Construction of p100 Knock Down Stable HepG2 Cell Line with a p100 shRNA Expression Lentiviral System

目的:用表达shRNA的干扰慢病毒载体系统构建p100基因稳定沉默的HepG2人肝癌细胞系.方法:通过RNAi序列设计软件进行p100干扰片段设计,筛选出p100基因的RNAi有效靶序列,进一步合成靶序列的Oligo DNA并构建p LKO.3G-p1001慢病毒载体,用双酶切及测序进行鉴定.将构建好的p100 shRNA表达质粒与慢病毒包装质粒共转染至包装细胞293T,包装产生病毒颗粒.运用病毒颗粒感染HepG2细胞,获得p100稳定沉默的细胞系.荧光显微镜下观察感染效率,利用Western blot方法检测稳定细胞株中p100的沉默效果.结果:成功构建了具有p100干扰效果的慢病毒载体,并获得了稳定沉默p100及对照的HepG2细胞株.慢病毒感染HepG2细胞后,荧光显微镜下观察到90％以上细胞发出绿色荧光,Western blot证实干扰p100后,HepG2细胞株中p100蛋白表达水平明显降低.结论:成功构建了具有p100干扰效果的慢病毒干扰载体及p100稳定沉默的HepG2细胞系.     

复发性抑郁患者的强迫症状及个性因素

目的探讨强迫症状对复发性抑郁障碍患者的影响以及复发性抑郁障碍患者个性因素分析。方法采用症状自评量表(SCL-90),结合临床,对56例复发性抑郁障碍患者进行评定;采用艾森克人格问卷(EPQ)对伴有强迫症状的复发性抑郁障碍患者42例与不伴强迫症状的复发性抑郁障碍患者14例进行对照研究。结果75%的患者伴有强迫症状,伴有强迫与无强迫患者相比EPQ的E分较低和N分较高,差异有显著性(P<0.01)。结论强迫症状是复发性抑郁障碍的一个重要的危险因素,人格因素中内向和神经质与强迫症状有关。     

SELEX与适配体在蛋白质研究中的应用

指数富集的配基系统进化(SELEX)是一种从大容量寡核苷酸文库中经反复分离扩增步骤得到针对靶分子的高亲和力高特异性核酸配基—适配体的体外筛选技术。SELEX技术自1990年发展至今,已涌现出多种筛选模式和分离方法。筛选特异结合蛋白质的SELEX技术发展直接影响和指导了适配体在蛋白质功能调控方面的应用。文章综述了多种SELEX技术在筛选蛋白质方面的发展近况,适配体在蛋白质功能研究中的应用,筛选过程中关键性因素的确定及适配体的前后期修饰。     

二氢卟吩p6醚类光敏剂的合成及光动力抗癌活性研究

目的 基于前期研究表明二氢卟吩f的3-乙烯基成醚修饰具有更强的光敏抗肿瘤活性而设计合成二氢卟吩p₆醚类光敏剂（1）,研究其初步体外光动力抗癌活性。方法 以蚕沙叶绿素a粗提物酸水解产物脱镁叶绿酸a（5）经碱及空气氧化降解制得的紫红素-18（4）为先导物,通过碱水解和CH₂N₂甲基化制得二氢卟吩p₆三甲酯（2）,2与33% HBr加成后再和烷氧醇发生亲核取代反应生成目标化合物（1）,并评价其对黑色素瘤B16-F10细胞的光动力杀伤效应。结果 6个目标化合物1a-1f对黑色素瘤B16-F10细胞的体外光动力抗癌活性均优于同类阳性对照药他拉泊芬和维替泊芬,其结构经电喷雾质谱（ESI-MS）、氢谱（¹H NMR）、碳谱（¹³C NMR）及电喷雾高分辨质谱（ESI-HRMS）确证。结论 二氢卟吩p₆醚类光敏剂具有光动力抗癌活性强、治疗指数（暗毒/光毒比）高等优点,值得进一步开发研究。     

Biosorption and bioaccumulation characteristics of cadmium by plant growth-promoting rhizobacteria

Plant growth-promoting rhizobacteria (PGPR) not only promote growth and heavy metal uptake by plants but are promising biosorbents for heavy metals remediation. However, there exist arguments over whether extracellular adsorption (biosorption) or intracellular accumulation (bioaccumulation) play dominant roles in Cd(ii) adsorption. Therefore, three cadmium-resistant PGPR, Cupriavidus necator GX_5, Sphingomonas sp. GX_15, and Curtobacterium sp. GX_31 were used to study bioaccumulation and biosorption mechanisms under different initial Cd(ii) concentrations, using batch adsorption experiments, desorption experiments, scanning electron microscopy coupled with energy dispersive X-ray (SEM-EDX) spectroscopy, transmission electron microscopy (TEM), and Fourier-transform infrared (FTIR) spectroscopy. In this study, with the increase of the initial Cd(ii) concentrations, the removal efficiency of strains decreased and the adsorption capacity improved. The highest Cd(ii) removal efficiency values were 25.05%, 53.88%, and 86.06% for GX_5, GX_15, and GX_31 with 20 mg l⁻¹ of Cd(ii), while the maximum adsorption capacity values were 7.97, 17.13, and 26.43 mg g⁻¹ of GX_5, GX_15, and GX_31 with 100 mg l⁻¹ of Cd(ii). Meanwhile, the removal efficiency and adsorption capacity could be ordered as GX_31 > GX_15 > GX_5. The dominant adsorption mechanism for GX_5 was bioaccumulation (50.66–60.38%), while the dominant mechanisms for GX_15 and GX_31 were biosorptions (60.29–64.89% and 75.93–79.45%, respectively). The bioaccumulation and biosorption mechanisms were verified by SEM-EDX, TEM and FTIR spectroscopy. These investigations could provide a more comprehensive understanding of metal-bacteria sorption reactions as well as practical application in remediation of heavy metals.

