Adjuvant Effects of Dual Co-stimulatory Molecules on Cellular Responses to HIV Multiple-epitope DNA Vaccination

Designing adjuvants that can induce strong cytotoxic T cell responses is largely required for preparing DNA vaccines. In this study we explored dual costimulatory molecules 4-1BBL and OX40L as adjuvants to improve the efficiency of the HIV multiple-epitope DNA vaccine. When explored in the human dendritic cell-T cell based coculture system, dual costimulatory molecules significantly enhanced the anti-HIV T cell response of the HIV multiple-epitope DNA vaccine, as detected by intracellular cytokine staining to HIV antigens, cytokines accumulation in the cultures, and antigen-specific cytotoxic T lymphocyte responses. These results suggest that dual costimulatory molecules 4-1BBL and OX40L can effectively increase the potential of the HIV multiple-epitope antigen DNA vaccine and may provide an exciting approach for HIV therapy.

共刺激分子B7H3通过调控DNA拓扑异构酶Ⅱα的表达对乳腺癌多柔比星化疗的影响

目的探讨共刺激分子B7H3(costimulatory molecule B7H3,B7H3)通过调控DNA拓扑异构酶Ⅱα(DNA topoisom eraseⅡalpha,TOP2A)的表达对乳腺癌多柔比星化疗敏感性的影响。方法收集临床标本,采用免疫组织化学染色验证B7H3表达与多柔比星化疗敏感性的相关性。采用成簇规律间隔短回文重复/Cas9基因编辑技术敲除B7H3,通过蛋白质印迹法和流式细胞术验证敲除结果。采用半抑制浓度值分析、集落形成、蛋白质印迹法检测凋亡蛋白、荧光双染色检测凋亡细胞以及细胞周期测定等方法验证B7H3表达与多柔比星作用的敏感性。小干扰TOP2A及其过表达TOP2A回复实验检测细胞凋亡。构建小鼠皮下成瘤模型验证体内B7H3表达与多柔比星化疗的作用。结果高表达B7H3的肿瘤组织对多柔比星治疗更加敏感。成功构建B7H3敲除的乳腺癌细胞。B7H3高表达的乳腺癌细胞对多柔比星作用更敏感。B7H3通过激活核因子κB(nuclear factor-κB,NF-κB)途径促进TOP2A的表达,进而增加乳腺癌细胞对多柔比星作用的敏感性。小鼠体内实验证明,B7H3高表达可增加乳腺癌对多柔比星化疗的敏感性。结论B7H3通过激活NF-κB通路调控TOP2A的表达使乳腺癌对多柔比星化疗更敏感。

Mechanics of Twisted DNA Molecule Adsorbed on a Biological Membrane

DNA is the carrier of all cellular genetic information and increasingly used in nanotechnology. The study of DNA molecule achieved in vitro while submitting the DNA to all chemicals agent capabilities to destabilize links hydrogen, such as pH, temperature. In fact, the DNA enveloped in the membrane cellular, so it is legitimate to study the influence of membrane undulations. In this work, we try to show that the fluctuations of the membrane can be considerate as a physics agent is also capable to destabilize links hydrogen. In this investigation, we assume that each pair base formed an angle an with the membrane’s surface. We have proposed a theoretical model, and we have established a relationship between the angle formed by the pair base θeq and an angle formed by the membrane and each pair base. We assume that DNA and biomembrane interact via a realistic potential of Morse type. To this end, use is made of a generalized model that extends that introduced by M. Peyrard and A. R. Bishop in the past modified by M. Zoli. This generalized model is based on the resolution of a Schrödinger-like equation. The exact resolution gives the expression of the ground state, and the associated eigenvalue (energy) that equals the free energy, in the thermodynamic limit. First, we compute the denaturation temperature of DNA strands critical temperature. Second, we deduce all critical properties that mainly depend on the parameters of the model, and we quantify the effects of the membrane undulations. These undulations renormalize all physical quantities, such as harmonic stacking, melting temperature, eigenfunctions, eigenvalues and regular part of specific heat.

