光动力作用可致人肝癌细胞HepG2凋亡的实验研究

为探讨光动力作用（PDT）对体外培养的人肝癌细胞HepG2的杀伤效应，应用体外培养的人肝癌细胞HepG2，采用血卟啉衍生物（HPD）为光敏剂，用He-Ne激光器（波长632．8nm，能量密度120J／cm^2）进行激光照射，以系列浓度经不同剂量的光照后用MTT法测定PDT对肝癌细胞的相对抑制率，流式细胞仪检测其凋亡率．发现随光敏剂浓度的升高和照光剂量的增加，光动力作用对细胞的相对抑制率逐渐增大，在低光动力剂量下明显上升，随后上升渐趋缓慢达平台期．取在特定的光动力剂量下，肝癌细胞出现明显的凋亡现象，与对照组有明显的差异．实验结果表明，光动力作用下对体外培养的人肝癌细胞具有明确的杀伤效应，其机理可能与细胞凋亡有关．     

活细胞应激反应过程中线粒体和核仁微环境动力学的荧光寿命成像研究

核仁和线粒体在维持细胞平衡发挥重要作用,研究其生理过程有助于深入了解生物学功能.本文采用一种红色荧光的芘罗丹明荧光探针在不同条件下靶向标记细胞线粒体和核仁.通过激光共聚焦成像和荧光寿命成像技术分析HeLa细胞在光照和药物刺激下细胞凋亡的形态变化,并利用相图定量分析了线粒体与核仁的微环境变化,确定在稳态HeLa细胞中探针标记到的线粒体的平均荧光寿命约为3.65 ns,线粒体黏度约为66×10–3 Pa·s.在激光光照后,探针标记到HeLa细胞线粒体的荧光寿命降至3.61 ns,对应线粒体黏度增至约131×10–3 Pa·s;使用紫杉醇和秋水仙碱诱导细胞凋亡,观察到探针标记于HeLa细胞核仁的荧光寿命先增加后降低,反映了在HeLa细胞凋亡过程中核仁微环境的变化,证明HeLa细胞在非稳态情况下核仁和线粒体的功能变化,为线粒体和核仁功能障碍相关疾病研究提供了新的研究方法.     

光动力作用促人肝癌细胞HepG2凋亡及对bax，bcl-2蛋白表达的影响

应用流式细胞仪分析光动力作用后细胞凋亡及免疫组化染色检测凋亡相关蛋白bcl-2蛋白，bax蛋白表达水平．发现血卟啉光动力实验组人肝癌细胞HepG2凋亡率达24．23±1．86％，与对照组相比，差异非常显著（P〈0．01）；光动力作用后凋亡相关蛋白bcl-2表达显著高于对照组（P〈0．05）；实验结果显示血卟啉光动力作用具有诱导人肝癌细胞HepG2凋亡的生物学效应，而凋亡调控蛋白bcl-2表达水平的降低可能是血叶林光动力作用诱导人肝癌细胞HepG2凋亡的一个重要机制．     

金纳米棒标记HepG2人肝癌细胞的荧光成像及其AFM探测

金纳米棒具有独特的光学性质,在生物医学领域有着广泛而重要的应用前景.本文制备了长径比为8∶1的金纳米棒,其在480 nm波长激发下,在560 nm和707 nm波长处有两个荧光发射峰.基于金纳米棒的荧光性质,将其标记于HepG2人肝癌细胞表面,利用激光扫描共聚焦显微镜对标记后的细胞进行荧光成像.在488 nm激发下,获...     

一种轴向核苷修饰硅酞菁的合成与光动力抗癌活性

合成了一种轴向核苷修饰硅酞菁,即二[5′-(2′,3′-O-异丙基)-5-甲基胞苷氧基] 硅酞菁,并通过1H NMR和HR-MS等手段进行了表征。该化合物在N,N-二甲基甲酰胺和含1% Cremophor EL水溶液中以单体形式存在,Q带最大吸收波长分别位于676和679 nm,荧光发射峰分别位于685和689 nm。离体光动力抗癌活性表明,该化合物具有显著的光动力抗癌活性,对人肝癌细胞HepG2的半致死浓度(IC₅₀)低至7.8×10-8mol·L-1。荧光共聚焦显微镜研究显示,SiPcG可定位于细胞线粒体中。研究表明,SiPcG是一种有发展潜力的新型抗癌光敏剂。     