Plant growth-promoting rhizobacteria (PGPR) not only promote growth and heavy metal uptake by plants but are promising biosorbents for heavy metals remediation.     

Carbon nanotube-based substrates for modulation of human pluripotent stem cell fate

We investigated the biological response of human pluripotent stem cells (hPSCs) cultured on a carbon nanotube (CNT) array-based substrate with the long term goal to direct hPSC germ layer specification for a wide variety of tissue engineering applications. CNT arrays were fabricated using a chemical vapor deposition system allowing for control over surface roughness and mechanical stiffness. Our results demonstrated that hPSCs readily attach to hydrophilized and extracellular matrix coated CNT arrays. hPSCs cultured as colonies in conditions supporting self-renewal demonstrated the morphology and marker expression of undifferentiated hPSCs. Conditions inducing spontaneous differentiation lead to hPSC commitment to all three embryonic germ layers as assessed by immunostaining and RT-PCR analysis. Strikingly, the physical characteristics of CNT arrays favored mesodermal specification of hPSCs. This is contradictory to the behavior of hPSCs on traditional tissue culture plastic which promotes the development of ectoderm. Altogether, these results demonstrate the potential of CNT arrays to be used in the generation of new platforms that allow for precise control of hPSC differentiation by tuning the characteristics of their physical microenvironment.     

SiRNA-phospholipid conjugates for gene and drug delivery in cancer treatment

Due to low charge density and stiff backbone structure, small interfering RNA (siRNA) has inherently poor binding ability to cationic polymers and lipid carriers, which results in low siRNA loading efficiency and limits siRNA success in clinical application. Here, siRNA-phospholipids conjugates are developed, which integrate the characteristics of the two phospholipids to self-assemble via hydrophilic siRNA and hydrophobic phospholipid tails to overcome the siRNA's stiff backbone structures and enhance the siRNA loading efficiency. In this study, the thiol-modified sense and antisense siRNA are chemically conjugated with phospholipids to form sense and antisense siRNA-phospholipid, and then these sense or antisense siRNA-phospholipids with equal amounts are annealed to generate siRNA-phospholipids. The siRNA-phospholipids can serve dual functions as agents that can silence gene expression and as a component of nanoparticles to embed hydrophobic anticancer drugs to cure tumor. siRNA-phospholipids together with cationic lipids and DSPE-PEG2000 fuse around PLGA to form siRNA-phospholipids enveloped nanoparticles (siRNA-PCNPs), which can deliver siRNAs and hydrophobic anticancer drugs into tumor. In animal models, intravenously injected siRNA-PCNPs embedded DOX (siPlk1-PCNPs/DOX) is highly effective in inhibiting tumor growth. The results indicate that the siRNA-PCNPs can be potentially applied as a safe and efficient gene and anticancer drug delivery carrier.     

思想政治课教学应遵循的原则

思想政治课程作为学校教育的必修课,能给学生正确的思想道德、文化价值等各个方面的理论指导,是学生素质教育的必要组成部分,尤其是面对当前新的社会发展形势,思想政治课显得尤为重要。就思想政治课教学过程中的一些心得体会供大家探讨。     

OCE对体外培养的视网膜组织的抗氧化作用

目的：观察不同成份的培养液对成年犬视网膜组织进行体外培养的前后相关生化指标的改变，旨在为视网膜移植提供抗氧化能力的保护性方法。方法：将健康成年家犬74眼随机分成六组，Ⅰ（新鲜视网膜组）Ⅱ（常规培养液组）Ⅲ（高浓度OCE组）Ⅳ（低浓度OCE组）Ⅴ（高浓度全脑组织提取液组Ⅵ（低浓度全脑组织提取液组）培养一周。六组视网膜取材后进行SOD、MDA、G－Px,GST,T－AOC生化指标的测定。结果：Ⅱ组与Ⅰ组SOD、G－Px、GST、T－AOC明显下降；DMA值增多非常显著；Ⅲ、Ⅳ组与Ⅱ组比较，T－AOC、SOD、G－Px、GST值显著增高，Ⅳ组明显高于其它各培养组；Ⅲ、Ⅳ组与Ⅱ组比较，MDA值降低明显，Ⅳ组降低最明显。结论：成年犬OCE中含有清除自由基、增强抗氧化能力的特殊物质。并且低浓度OCE对体外培养的视网膜组织抗氧化作用显著。