Naked DNA in cells:An inducer of major histocompatibility complex molecules to evoke autoimmune responses?

The major histocompatibility complex(MHC) is the exclusive chaperone that presents intracellular antigens,either self or foreign to T cells.Interestingly,aberrant expression of MHC molecules has been reported in various autoimmune target tissues such as thyroid follicular cells in Grave's disease.Herein,we review the discovery of an unexpected effect of cytosolic doublestranded DNA(ds DNA),despite its origins,to induce antigen processing and presenting genes,including MHC molecules,in non-immune cells.Moreover,we highlight several recent studies that suggest cell injury endows thyroid epithelial cells with a phenotype of mature antigen presenting cells by inducing multiple antigen processing and presenting genes via releasing genomic DNA fragments into the cytosol.We discuss the possibility that such cytosolic ds DNA,in naked form without binding to histone proteins,might be involved in the development of cell damage-triggered autoimmune responses.We also discuss the possible molecular mechanism by which cytosolic ds DNA can induce MHC molecules.It is reasonable to speculate that cytosolic ds DNA-induced MHC class Ⅰ is partially due to an autocrine/paracrine effect of type Ⅰ interferon(IFN).While the mechanism of cytosolic ds DNA-induced MHC class Ⅱ expression appears,at least partially,distinct from that mediated by IFN-γ.Further in-depth are required to clarify this picture.

DNA-卟啉杂化分子合成技术研究进展

DNA-卟啉杂化分子作为一种新型的有机-生物杂化材料,因其卓越的光学性质、良好的水溶性等独特的物理和化学性质,在化学和生物科学领域引起广泛关注。该类杂化分子融合了DNA的生物活性和识别能力,以及卟啉分子的光敏性和荧光特性,使得其在光动力疗法、生物传感、荧光成像等多个领域展现出巨大的应用潜力。本文论述了DNA-卟啉杂化分子的非共价和共价合成技术的研究进展,并对其应用前景进行了展望。

DNA-有机分子杂化体的合成及应用研究

随着DNA化学修饰技术的发展,疏水分子和DNA通过固相反应合成了两亲性DNA有机杂化物,两亲性有机杂化物可以被视为新的构建块,然后在溶液中自发组装成各种纳米结构,组装体在药物输送、靶向递送及生物传感等领域有着重要的应用。论述了DNA-有机分子杂化体的组装、合成方法以及在生物医药方面的研究进展。

Modified "DMC" technique for stretching DNA molecules

A modified "dynamic molecular combing"(DMC) technique used for stretching double-stranded DNA is reported. DNA molecules were stretched on the silanized mica surface by this technique, its speed being precisely controlled with a computer. This approach combined the precise DNA stretching method with high resolution AFM imaging at nanometer scale, thus making it useful for DNA alignment manipulation and subsequent gene research.

Catenary-like pattern of stretching DNA molecules:experiments and models

It was verified that our "s-suspension bridge" model may be not only responsible for catenary-like patterns of DNA molecules which were formed in our experiments, but also for catenary-like patterns of DNA molecules in literature.

Nano-manipulation of single DNA molecules

Nano-manipulation of single atoms and molecules is a critical technique in nanoscience and nanotech- nology. This review paper will focus on the recent development of the manipulation of single DNA molecules based on atomic force microscopy (AFM). Precise manipulation has been realized including varied manipulating modes such as “cutting”, “pushing”, “folding”, “kneading”, “picking up”, “dipping”, etc. The cutting accuracy is dominated by the size of the AFM tip, which is usually 10nm or less. Single DNA fragments can be cut and picked up and then amplified by single molecule PCR. Thus positioning isolation and sequencing can be performed.