飞秒激光对高反膜的破坏及其超快动力学过程

研究了800nm飞秒激光照射下45°高反膜ZrO2-Si O2的破坏及其超快动力学过程。利用原子力显微镜和扫描电镜观察了材料的烧蚀形貌,测量了破坏阈值与脉冲宽度、烧蚀深度与脉冲能量的依赖关系。随着脉冲宽度从50fs增加到900fs,其烧蚀阈值从0.35J/cm2增加到1.78J/cm2。烧蚀深度与激光能流密度近似成对数关系。当激光强度略高于烧蚀阈值时,材料很快被烧蚀到几百纳米,烧蚀深度表现出明显的层状特性。同时,利用建立的抽运探针实验系统,测量了高强度抽运脉冲作用下材料对探针光的反射率随延迟时间的变化,揭示了薄膜烧蚀的超快动力学过程。实验结果表明高反膜表层的材料对烧蚀特性有重要影响。     

几种新型金属配合物热致三阶非线性光学特性

利用连续激光Z-扫描方法研究了过渡金属Ni、Cu、Pd的新型有机配合物的热致三阶非线性光学特性。结果表明：在488nm和632．8nm波长连续激光激发下,这几种配合物的光学非线性折射主要来源于热效应。并由实验曲线计算了它们的热光系数dn／dT及热致非线性折射率n2。通过比较,发现热致非线性折射率与配合物分子结构、激发波长及在该波长下的线性吸收强弱有关。金属中心原子的原子序数越大,热光系数和折射率也相应的越大。在488nm波长激光激发下的dn／dT和n2大于该配合物在632．8nm波长激发下的dn／dT和n2,在同一波长下,化合物溶液在该波长下的吸收越强,其dn／dT和n2值也越大。     

二氧化硅增透膜微观结构与激光损伤阈值的调控

采用溶胶-凝胶技术制备了二氧化硅增透膜,通过向溶胶中添加高分子聚乙烯醇缩丁醛（PVB）,调控胶体的粒径,进而控制膜层微观结构,研究膜层微观结构与激光损伤阈值的关系。纳米粒度仪和扫描探针显微镜测试表明:PVB加入溶胶后,控制了二氧化硅胶粒的生长,使二氧化硅胶粒生长更均匀,因而膜层的微观结构更均匀。当PVB质量分数为1%时,胶体粒径为15 nm,分散系数小于0.1。用该胶体镀膜,膜层均匀,表面粗糙度小于3.25 nm。并且PVB加入后增加了膜层胶粒间的黏附性,使得膜层强度增大。PVB加入使膜层的激光损伤阈值有所增加。当PVB的添加量为1%时,膜层的激光损失阈值从30.0 J/cm2增加到40.1 J/cm2。膜层激光损伤阈值的增加与膜层微观均匀性和物理强度的增加有关。     

1064nm纳秒激光对石墨烯的损伤效应研究

通过化学气相沉积法制备,并转移到基片得到1~3层石墨烯样品。利用霍尔效应及微区拉曼光谱测量,结合光学显微镜观察,分析了不同层数石墨烯在1064nm纳秒激光辐照下的损伤特性。实验发现1~3层石墨烯的激光损伤阈值依次降低,分别为:单层0.45J/cm2,2层0.34J/cm2,3层0.23J/cm2。激光强度超过阈值时,石墨烯薄膜电阻增大,载流子迁移率降低。通过光学显微镜观察发现局部区域破损,破损区域的拉曼光谱中1580cm-1左右的G峰和2700cm-1左右的2D峰高度比发生变化。实验结果表明1064nm纳秒激光辐照石墨烯主要为剥离作用。     

类锂铝10.57和15.47纳米X光激光增益研究

在LF-11(10^(11)W)激光装置上,开展了类锂铝(Al^(10+))的X光激光实验研究。实验中,激光器运行在线聚焦工作状态,波长1.06μm,脉冲宽度约为200ps,能量约20J。线聚焦长12mm,宽约100μm。实验中使用了厚铝靶(1.2μm)和薄膜铝靶(60和94.7nm),用时间积分掠入射光栅谱仪测量线状等离子体轴向XUV谱,用针孔照相机监视线聚焦状态。结果表明,Al^(10+)离子的10.57(3d-5f)和15.47(3d-4f)nm线的强度随等离子体的长度呈现明显的非线性增长。这两条激光跃迁线,用60nm铝靶时,增益系数分别为3.18和2.26cm^(-1);用1.2μm铝靶时,增益系数分别为1.67和0.91cm^(-1);用94.7nm铝靶时,波长为10.57nm线的增益系数为1.78cm^(-1)。说明厚度合适的薄膜靶能获得较高的增益。     

Confocal fluorescence lifetime imaging of free calcium in single cells

Ca²⁺ concentrations in biological cells are widely studied with fluorescent probes. The probes have a high selectivity for free calcium and exhibit marked changes in their photophysical properties upon binding. The differences in the fluorescent lifetime of the probes can now be used as a contrast mechanism for imaging purposes. This technique can be further exploited for the quantitative determination of ion concentrations within the cells. We describe the use of a fast fluorescence lifetime imaging method in combination with a standard confocal laser scanning microscope for the determination of Ca²⁺ concentrations in single rat cardiac myocytes using the intensity probe Calcium Green.     

