力。结果表明,辽宁省的人均生态足迹由2003年的2.5875hm2/人增加到2007年的3.3401hm2/人,而同期的人均生态承载力却由0.5652hm2/人降低到0.5590hm2/人,人均生态赤字则由2.0223hm2/人增长到2.7811hm2/人。高额的生态赤字反映出辽宁省人口对自然资源的利用已超出了生态承载力范围。生态赤字的快速增长则反映出当前的发展模式是不可持续的,生态环境处于不安全状态。通过对生态足迹的研究,提出发展中应调整产业结构,引进高新技术,改变生产和消费模式,以实现可持续发展。

Enhanced electrochemiluminescence sensor from tris(2,2 '-bipyridyl) ruthenium(II) incorporated into MCM-41 and an ionic liquid-based carbon paste electrode

The electrochemiluminescence (ECL) of tris(2,29-bipyridyl) ruthenium(II) [Ru(bpy)(3)(2+)] ion-exchanged in the sulfonic-functionalized MCM-41 silicas was developed with tripropylamine (TPrA) as a co-reactant in a carbon paste electrode (CPE) using a room temperature ionic liquid (IL) as a binder. The sulfonic-functionalized silicas MCM-41 were used for preparing an ECL sensor by the electrostatic interactions between Ru( bpy)(3)(2+) cations and sulfonic acid groups. We used the IL as a binder to construct the CPE (IL-CPE) to replace the traditional binder of the CPE (T-CPE)-silicone oil. The results indicated that the MCM-41-modified IL-CPE had more open structures to allow faster diffusion of Ru( bpy)(3)(2+) and that the ionic liquid also acted as a conducting bridge to connect TPrA with Ru( bpy)(3)(2+) sites immobilized in the electrode, resulting in a higher ECL intensity compared with the MCM-41-modified T-CPE. Herein, the detection limit for TPrA of the MCM-41-modified IL-CPE was 7.2 nM, which was two orders of magnitude lower than that observed at the T-CPE. When this new sensor was used in flow injection analysis (FIA), the MCM-41-modified IL-CPE ECL sensor also showed good reproducibility. Furthermore, the sensor could also be renewed easily by mechanical polishing whenever needed.

Potential energy landscape and robustness of a gene regulatory network: Toggle switch

Finding a multidimensional potential landscape is the key for addressing important global issues, such as the robustness of cellular networks. We have uncovered the underlying potential energy landscape of a simple gene regulatory network: a toggle switch. This was realized by explicitly constructing the steady state probability of the gene switch in the protein concentration space in the presence of the intrinsic statistical fluctuations due to the small number of proteins in the cell. We explored the global phase space for the system. We found that the protein synthesis rate and the unbinding rate of proteins to the gene were small relative to the protein degradation rate; the gene switch is monostable with only one stable basin of attraction. When both the protein synthesis rate and the unbinding rate of proteins to the gene are large compared with the protein degradation rate, two global basins of attraction emerge for a toggle switch. These basins correspond to the biologically stable functional states. The potential energy barrier between the two basins determines the time scale of conversion from one to the other. We found as the protein synthesis rate and protein unbinding rate to the gene relative to the protein degradation rate became larger, the potential energy barrier became larger. This also corresponded to systems with less noise or the fluctuations on the protein numbers.

高分子科学实验中异常数据的判别与剔除

简单介绍了对测量值中异常数据进行判别与剔除的三种统计判别方法——3σ准则、格鲁布斯准则和Q检验方法的基本原理,并以高分子科学实验中的两个实例为代表,应用Visual Basic for Applica-tions(VBA)编写的程序模块,对测量值中异常数据的判别与剔除进行了详细讨论。