Improved data analysis method of single-molecule experiments based on probability optimization

To extract the dynamic parameters from single molecule manipulation experiments, usually lots of data at different forces need to be recorded. But the measuring time of a single molecule is limited due to breakage of the tether or degradation of the molecule. Here we propose a data analysis method based on probability maximizalion of the recorded time trace to extract the dynamic parameters from a single measurement. The feasibility of this method was verified by dealing with the simulation data of a two-state system. We also applied this method to estimate the parameters of DNA hairpin folding and unfolding dynamics measured by a magnetic tweezers experiment.

Molecular Dynamics Simulations of the DNA-CNT Interaction Process: Hybrid Quantum Chemistry Potential and Classical Trajectory Approach

In this work the quantum chemistry Tersoff potential in combination with classical trajectory calculations was used to investigate the interaction of the DNA molecule with a carbon nanotube (CNT). The so-called hybrid approach—the classical and quantum-chemical modeling, where the force fields and interaction between particles are based on a definite (but not unique) description method, has been outlined in some detail. In such approach the molecules are described as a set of spheres and springs, thereby the spheres imitate classical particles and the spring the interaction force fields provided by quantum chemistry laws. The Tersoff potential in hybrid molecular dynamics (MD) simulations correctly describes the nature of covalent bonding. The aim of the present work was to estimate the dynamical and structural behavior of the DNA-CNT system at ambient temperature conditions. The dynamical configurations were built up for the DNA molecule interacting with the CNT. The analysis of generated МD configurations for the DNA-CNT complex was carried out. For the DNA-CNT system the observations reveal an encapsulation-like behavior of the DNA chain inside the CNT chain. The discussions were made on possible use of the DNA-CNT complex as a candidate material in drug delivery and related systems.

Interaction between human telomeric G-quadruplexes characterized by single molecule magnetic tweezers

Human telomeric G-quadruplex plays a crucial role in regulating the genome stability. Despite extensive studies on structures and kinetics of monomeric G-quadruplex, the interaction between G-quadruplexes is still in debate. In this work,we employ magnetic tweezers to investigate the folding and unfolding kinetics of two contiguous G-quadruplexes in 100-mM K~＋buffer. The interaction between G-quadruplexes and the consequent effect on the kinetics of G-quadruplex are revealed. The linker sequence between G-quadruplexes is further found to play an important role in the interaction between two G-quadruplexes. Our results provide a high-resolution insight into kinetics of multimeric G-quadruplexes and genome stability.

DNA损伤修复的单分子水平研究进展

外源或内源的DNA损伤在生物体内持续发生。DNA损伤修复的缺陷与很多疾病甚至癌症等息息相关,而生物细胞进化出一系列精密的修复机制以耐受或切除这些损伤。单分子技术区别于常规的生化、分子生物学等手段,可以在体外和活细胞内研究DNA修复相关生物分子的动态反应特征,从而对DNA修复机制进行更充分的剖析。文章围绕常见的DNA损伤及其修复类型,阐述了近年来利用原子力显微镜、磁镊、光镊等单分子操控技术,以及全内反射荧光显微镜、光激活定位显微镜和超分辨显微示踪等单分子荧光成像技术在DNA修复机制研究中取得的进展,梳理了利用单分子技术解决的长期存在的关于DNA修复难题,并展望了单分子技术联合其他交叉学科技术在研究DNA修复机制方面的前景。

表征不同DNA高阶结构的单分子方法

DNA不仅是生命遗传信息的载体,而且是一种高度可编程和自组装的纳米材料。不同的DNA结构与其生物和化学功能有关。因此,了解各种DNA结构的物理和化学性质在生物学和纳米化学中具有重要意义。然而,批量分析忽略了溶液中DNA结构的异质性。单分子方法是观察单个分子行为和探测自由能态高度非均质性的有力工具。在本文中,介绍了单分子方法,包括单分子检测和操作方法,并讨论了这些方法如何有助于测量单链/双链DNA(ss/dsDNA)、DNA高阶结构和DNA纳米结构的分子性质。本文为DNA纳米技术与单分子方法的结合提供了一个新的视角,以了解DNA和其他生物物质和软物质的生物物理特性。

