VLT spectroscopy of globular cluster in NGC 3115

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【天文学】第一讲_引言

【天文学】第一讲_引言
形态,内部结构,物理状态,化学组成,相互关系 起源,演化和死亡 ▲ 实测天体物理 天体的观测方法,观测结果
天体的位置及其变化 太阳系天体的运动规律 天体物理本质研究
▲ 理论天体物理 观测现象的解释
天体物理学: 天体物理本质研究
1. 太阳物理:太阳的本质,能源问题 2. 太阳系物理学(行星物理):
宇宙大尺度结构 致密天体,如中子星,白矮星,黑洞等 引力波,引力透镜
9. 宇宙学:宇宙的整体研究;观测和理论
大爆炸理论 宇宙大尺度结构的形成和演化
10. 天体演化学:太阳系,恒星,星系,宇宙
动力学演化,化学演化 化学元素的起源,宇宙中的分布,变化规律
天体物理学: 天体物理本质研究
从观测方法上分:
光学天文学 红外天文学 射电天文学 空间天文学 : 地面,空间 :地面,空间 :地面,空间 :空间探测器
厄尔尼诺现象与地球自转有关
亚洲 东太平洋
西太平洋
南美洲
天文与气象
地球绕太阳公转 黄道与赤道 存在23°交角 气候的四季变化
天文与气象
月球对地球 的引力作用 海水每天的 潮起潮落
问题二: 天文学有什么用 ?
天文学是一门古老的学科,是一门观测的科学, 在历史上它与人类的生产活动和日常生活密切 相关, 例如: 季节的变化, 潮水涨落, 野外方向的确定等等
3。天文学对哲学的意义 人类如何认识宇宙
地心说 日心说 大爆炸理论 太阳系的起源到宇宙的起源
4。天文学对工农业生产的作用
计量时间
星表,年历的编制 精密定轨, 测距
时间标准,用于尖端科学
农业生产,航海,航空,航天 卫星轨道
天文高灵敏度探测器
遥感, 军事

Molecular SpectroscopyVisible and Ultraviolet Spectroscopy

Molecular SpectroscopyVisible and Ultraviolet Spectroscopy
Total Spin Multiplicities
Orbital Spin States


For triplet state: Under the influence of external field, there are three values (i.e. 3 energy states) of +1, 0, -1 times the angular momentum. Such states are called triplet states (T). According to the selection rule, S→S, T→T, are allowed transitions, but S→T, T→S, are forbidden transitions.
UV and Visible Spectroscopy

In structure determination : UV-VIS spectroscopy is used to detect the presence of chromophores like dienes, aromatics, polyenes, and conjugated ketones, etc.

Instrumentation
光源
分光器樣品偵測器源自記錄器Components of a Spectrophotometer Light Source
Deuterium Lamps-a truly continuous spectrum in the ultraviolet region is produced by electrical excitation of deuterium at low pressure. (160nm~375nm) Tungsten Filament Lamps-the most common source of visible and near infrared radiation.

Vibrational Spectroscopy - NUI Galway National University of 振动光谱-高威国立大学

Vibrational Spectroscopy - NUI Galway National University of 振动光谱-高威国立大学

RAIRS examples
n(N-O) spectra obtained from a Pt
surface subjected to a fixed exposure of NO at various temperatures Note - the surface coverage of adsorbed NO molecules decreases as the temperature is raised and little NO remains adsorbed at temperatures of 450 K and above.
CO on Pt
Vibrational Spectroscopy
IR Spectroscopy (of various forms, e.g. RAIRS, ATR) Electron Energy Loss Spectroscopy ( EELS )
IR Spectroscopy
• Transmission IR Spectroscopy : studies where large surface area permits high concentration of adsorbed species. • Diffuse Reflectance IR Spectroscopy ( DRIFTS ) : diffusely scattered IR collected, refocused and analysed. Employed with high surface area samples not sufficiently transparent to be studied in transmission. • Reflection-Absorption IR Spectroscopy ( RAIRS ) : IR specularly reflected from front face of a highly-reflective sample, such as a metal single crystal surface. • Multiple Internal Reflection Spectroscopy ( MIR ) : IR passed through IR transmitting sample and undergoes total internal reflection. IR radiation may be absorbed by species adsorbed on the solid surface - alternative name of Attenuated Total Reflection (ATR).

211126668_超声波辅助酶法提取牛皮胶原蛋白及其结构表征

211126668_超声波辅助酶法提取牛皮胶原蛋白及其结构表征

赵改名,王壮壮,祝超智,等. 超声波辅助酶法提取牛皮胶原蛋白及其结构表征[J]. 食品工业科技,2023,44(9):190−199. doi:10.13386/j.issn1002-0306.2022070219ZHAO Gaiming, WANG Zhuangzhuang, ZHU Chaozhi, et al. Ultrasound-Assisted Enzymatic Extraction and Structural Characterization of Cowhide Collagen[J]. Science and Technology of Food Industry, 2023, 44(9): 190−199. (in Chinese with English abstract). doi: 10.13386/j.issn1002-0306.2022070219· 工艺技术 ·超声波辅助酶法提取牛皮胶原蛋白及其结构表征赵改名1,王壮壮1,祝超智1, *,余小领1,张秋会1,祁兴山2(1.河南农业大学食品科学技术学院,河南郑州 450002;2.恒都综合试验站,河南驻马店 463000)摘 要:以牛皮为原料,优化超声波辅助酶提取牛皮中胶原蛋白的工艺。

在单因素实验的基础上设计响应面试验,以牛皮胶原蛋白提取率为响应值,优化得到胶原蛋白的最佳提取工艺,并对其进行结构表征。

结果表明:牛皮中胶原蛋白的最佳提取工艺条件为超声波功率161 W 、超声波处理时间64 min 、胃蛋白酶添加量109 U/g 、料液比1:16 g/mL ,在此条件下胶原蛋白的提取率为63.77%。