Chromatin condensation and sensitivity of DNA in situ to denaturation during cell cycle and apoptosis--a confocal microscopy study

The goal of this study was to construct high resolution 3D confocal images of regions of condensed and extended chromatin in cell nuclei and individual chromosomes. It has been shown previously that sensitivity of DNA in situ to denaturation correlates with chromatin condensation and varies during cell cycle and apoptosis. Thus, detection of DNA which was partially denatured in situ provided a means to image areas of condensed chromatin. DNA denaturation was detected using a metachromatic dye acridine orange (AO) which differentially stains single stranded (ss) and double-stranded (ds) DNA sections. Early studies of denaturability of cellular DNA utilized flow cytometry and standard fluorescence microscopy. These techniques could not reveal small local differences in DNA denaturability within cell nucleus or in individual chromosomes. For instance, it was not possible to detect the initial points of chromosome condensation in G2-phase of the division cycle or in apoptosis. In order to achieve this goal we have recently extended these studies by applying confocal microscopy. We investigated DNA denaturability in normal human fibroblasts and HL-60 leukemic cells, at different stages of cell cycle and apoptosis. Following removal of RNA and partial denaturation of DNA with acid cells were stained with AO. Green (530 nm) and red (640 nm) fluorescence (exc. 457 nm) of non-denatured and denatured DNA was imaged by confocal microscopy. Blind deconvolution was used to further improve the quality of 3D images. Photobleaching of AO fluorescence was minimized and a correction for chromatic aberration and register shift was implemented. Nuclei of interphase cells exhibited predominantly green fluorescence representing AO binding to ds DNA. Punctuate areas of red fluorescence representing AO binding to denatured DNA and most likely associated with local regions of condensed chromatin were also present in all interphase nuclei. The proportion of denatured DNA increased in cells entering mitosis. In prophase individual condensing chromosomes exhibited varied proportions of green and red fluorescence indicating different content of denatured chromatin. In some chromosomes bands of denatured and denaturation-resistant chromatin were clearly resolved. In metaphase and anaphase chromosomes exhibited red fluorescence along all length of their arms indicating the highest and uniform susceptibility to denaturation. In telophase chromosomes contained predominantly denaturation-resistant DNA again and denaturated regions were significantly less abundant. At cytokinesis some decondensing chromosomes were still resolved. At this stage almost all regions of denatured DNA were located close to nuclear envelope. These regions may correspond to pockets of heterochromatin reforming at nuclear periphery. In early apoptosis condensation of chromatin appeared to commence in several distinct regions within nucleus. Some apoptotic bodies contained condensed chromatin surrounding central regions of extended chromatin. At late stages of apoptosis the whole volume of apoptotic bodies was occupied by condensed chromatin.     

LiF∶F2晶体中实现三维光数据存储的实验研究

基于LiF∶F2晶体在近红外飞秒脉冲激光作用下产生F2色心向F+3色心的转变,以及两种色心荧光光谱的不同,实现了荧光反射共焦读出和多光子写入的三维光数据存储的原理性实验.钛宝石再生放大器输出的脉冲宽度100 fs、中心波长800 nm、重复频率1 kHz的超短脉冲激光束,用数值孔径0.68的显微物镜聚焦到LiF：F2晶体内部,通过移动晶体实现了三维逐位式数据写入；用405 nm的连续蓝光激发存储位,通过探测F+3色心产生的540 nm荧光,实现了对信息位进行快速非破坏性的反射共焦读出.与多光子三维存储的透射共焦散射读出和相衬读出相比,格式与现存光盘技术兼容,结构简单；更重要的是,存储信息位是依靠荧光光谱的变化,折射率变化很小,可以有效增加读出层数.     

一种示踪干细胞的磁共振-荧光双模成像探针

本文设计并合成了Gd基磁共振-荧光双模成像探针——Gd-DOTA-PEG-GA,通过电穿孔的方式标记人源间充质干细胞（hMSCs）.电穿孔标记诱导细胞将探针组装成团簇状纳米粒子进入细胞质,显著延长其与细胞结合的时间,并呈现出明显的T₂信号减弱效应,且信号减弱效应可以持续7天以上.在水溶液中,该探针的发射带集中在498 nm,并且荧光强度在一周内无明显衰减.该探针标记的细胞在荧光倒置显微镜下呈现绿色荧光.这些结果表明该探针可以作为磁共振-荧光双模成像探针用于干细胞示踪.     