HBV DNA聚合酶在介导HBV相关肝细胞癌肿瘤细胞免疫逃逸中的作用

目的本研究旨在明确HBV DNA聚合酶是否与T淋巴细胞功能衰竭相关,进而介导HBV相关肝细胞癌(HCC)肿瘤细胞的免疫逃逸以及探寻具体的分子机制。方法稳定转染带有Flag标签的HBV DNA聚合酶表达质粒(FlagHBV-P)和细胞间黏附分子-1(ICAM1)的肝癌细胞系Huh7、HepG2与Jurkat细胞共培养,MTT、qRT-PCR、酶联免疫吸附测定(ELISA)实验分别检测Jurkat细胞增殖、活化(CD69表达)以及细胞因子IFN-γ分泌情况。RNA-seq筛选稳转细胞系与对照细胞中表达差异的免疫相关分子,mRNA半衰期及蛋白半衰期实验确定HBV DNA聚合酶对免疫相关分子具体在何种水平产生影响。网站预测此免疫相关分子启动子区域可能结合的转录因子,Western Blot实验验证转录因子对免疫相关分子的影响,挽救实验确定HBV DNA聚合酶是否通过此转录因子影响免疫相关分子的表达水平。两组间比较采用成组t检验。结果与对照组相比,实验组细胞(Huh7、HepG2)增殖、活化及细胞因子分泌明显降低(P值均<0.01)。与对照细胞相比,实验组细胞(Huh7、Hep G2)的ICAM1 mRNA和蛋白水平均降低(P值均<0.01)。网站预测ICAM1启动子并初步筛选锚定了NFKB1、RELA和STAT3。与对照组相比,实验组细胞(Huh7、Hep G2)p65蛋白的表达水平明显降低(P值均<0.01)。过表达p65后,ICAM1蛋白表达水平明显升高,降表达p65后,ICAM1蛋白表达水平明显降低(P值均<0.01),挽救实验中过表达p65后,对照组与实验组细胞的ICAM1表达水平无明显差异(P值均>0.05)。过表达ICAM1后,对照组与实验组细胞(Huh7、Hep G2)的增殖、活化及细胞因子分泌无明显差异(P值均>0.05)。结论HBV DNA聚合酶通过抑制NF-κB中p65的表达来下调ICAM1的水平以介导HCC免疫逃逸。

RNA聚合酶动态调控DNA转录的单分子水平研究进展

真核生物的RNA聚合酶Ⅱ(Pol Ⅱ)和原核生物的RNA聚合酶(RNAP)主要负责转录合成信使RNA(mRNA),调控不同基因的转录水平,以调节生物体的生长发育和应对复杂多变的环境。研究者采用传统的荧光显微镜观测到RNAP可形成团簇,据此针对DNA转录调控提出“转录工厂”模型。随着单分子技术的发展,研究者在单分子水平上观测到了活细胞中RNAP动态调控DNA转录,提出RNAP可以通过液-液相分离机制进行转录调控。该综述总结了不同单分子荧光显微镜的技术原理,以及相关的荧光探针标记方法,并介绍了在真核生物和原核生物中应用单分子成像技术可视化RNA聚合酶动态调控DNA转录过程的研究进展,最后展望了单分子技术在转录调控研究中的应用前景。

DNA与其靶向分子相互作用研究进展

DNA与其靶向分子相互作用的研究不仅对阐述一些抗肿瘤、抗病毒药物及致癌物的作用机理 ,而且对进一步指导人工核酸酶的合成及 DNA高级结构研究等方面的工作都具有重要意义 .本文着重评述了近年来不同结构类型的 DNA靶向分子与 DNA相互作用研究方面的进展 .

有机药物分子与DNA相互作用的研究进展

综述了有机药物分子与DNA相互作用的模式、影响因素、并着重介绍了其相互作用的电化学等研究方法,概括了其近几年的研究进展,并在此基础上提出了在药物分子与DNA相互作用的研究工作中应解决的问题。