十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE )、紫外光谱(UV )和傅立叶红外光谱(FTIR )分析表明,超声波辅助酶提取的胶原蛋白符合Ⅰ型胶原蛋白的特征,保持了其完整的三螺旋结构,氨基酸组成和扫描电镜(SEM )分析得到超声波辅助酶提取的胶原蛋白三螺旋稳定程度略微下降。

【材料研究方法】光谱分析(英文)

【材料研究方法】光谱分析(英文)

Vibrational spectroscopy6.3.1 Fundamentals of vibrational spectroscopyDefinition Vibrational spectroscopy:is concerned with the d t ti f t iti detection of transitions between energy levels in molecules that result from stretching and bending vibrations of the interatomic bonds.asymmetricalVibrational spectroscopyKinds of vibrational spectroscopy ¾Infra-red spectroscopy(more sensitive to polarized group)6.3.1 Fundamentals of vibrational spectroscopysymmetrical¾Raman spectroscopy (moresensitive to non-polarized)Both methods are concerned with vibrations in molecules , they differ in the manner in which interaction with the exciting radiation occurs .Linear PE: (a) IR, (b)RamanFig. 6-14 Dipole moment of HClVibrational spectroscopyVibrating of Disulfide carbonSymmetrical stretchingInfrared inactive 6.3.2 Infrared spectroscopyAsymmetrical stretchingBendingInfrared activeInfrared inactive Fig. 6-15 Vibration of Disulfide carbonm1lowHigh/cm-1High/cm-1lowVibrational spectroscopy Methylbenzene(甲苯)2005.2 S. Guv =0 represents the ground state v =l the excited vibrational state6.3.3 Raman spectroscopy(1)(2)(3)Vibrational spectroscopy ¾The essential prerequisite for Raman scattering is a change in the polarizability of the bond when vibrations occur.Polarizability may be thought of as a measure of 6.3.3 Raman spectroscopy¾Polarizability may be thought of as a measure of theFig. 6-16 Motion state of linear molecules Degrees of freedom (H2O) : 3×3−6 = 3Vibraitonal modes (methylene group):2926cm-1(s)asνAsymmetricalsν: 2853 cm Symmetricalδ:1468 cm-1(m) δr:720 cm-1(CH1306~1303cm-1(w)γt :1250cmscissoring rocking waggingHexaneFour peakspSpectral interpretation always starts at the high end, because there are the best group frequencies and they are the easiest to interpret. No peaks appear above 3000 cm-1, the cut-off for unsaturated C-H. the four peaks below 3000 cm-1 are saturated C-H stretching modes.HexaneThe peak at 2962 cm-1 isassigned to the antisymmetricassigned to the antisymmetricstretch of the CH3group. Thisvibration is always found inthe range 2962±10 cm-1. thereare actually two degenerateantisymmetric stretchingmodes (only one shown).HexaneAt 2926cm-1, the CH2antisymmetric stretchabsorbs.Normal range:2926±10 cm-1.HexaneAt 2872cm-1, the CH3symmetric stretchabsorbs.Normal range:2872±10 cm-1.HexaneAt2853-1,the CHAt 2853cm, the CH2symmetric stretchabsorbs.Normal range:2853±10 cm-1.Vibrational spectroscopy Hexane1470cm-1This is the C-H bendingregion, expanded to show thenearly overlapping peaks forthe CH3and CH2bends.Vibrational spectroscopyHexanerocking When four or more CH2groups arein a chain, a vibration at 720±10cm-1corresponds to concertedrocking of all of the CH2’s.Vibrational spectroscopyHexanol3334 cm-1–OH stretch. Normal range: 3350±150 cm-1.This is a very characteristic group frequency. All of thepeaks due to the OH group are broad due to hydrogenbonding.Vibrational spectroscopy Hexanol 1430 cm -1–OH bend . Normal range: 1400±100 cm -1. This broad peak is buried under the CH bending modes.Vibrational spectroscopyHexanol660 cm -1–OH wag. While not a group frequency, this is another band due to the OH.Vibrational spectroscopy Aromatic ring expansion (Methylbenzene )At 1601 cm -1, thesymmetric ring strethch absorbs. Normal range: 1590±10 cm -1. This ib ti h di lOnly notsymmetrically substituted.vibration has a dipole change (and absords in IR) only when notsymmetrically substituted. The intensity of this band also varies with thesubstituent. Compare to p-xylene from the overlay menu.Vibrational spectroscopyAromatic ring expansion (Methylbenzene )At 1500cm -1, a different ring stretch absorbs. Range: 1500±10cm -1. Variable intensityVibrational spectroscopy 6.3.6 Comparing of IR and Raman SpectroscopyasymmetricalsymmetricalFig. 6-17 Linear PE: (a) IR, (b) Raman。

Vibrational spectroscopy振动光谱

Vibrational spectroscopy振动光谱

0
0 incidence angle 90
s-polarized light at the surface - uniform phase shift - vanishing E field at the surface
p-polarized light at the surface
Absorbance is proportional to
a large reflectivity change at high incidence angle
Surface selection rule
mfi = <f|m|i> 0, dm/dr 0
-The electric field of light and the molecule interact with surface electrons -The incident light must be p-polarized -Only vibrations with a dipole moment perpendicular to the surface -The incident light should be reflected at grazing incidence
mM +-
-+ mimage
+ mM
-
-
+
mimage
IR inactive
IR active
- for lying down molecules, molecular and image dipoles are cancelled out
- for upright molecules, molecular and image dipoles are enhanced