多糖对白细胞荧光发射特性及变化机制研究

细胞微环境的稳定是保持细胞正常增殖、代谢和功能活动的重要条件,微环境成分的异常变化可使细胞发生病变。采用荧光光谱技术研究离体白细胞在多糖微环境中荧光发射特性的变化规律和发光机制,并进一步的分析了多糖对白细胞的生物活性的影响。实验结果表明：当白细胞受波长为407 nm的激光照射时,发射位于450 nm的荧光。在加入脂多糖或葡聚糖时,白细胞的荧光发射峰的位置不会变化,峰值受到影响。脂多糖的加入会减弱白细胞荧光峰强度,且荧光强度随脂多糖浓度（0～500 μg·mL-1范围内）的增加而持续减弱。而葡聚糖可以一定程度增加白细胞的荧光强度,浓度越高,荧光强度越大。分析认为白细胞发射的450 nm荧光来自发射物质烟酰胺腺嘌呤二核苷酸(NADH)。白细胞内NADH随着离体时间的增长,被氧化成不发荧光的烟酰胺腺嘌呤磷酸二核苷酸（NAD+）,导致细胞荧光峰值下降,从而引起细胞凋亡。加入脂多糖产生的羟自由基（·OH）会与NADH发生氧化反应,因此脂多糖加速了NADH的消耗,导致白细胞荧光减弱,加快细胞凋亡。而葡聚糖主要是由葡萄糖单体组成,葡聚糖的加入会将NAD+还原成NADH,因此延缓了白细胞的凋亡。分析认为脂多糖可以加速白细胞的凋亡,提高细胞发生炎症甚至是肿瘤的机率,而葡聚糖对白细胞有保护作用。该研究为研究肿瘤的发生和发展过程以及治疗提供有价值的参考。     

Effects of low-intensity laser irradiation on the apoptosis of rabbit vascular smooth muscle cells in culture

Restenosis is a major complication after coronary intervention therapy. Excessive proliferation of vascular smooth muscle cells (VSMCs) and a decline in their apoptosis, which eventually leads to excessive neointimal thickening in coronary arteries, are the main causes of restenosis. Induction of the apoptosis of VSMCs and inhibition of excessive proliferation of VSMCs are therefore crucial for the prevention of restenosis, and low-intensity laser irradiation of coronary arteries may play a promising role in keeping this in balance. In this study, we used in vitro cultured rabbit VSMCs to investigate the effects of low-intensity laser irradiation at a wavelength of 532 nm on the apoptosis of VSMCs via morphological observation and molecular biology. The results showed that apoptotic bodies and obvious intranuclear apoptosis-positive particles formed within VSMCs 24 h after laser irradiation, suggesting that low-intensity laser irradiation at certain doses can inhibit the proliferation of VSMCs by promoting their apoptosis. This experiment provides evidences for further animal experiments and clinical trials on prevention and treatment of restenosis by intracoronary low-intensity laser irradiation at a wavelength of 532 nm.     

ANXA2 enhances the progression of hepatocellular carcinoma via remodeling the cell motility associated structures

Hepatocellular carcinoma (HCC) ranks as the fifth most common malignancy worldwide. The detailed mechanism of signal regulation for HCC progression is still not known, and the high motility of cancer cells is known as a core property for cancer progression maintenance. Annexin A2 (ANXA2), a calcium-dependent phospholipids binding protein is highly expressed in HCC. To study the roles the excessively expressed ANXA2 during the progression of HCC, we inhibited the ANXA2 expression in SMMC-7721 cells using RNAi, followed by the analysis of cell growth, apoptosis and cell motility. To explore the relationship between the cell behaviors and its structures, the microstructure changes were observed under fluorescence microscopy, laser scanning confocal microscopy and electron microscopy. Our findings demonstrated that down-regulation of ANXA2 results in decreased the cell proliferation and motility, enhanced apoptosis, suppressed cell pseudopodia/filopodia, inhibited expression of F-actin and β-tubulin, and inhibited or depolymerized Lamin B. The cell contact inhibition was also analyzed in the paper. Take together, our results indicate that ANXA2 plays an important role to enhance the malignant behaviors of HCC cells, and the enhancement is closely based on its remodeling to cell structures.     

用于微流控液滴检测的共焦激光诱导荧光检测系统

以发射波长为473nm的二极管泵浦激光器为激发光源,搭建适于微流控液滴检测的共聚焦激光诱导荧光检测系统.系统玻璃芯片采用简易加工技术加工而成.并用2%十八烷基三氯硅烷将微通道处理为具有强疏水性质.荧光素钠溶液在T形通道交叉处被十四烷剪切形成液滴,液滴流经检测点时被激光诱导荧光系统检测.荧光素钠的检出限为1.1×10<'...     

用于钙原子光频标的半导体倍频激光器及荧光谱

利用BIBO(BiB3O6)晶体的倍频效应,由半导体激光器产生的波长为846 nm激光可以获得波长为423 nm的蓝光。真空室内的钙炉在加热到600 ℃时产生钙原子束。将423 nm激光垂直照射到钙原子束上,用光电探测器可以获得钙原子束的荧光谱,谱线的半峰全宽(FWHM)为100 MHz。