变异链球菌的VicRK双组分信号传导系统

变异链球菌的VicRK双组分信号传导系统

变异链球菌的VicRK双组分信号传导系统变异链球菌是人类龋病的主要病原菌,它通过蔗糖依赖性黏附形成生物膜并在其中产酸耐酸,最终导致龋病。

VicRK是变异链球菌13种双组分信号传导系统之一,可调节变异链球菌致龋性毒力相关因子的表达。

本文就VicRK的作用机制、结构组成、生理特性,及其对变异链球菌致龋性的影响,VicRK和VicX 间的关系等研究进展作一综述。

标签:变异链球菌;双组分信号传导系统;基因表达;VicRKVicRK two-component signal transduction system of Streptococcus mutansTian Yuanyuan, Hu Tao.(State Key Laboratory of Oral Diseases, Sichuan University, Chengdu 610041, China)[Abstract]Streptococcus mutans(S.mutans), which is considered as the chief pathogen of human caries, possesses the ability to form biofilm via sucrose-dependent adhesion, genesis and endure acids in the biofilm that may ultimately lead to dental caries. VicRK is one of the 13 putative two-component signal transduction systems of S.mutans that modulate the expression of cariogeneisis related virulence factors. This review summarized the mechanism, structural organization, physiological characteristics and the impact on the cariogenesis capabilities of VicRK,as well as the correlation between VicRK and VicX in S.mutans.[Key words]Streptococcus mutans;two-component signal transduction system;gene expression;VicRK2005年,Ulrich等[1]对145种细菌基因组进行了测序,在此过程中他们发现了至少 4 000对双组分信号传导系统(two-component signal transduction system,TCSTS)。

Broad Band Dielectric Spectroscopy -williams

Broad Band Dielectric Spectroscopy -williams
2-D Cartoon: Penrose Tiling of a mesoscopic region No symmetry, so it is a disordered system Paint in the energies of the small regions at t = 0
Snapshot of an ensemble at time t = 0 The local regions span the range of dynamical behaviour - giving slow to fast relaxations Motions occur in the local regions; continual exchange of energies between the regions changes their dynamics continually
ε (ω ) =
σ (ω ) 1 = M (ω ) iωε v
C0
Broadband Dielectric Spectroscopy
Complex Permittivity
ε = ε '−iε ' ' ε '= ε∞ +
∆ε 1 + ω 2τ 2
ε ''=
log f/Hz 1. Relaxation Strength ∆ε 3. Shape of process Low frequencies
Electrical Properties of Polymers
E Riande & R Diaz-Calleja, Marcel Dekker 2004
Introductory Review
G Williams & DK Thomas, Novocontrol Application Notes, No.3, 1998.

Vibrational_Spectroscopy_I

Vibrational_Spectroscopy_I

10
Vibrational Spectrum of HCl
Vibrational spectrum of HCl is based on the harmonic oscillator model with ωe = 2989 cm-1.
v
ν (cm-1)
11
Vibrational Spectrum of HCl
• Exceptions to the infrared ΔJ ≠ 0 selection rule are found for some diatomic molecules such as NO.
30
Vib-Rot Infrared Spectrum of the DCl Molecule
• νvib(HCl) > νvib(DCl) because of the differences in force constants and reduced massed between the two molecules. • B0 = 5.392263 cm-1 B1 = 5.279890 cm-1
12
Vibrational Spectrum of HCl
13
Vibrational Spectrum of HCl
14
Morse Potential
15
Morse Potential
• It is a better approximation for the vibrational structure of the molecule than the quantum harmonic oscillator because it explicitly includes the effects of bond breaking, such as the existence of unbound states.

天文术语 G-I

天文术语 G-I
high-metallicity cluster 高金属度星团;
高金属度星系团
high-resolution spectrograph 高分辨摄谱仪
high-resolution spectroscopy 高分辨分光
high - z 大红移
Hinotori 〈火鸟〉太阳探测器
Hipparcos, High Precision Parallax 〈依巴谷〉卫星
impulsive solar flare 脉冲太阳耀斑
infralateral arc 外侧晕弧
infrared CCD 红外 CCD
infrared corona 红外冕
infrared helioseismology 红外日震学
infrared index 红外
infrared observatory 红外天文台
ISO, Infrared Space Observatory 〈ISO〉红外空间天文台
isochrone method 等龄线法
IUE, International Ultraviolet 〈IUE〉国际紫外探测器(责任编辑:admin)
IRPS, Infrared Photometer / Spectro- 红外光度计/分光计
meter
irregular cluster 不规则星团; 不规则星系团
IRTF, NASA Infrared Telescope 〈IRTF〉美国宇航局红外
Facility 望远镜
IRTS, Infrared Telescope in Space 〈IRTS〉空间红外望远镜
intranetwork element 网内元

POE-g-MAH_对PLA

POE-g-MAH_对PLA

第4期71李梦云,等:四环素和铜离子对生物除磷中微生物胞外聚合物的影响合物中蛋白质和多糖增加量三种配比下相对最高;四环素和铜离子配比为0.894时,四环素与铜离子浓度相当,混合物投加初期胞外聚合物中蛋白质和多糖增加量三种配比下相对最低。

三种配比混合物作用下,随混合物浓度增加,微生物胞外聚合物中蛋白质三维荧光强度逐渐减弱。

参考文献:[1] 李金璞,张雯雯,杨新萍.活性污泥污水处理系统中胞外多聚物的作用及提取方法[J].生态学杂志,2018, 37(9):2825-2833.[2] Mohite B V,Koli S H,Patil S V. Heavy metal stress and its consequences on exopolysaccharide (EPS)-producing pantoea agglomerans[J].Applied Biochemistry and Biotechnology,2018,186(1):199-216.[3] Grabert R,Boopathy R,Nathaniel R,et al. Effect of tetracycline on ammonia and carbon removal by the facultative bacteria in the anaerobic digester of a sewage treatment plant[J].Bioresource Technology,2018,267:265-270.[4] Li J Y,Du Q P,Peng H Q,et al. Spectroscopic investigation of the interaction between extracellular polymeric substances and tetracycline during sorption onto anaerobic ammonium-oxidising sludge[J].Environmental Technology,2021,42(11):1787-1797.[5] Li W W,Yu H Q. Insight into the roles of microbial extracellular polymer substances in metal biosorption[J].Bioresource Technology,2014,160:15-23.[6] Li C X,Xie S Y,Wang Y,et al. Simultaneous heavy metal immobilization and antibiotics removal during synergetic treatment of sewage sludge and pig manure[J].Environmental Science and Pollution Research,2020,27(24):30323-30332.[7] Zhang Y H,Liu S S,Liu H L,et al. Evaluation of the combined toxicity of 15 pesticides by uniform design[J].Pest Management Science,2010,66(8):879-887.[8] Abbondanzi F,Cachada A,Campisi T,et al. Optimisation of a microbial bioassay for contaminated soil monitoring: bacterial inoculum standardisation and comparison with Microtox ® assay[J].Chemosphere,2003,53(8):889-897.[9] 陶梦婷,张瑾,姜慧,等.3种农药对青海弧菌Q67的联合毒性作用特征[J].环境科学与技术,2019,42(6):12-20.[10] Sutherland I W. Biofilm exopolysaccharides:a strong and sticky framework[J].Microbiology,2001,147(1):3-9. L1-3* L1-5* L1-7* L1-9* L2-3* L2-5* L2-7* L2-9* L3-3* L3-5* L3-7* L3-9*图4 三种浓度配比混合物作用下生物除磷中微生物胞外聚合物的三维荧光光谱图(下转第91页)第30卷第4期2022年8月V ol.30 No.4Aug.2022安徽建筑大学学报Journal of Anhui Jianzhu UniversityDOI:10.11921/j.issn.2095-8382.20220412POE-g-MAH对PLA/PP共混材料界面状态及机械性能的影响陈鑫亮1,高 尚1,樊炳宇1,刘 瑾1,2,王 平1,2(1.安徽建筑大学 材料与化学工程学院,安徽 合肥 230601;2.安徽省先进建筑材料国际联合研究中心,安徽 合肥 230601)摘‌要:聚乳酸(PLA)与聚丙烯(PP)的相容性较差,界面相互作用较弱,导致PLA/PP共混材料的力学性能较差。

UV-vis Spectroscopy and Cyclic Voltammetry Investigations of Tubular JAggregates

UV-vis Spectroscopy and Cyclic Voltammetry Investigations of Tubular JAggregates

10.1149/1.3104058 © The Electrochemical SocietyUV-vis Spectroscopy and Cyclic Voltammetry Investigations of Tubular J-Aggregates of Amphiphilic Cyanine DyesJ. L. Lyon a,c, D. M. Eisele b, S. Kirstein b, J. P. Rabe b,D. A. Vanden Bout a, and K. J. Stevenson aa Department of Chemistry, University of Texas at Austin, Austin, TX 78712, USAb Department of Physics, Humboldt University, D-12489 Berlin, Germanyc Current Affiliation: Office of Biotechnology, The University of Texas Health ScienceCenter at Houston, Houston, TX 77030, USAThe redox and light absorption properties of immobilizedaggregates of the cyanine dye 3,3'-bis(2-sulfopropyl)-5,5',6,6'-tetrachloro-1,1'-dioctylbenzimidacarbocyanine (C8S3) ontransparent, conductive indium tin oxide (ITO) electrodes havebeen studied directly using cyclic voltammetry (CV) inconjunction with UV-vis spectroscopy to elucidate uniquemechanistic features of J-aggregate oxidation. C8S3 self-assemblesin aqueous solution to form double-walled, tubular J-aggregateswith ~13 nm diameter and lengths up to several hundrednanometers. Spectroelectrochemical measurements indicate thatirreversible J-aggregate oxidation occurs primarily along the outerwall of the tubular structure as evidenced by the potential-inducedirreversible bleaching of J-band absorption. Additionally, J-aggregate oxidation involves both electrochemical and chemicalsteps in which dimerization and subsequent dehydrogenation of theJ-aggregate leads to the formation of a new dehydrogenated dimeroxidation product.IntroductionTubular J-aggregates of amphiphilic carbocyanine dyes have drawn interest as a model system to study the efficient excitonic energy migration properties similar to those in Light Harvesting Systems (LHS) of plants and bacteria (1, 2). In particular, the double-walled tubular structure of the cyanine dye 3,3'-bis(2-sulfopropyl)-5,5',6,6'-tetrachloro-1,1'-dioctylbenzimidacarbocyanine (C8S3) J-aggregates is an extremely attractive supramolecular model for an artificial LH S due to its ordered, quasi-one-dimensional structure that allows investigation of energy migration and charge transfer processes.H owever, a general problem of aqueous based J-aggregate systems is oxidation, especially in the presence of metal cations such as Ag+ where selective oxidation of the J-aggregate has been observed to occur along with the formation of Ag nanoparticles along the outer wall of the tube (3). Still, the exact mechanism of particle nucleation at the J-aggregate, and its effect on the intrinsic energy migration properties of the aggregate, is poorly understood. For future applications in the context of artificial LH S, more knowledge about the oxidation process is necessary to prevent or control the destructive oxidation of the light harvesting antenna system.H erein, we report electrochemical and spectroscopic properties of immobilized J-aggregates on transparent, conductive indium tin oxide (ITO) electrodes using cyclic voltammetry (CV) in conjunction with UV-vis spectroscopy to elucidate uniquemechanistic features of J-aggregate oxidation. This work contributes to the limited number of electrochemical studies of immobilized cyanine dye J-aggregates in aqueous solution (4,5).ExperimentalC8S3 J-aggregate PreparationThe amphiphilic cyanine dye 3,3’ bis(2 sulfopropyl) 5,5’,6,6’ tetrachloro 1,1’ dioctylbenzimidacarbocyanine (C8S3) is available as a sodium salt from FEW Chemicals (Wolfen, Germany) and was used as received. A 3.00 mM stock solution of monomeric C8S3 was prepared by dissolving an appropriate amount of C8S3 (MW = 902.8 g/mol) in pure methanol (Fisher Scientific) by stirring to form a clear red solution. To prepare C8S3 J aggregates, 130 ȝL of the C8S3 stock solution was added to 500 ȝL of ultrapure H2O (>18.2 Mȍ cm, Barnstead) and agitated to ensure even mixing. An immediate color change from clear red to opalescent, bright pink was observed by eye, indicating the formation of double-walled tubular C8S3 aggregates (6). The solution was stored in the dark for 24 h before adding an additional 500 ȝL H2O to stabilize the aggregation process, resulting in a final dye concentration of 3.36 x 10-4 M. Solutions of J-aggregates were typically used for experiments within five days of preparation, and stored in the dark when not in use. Absorption measurements of C8S3 monomer and J-aggregate solutions were collected using an Agilent Instruments 8453 UV-Visible spectrometer with a photodiode array detector. The measurement cell consisted of two quartz slides with a fixed Teflon gasket (0.1 mm path length) containing 45 ȝL of either monomer or J-aggregate solution.Electrochemical MeasurementsElectrochemical experiments were conducted on J-aggregates immobilized on transparent, conductive indium tin oxide (ITO)-coated glass electrodes (Delta Technologies, Ltd.; 15 ȍ/Ƒ) immersed in an aqueous solution containing 1 M KNO3 supporting electrolyte (Fisher Scientific). The pH of this supporting electrolyte was 5.78 ± 0.03 as prepared. Prior to use, the ITO working electrode substrates were cleaned by immersion in 30% (v/v) aqueous ethanolamine (Aldrich) at 80 °C for 20 min, followed by rinsing with methanol and sonicating in ultrapure H2O for 30 min. The substrates were then dried under a stream of nitrogen. Films of J aggregates were prepared on ITO by drop casting 10 ȝL of J aggregate solution on a 0.5 cm2 area of ITO followed by drying in the dark for ~ 2 h. Once dried, they remained at the ITO surface when immersed in supporting electrolyte. ITO working electrodes, either bare or with dried J aggregate films, were fitted in a 20 mL glass cell containing ~5 mL supporting electrolyte, Au wire counter electrode, and a Hg/Hg2SO4 (sat’d. K2SO4) reference electrode (CH Instruments; E° = +0.640 V vs. NHE) with the ITO surface oriented perpendicular to the bottom of the cell. Electrochemical measurements were performed at room temperature (23 ± 2 °C) using an Autolab PGSTAT30 potentiostat interfaced with Autolab GPES version 4.9 software. Prior to each experiment, the cell was purged with Ar for at least 5 min. All experiments were conducted under flowing Ar.Spectroelectrochemical MeasurementsSpectroelectrochemical experiments were performed using a CH700 bipotentiostat (CH Instruments) interfaced to an Agilent Instruments 8453 UV-Visible spectrometer with a photodiode array detector. A homemade electrochemical cell with a fixed working electrode area of 0.45 cm2 and ~1 mL 1 M KNO3 supporting electrolyte was placed in the spectrometer sample holder with the J-aggregate film-coated ITO working electrode in the beam path. The cell also contained a Pt wire counter electrode and H g/H g2SO4 (sat’d. K2SO4) reference electrode. The potential was cycled between -0.7 and +0.45 V for three consecutive cycles while absorption spectra between 350 and 800 nm were collected every 4.0 s with a 0.5 s integration time. A slow scan rate of 0.01 V/s was used, permitting spectral acquisition every 45 mV.Results and DiscussionC8S3 self-assembles in aqueous solution in the presence of 10 wt % methanol to form double-walled, tubular J-aggregates with ~8 nm inner diameter and lengths up to several hundred nanometers (6). The aggregation of the dyes produces new visible absorption bands that result from the dipolar coupling of the cyanine chromophores. As shown in Figure 1, the key features of the spectrum are two sharp J-aggregate bands at 590 and 600 nm that have been assigned to absorption by the outer and inner wall, respectively. Since each of the tubes has a distinct absorption feature, it is possible to follow the redox chemistry of the inner and outer wall independently. This double walled tubular structure has been confirmed by cryogenic TEM, polarized spectroscopy and theory (6). The morphology of the tubules is strongly dependent on the solvent conditions and a variety of other structures can be obtained by altering the chemical composition of the solvent (7).Figure 1. UV-Vis spectrum of the C8S3 monomer and tubular aggregate showing the inner and outer wall J-bands that form upon aggregation.Figure 2 depicts cyclic voltammograms (CVs) showing the irreversible oxidation of immobilized J-aggregates at ITO in aqueous pH 5.78, 1 M KNO3. The J-aggregates are significantly easier to oxidize as evidenced by the appearance of an irreversible J-aggregate oxidation (labeled I, black trace) as the potential is scanned positive. A single oxidation peak for the J-aggregate appears at +0.22 V compared to that reported for the monomer at +1.04 V (8) and consistent with those reported for other J-aggregate systems (4), in which a lower oxidation potential is reflective of a lower activation energy for J-aggregate oxidation relative to the cyanine monomer due to a shift in HOMO-LUMOenergy levels upon aggregation. Subsequent scans between positive and negative potential extremes reveal a second, reversible set of peaks centered at E1/2= -0.378 V, labeled II/II’. Note that the current response for this new reduction/oxidation process continues to increase until a steady state response is obtained after ~15 consecutive cycles. Additionally, voltammetric studies were conducted in which the potential window of the immobilized J-aggregate ITO electrode was cycled 15 times without inducing J-aggregate oxidation (i.e. from the same negative extreme as that shown in Figure 2 to +0.025 V, just before the onset of J-aggregate oxidation). In this case, we do not see the appearance of the redox response for II/II’. However, if the potential was then scanned positive (anodically) to +0.45 V to induce J-aggregate oxidation, the reduction peak II’ appears on the return sweep. This data indicates that peaks II/II’ are indeed related to the formation of a new species as a result of J-aggregate oxidation; this species is most likely a J-aggregate oxidation product whose redox activity behaves in accordance with a previously proposed electrohydrodimerization (EHD) mechanism (9).Figure 2. Cyclic voltammograms (15 cycles) displaying the initial irreversible oxidation of the J-aggregate (peak I) and the formation of a second, reversible couple, (peaks II/II’), during potential cycling of a J-aggregate/ ITO electrode in pH 5.78, 1 M KNO3. Scan rate 0.1 V/s.In a previous study (8) we conducted several other pH-dependent measurements to understand this complicated, proton-coupled electron transfer process. Scheme 1 depicts our proposed potential-dependent mechanism of C8S3 J-aggregate oxidation and subsequent formation of other redox active products, which support an electrochemical-chemical-electrochemical (ECE) process that consists of a net 4-electron transfer per dye molecule to yield a dehydrogenated dimer, labeled as DHD ox4+.Scheme 1. Depiction of the overall EH D mechanism involving ECE steps for C8S3 J-aggregate oxidation and subsequent formation of a dehydrogenated dimer product (DHD).We note that while the DH D ox4+ formation mechanism is derived from studies of cyanine dye monomers, it may be even more probable in the case of J-aggregates, in which monomer units of dye are spatially arranged near each other, promoting dimer associations of oxidized dye and the subsequent formation of DH D ox4+. If the species responsible for II/II’ in Figure 2 is a DH D ox4+product formed from neighboring dye molecules in the immobilized J-aggregates at ITO, then its voltammetric behavior should follow that predicted for a surface-confined redox species. As reported previously (8), the peak currents for both II and II’ at steady state do, in fact, scale linearly with increasing scan rate between 0.01 and 3 V/s, as predicted for an immobilized system.To correlate the electrochemical oxidation of C8S3 J-aggregates with their well-defined absorption spectrum, spectroelectrochemical experiments were carried out in which the potential of an ITO working electrode containing a J-aggregate film was scanned concurrently with the acquisition of absorbance spectra in the visible region from 350-800 nm. Similar studies have been undertaken previously to investigate J-aggregates organized on modified Au(111) (5). To maximize the number of spectra acquired for correlation to the CV, a slow scan rate of 0.01 V/s was used, permitting spectral acquisition every 45 mV. Figure 3 depicts the decrease in absorbance observed upon electrochemical oxidation of the J-aggregate film. As more anodic potentials are applied, the absorbance at 590 nm corresponding to the outer wall of the J-aggregate tubular structure decreases dramatically. The inner wall absorbance at 600 nm also decreases, though not as substantially as that of the outer wall; the relative intensity of the inner wall in fact surpasses that of the outer wall as a result of electrochemical oxidation. This change in relative absorbance most likely reflects a more facile oxidation of the outer wall, due to outer wall dye molecules being in closer electrical contact with the electrode, along with a more hindered oxidation of the inner wall dye molecules due to their occlusion by the C8 chains, and a possible charge compensation hindrance due to slower ion transport within the interior of the tube.Figure 3. Spectroelectrochemical data for the oxidation of immobilized C8S3 J-aggregates on ITO: (a) absorption spectra versus applied potential, (b) cyclic voltammogram at 0.01 V/s in pH 5.78, 1 M KNO3, (c) differential absorbance (įA/įV) plot for outer wall and inner wall J-aggregate absorbance bands centered at 590 and 600 nm, respectively.Importantly, the absorption maxima do not shift as a function of applied potential, but rather remain fixed at 590 and 600 nm, respectively. Previous spectroscopic studies by Lenhard and H ein correlated wavelength shifts with disruption of the J-aggregate dyeupon chemical oxidation (10); however, these results suggest that the J-aggregates’ morphology is not disturbed as a result of electrochemical oxidation. The differential absorbance (įA/įV) plots for both inner and outer wall oxidation correlate well with the voltammetric response shown in Figure 3c. The įA/įV plots nearly perfectly overlay the CV data, showing the direct correlation between electrochemical oxidation of the J-aggregates and changes in the J-band absorbance. At this slower scan rate, the J-aggregate oxidation peak in Figure 3b features a shoulder near +0.230 V in addition to the sharp oxidation at +0.166 V. Closer inspection of Figure 3c reveals that the inner wall of the J-aggregate structure passes through a maximum in differential absorbance a few seconds after the outer wall does, and that the resulting differential absorbance plot very closely resembles the shouldered peak in Figure 3b. On the return sweep of the voltammogram shown in Figure 3b, the reduction peak II’ corresponding to the reduction of DH D ox4+occurs at -0.378 V, and a response in the įA/įV plots is observed concurrently. Assuming that only the absorbance bands centered at 590 and 600 nm are present in spectra during the course of this experiment, this data suggests that the J-band absorbance is increasing as DH D ox4+is reduced. H owever, examination of the full absorption spectrum reveals a new broad absorption generated upon reduction of II’ that overlaps with the aggregate peaks. This absorption correlates to the formation of DHD red2+.Our spectroelectrochemical studies are also consistent with spectral changes observed in the reaction of J-aggregates in the presence of Ag+. The kinetics of the chemistry can be directly monitored because the effects of the silver growth can be clearly seen in the absorption spectra before and after particle formation, Figure 4.Figure 4. UV-Vis spectrum of a C8S3 aggregate solution after the addition of Ag(NO3). Initial spectrum (black), spectrum with Ag(NO3) after 3 hours in the dark (gray), spectrum with Ag(NO3) after 15 minutes with 5 minutes of illumination (orange). Inset: red arrow indicates the presence of Ag plasmon absorbance near 410 nm.The formation of the silver particles results in the loss of absorption from the aggregates and an increase in absorption due to a silver plasmon. The decay of the inner and outer wall absorptions differ from one another with the outer wall decaying nearly to zero while some portion of the inner wall remains after the chemistry is complete. However, at this stage the resulting effect of photo-oxidation upon the aggregates’ morphology remains unclear, as well as the mechanistic details of particle nucleation and growth.Last, we also performed additional experiments in which a solution of J-aggregates had been illuminated with UV-light in the presence of Ag+. This solution was then evaporated on an ITO electrode. Cyclic voltammetry was subsequently conducted on the J-aggregate film in pH 5.78, 1 M KNO3. As shown in Figure 5, a distinct anodic response associated with the oxidation of Ag0 is observed at +0.03 V, as well as a diminished oxidative response at +0.2 V associated with the residual oxidation of un-reacted J-aggregates. Integration of the Ag oxidation peak gives 9.2x10-7 C, which correlates to ~95 pmol Ag. We also tried to illuminate a dried J-aggregate film in the presence of Ag+ while in the spectroelectrochemical cell, but the UV-Vis spectrum showed no decrease in absorbance at 590 or 600 nm. However, these preliminary experiments indicate that the J-aggregate acts as a reducing agent in the presence of Ag+ which leads to nucleation and growth of Ag nanoparticles at outer wall sites along the J-aggregate. More studies are necessary to understand how the J-aggregates act as a localized photoreductant to facilitate and confine the reduction of metal ions.Figure 5. Cyclic voltammogram of immobilized J-aggregates on an ITO electrode prepared from UV illumination of a J-aggregate solution containing Ag+.SummaryThe spectroelectrochemical studies of tubular J-aggregates immobilized at ITO electrodes has been described. The electrochemical behavior of C8S3 J-aggregates is comparable to that observed in previous studies performed on solution-phase J-aggregates and cyanine dye monomers. An electrohydrodimerization mechanism was proposed to explain the unique potential dependent redox response. Both irreversible dimerization and dehydrogenation chemical steps follow the initial J-aggregate oxidation, which leads to the formation of a new dehydrogenated dimer oxidation product that is most likely confined within the J-aggregate itself. UV-vis spectroscopy studies demonstrate that the J-aggregate oxidation is mostly confined to the outer wall of the tubular structure, and that the inner wall is oxidized to a lesser extent relative to the outer wall. Future studies involve the development of a model of all the species involved in the redox chemistry and verify this model with spectral measurements acquired in the absence and presence of metal salts (chemical oxidants) and with and without illumination to UV light.AcknowledgmentsFinancial support for this work was provided by the R.A. Welch Foundation (grants F-1377 and F-1529), the National Science Foundation (CH E-0809770), and Deutsche Forschungsgemeinschaft [Sfb 448: Mesoscopically Organized Composites].ReferencesMcDermott et al.,Nature 374, 517 (1995).1. G.2. H. v. Amerongen, L. Valkunas, R. v. Grondelle, in Photosynthetic Excitons.(World Scientific, Singapore, 2000).3. S. Kirstein, H. v. Berlepsch, C. Böttcher, Int. J. Photoener. 2006 (47917), 1(2006).4. M. Kawasaki, T. Sato, J. Phys. Chem. B 105, 796 (2001).5. M. Kawasaki, D. Yoshidome, T. Sato, M. Iwasaki, J. Electroanal. Chem. 543, 1(2003).Didraga et al.,J. Phys. Chem. B 108, 14976 (2004).6. C.7. H. von Berlepsch, S. Kirstein, R. Hania, A. Pugzlys, C. Böttcher, J. Phys. Chem.B 111, 1701 (2007).8. J. L. Lyon et al.,J. Phys. Chem. C 112, 1260 (2008).9. J. R. Lenhard, A. D. Cameron, J. Phys. Chem. 97, 4916 (1993).10. J. R. Lenhard, B. R. Hein, J. Phys. Chem. 100, 17287 (1996).。

【doc】一氧化氮合成酶(NOS)基因表达的半定量检测及其运用

【doc】一氧化氮合成酶(NOS)基因表达的半定量检测及其运用

一氧化氮合成酶(NOS)基因表达的半定量检测及其运用4生理,1994,4B(4),347354ActaPhysiologicaSinica一氧化氮合成酶(NOS)基因表达的半定量检测及其运用1张晨晖李肯虹,继峰周洪张新波场健r_元j五吾础研究所分子生物学研究室北京I.口舶3)-三Jl摘要'/4'本工作参照Mi~lin(1993)定量RTPeR方法,建立了一种灵敏,简捷,特异的定量NOSmRNA测定方法}证明了NOSmRNA不仅存在于脑组织,亦广泛分布于心,肾,肺和肝组织中,其中以脑组织含量最高,肾,心次之.除内皮细胞以外,平滑肌细胞中亦有NOS基因的表达}此外,本工作还观察到,自发性高血压大鼠的脑,肾,肝和平精肌细胞中NOSmRNA水平下降,提示NOS基因表达受抑,可能与高血压的病国密切相关.f上一,关键词:.=墨些窒盛堕差里室垄自发性高血压太最;半定量pcRzfl兰,-近年来的研究证明,一氧化氮(nitricoxide,NO)是一种血管内皮细胞释放的内皮衍化舒张因子(endothelium—delvedrelaxingfactors,EDRFs),它具有舒张血管,降低血压,抑制血管平滑肌细胞增殖和血小板粘附等重要的生理作用,在高血压,心肌缺血等许多心血管疾病的发病中具有重要意义.NO是由L广精氨酸(L—ARG)和分子氧在一氧化氮合成酶(nitricoxidesynthase,NOS)催化下合成的,NOS则是NO生成的关键酶.最近,NOS cDNA已经在大鼠内皮细胞,小脑中克隆出来0一:.但是关于它在体内不同组织的分布以及在心血管疾病高血压发病中的作用,迄今还未见报道.本工作应用聚合酶链式反应(po]y—merasechainreaction,PCR)技术建立了一种新的,简捷,特异的定量NOSmRNA的测定方法,研究了NOS在大鼠体内的分布,发现自发性高血压大鼠(spontaneoushyl~rtenslverat,SHR)脑,肾和肝中NOSmRNA的表达明显降低,其内皮依赖性的血管舒张反应亦明显下降提示,N0S基因表达下降,可能与高血压的病因密切相关.1材料和方法动物本工作选用8—1o周龄的雄性SHR和WKY(Wistar-Kyotorat,WKY)大鼠进行实验,SHR大鼠由Okamoto等选育成功,其特点为1oo%高血压自发率(BP=22.7士1.3kPa),同时具有高血压性心血管病变,适用于人类高血压病的研究,WKY为其正常对照模型,实验所用动物由中国医学科学院心血管研究中心提供车文19日3年6月8日收到1993年9月12日修回生理46卷试剂本工作所用的反转录酶(MoMLV)和TaqDNA聚台酶分别购自美国的Stret- gene公司和Perkin—Klmer公司,[P]dCTP为北京福瑞公司产品(放射比活度为3O00Ci/mmo1)RandomPrimer和dNTP购自美国Promega公司,其它试剂均购自美国Sigma 公司.细胞培养取用6—8周龄雄性SHR,WKY大鼠和新生小牛的胸主动脉段,分别按Hofman和Booyse的方法培养血管平滑肌细胞和内皮细胞.本实验选用5—8代细胞进行实验.总RNA的制备取10的平滑肌细胞或内皮细胞,用RNAzolBRNA提取试剂盒(美国TEL—TEST公司)提取细胞总RNA.各取1g脑,心,肺,肾,肝组织采用异硫氰酸胍一酚一氯仿方法:提取组织总RNA.总RNA提取完成后,分别用紫外分光光度计测定总RNA的浓度,重复测定三次.oD:OD.比值在1.8—2.0之间.计算样品总RNA的浓度.反转录(reversetranscription,RT)取2lag总RNA,在65℃变性3min,依次加入0.5mmol/LdNTP,0.01mol/LDTT,10pmol/LRandomPr/mer,15unit反转录酶和oH 8.350mmot/LTris—HCI,75mmol/LKCI,3mmol/LM鲁cl2缓冲液,总反应体积为201.在37℃保温90min.反应结束后,加入50TE缓冲液(pH8.010mmol/LTris—HCI,1mmol/ LEDTA),于70℃加热10min以终止反应定量PCRNOS寡核苷酸的引物合成是根据Bredt等E3]发表的大鼠小脑NoscD- NA序列应用DNA合成仪合成的.引物A,B序列分别为:5GG:GAATCCATACCAGCCTGATCCATGGAACC35TACTCGAAACGCCTGAA TGG3用这一对引物可以扩增出自2463bp-3124bp(661bp)的NOScDNA序列,长度相当于821aa一1041aaNOS氨基酸序列.取1/lo体积(71)反转录反应液,分别加入200mmol/LdNTP,50pmol/LNOS寡核苷酸引物,0.5ttCi32P]dCTP和pH8.310mmol/LTr/s—HCI.50mmol/LKC1,1.5mmol/ LMsCl缓冲液,2.5unitTaqDNA聚合酶;总反应体积为100l.最后加入50山轻矿物油.PCR反应所需的变性,退火和延伸温度分别为:94℃,55℃和72℃,反应时间分别为:1,2和3min.共进行25—35次循环.首次循环.94℃需要3min.反应完成后取10山PCR反应物进行1.5琼脂糖凝胶电泳(含0.5~tg/ml的溴化乙锭)分析,以0xl74/HaeⅢ为分子量标准,电泳缓冲液为1xTBE(0.09mol/LTris一硼酸,0.002mol/LEDTApH8.0). 电泳完毕后在紫外灯下照片,并切取含有荧光条带的凝胶,应用液体闪烁计数仪进行放射性测量.2实验结果2.1NosmRNA刹量方法的建立本工作依据Martin(1993)的方法建立了定量NOSmRNA测量方法.其结果如图1. 2所示.由图1可以看出,脑组织总RNA反转录后经25—35次PCR扩增后,其NOScDNA片段约为660bp与所设计的NOScDNA相同.其扩增量与扩增循环次数密切相关.由图2可以看出,脑组织总RNA样本量不同,RT—PCR后NOScDNA产率亦不同.具有明显的4期氧化氮合成酶(Nos)基因表达的半定量捡涮及其运用349剂量一效应关系;应用0.25ug的总RNA,即可检测到NOSmRNA圉1PCR扩增产率与循环次数之间的剂量一效应关系.1Dose—effectrelationshipbetwec"theyieldofPCRproductandthenumberofPCRampJificationcycle(n一3,:K2:SE).M:xl74/Ha~Il000Am0u"【ofIOtillRNA2530圉2PCR产量和总P,NA量之间的剂量一效应关系Fig.2D.se—effectrelationshipbetwoentheyieldofPCRproductandtheamountoftotalRNAn一3,士SE.0,l?2,3,4tamountoftOtalRNA(0,0t25,0.5,1.0,1.5.2,respec-tire竹).MI.xl74I/HaeI.…取浓度脑组织总RNA(2g),分别经过5RT—PCR后.NOScDNA产率基本相同?具有良好的重复性.结果见图3所示.圈32恒量总RNART-PCR重复实验Fjg?3RT-PCRrepeatexperimentwithaconstant~Jnount.ftotalRNA(2ug)l,2,3-4,5:RT—PCRrepeattimesM:似174/Ha~I.2?2N0SmRNA在大鼠体内分布取正常Wistar大鼠小脑,心,肺,肾,肝组织,提取总RNA,应用RT-PC~,测定不同组织中NOSmRNA的分布,其总RNA用量均为2.0ug,其结果如图4所示.由图可l}{∞一.r.....mr....L.................._-畜●吾j墨lu.u0u芍fd2自言q孙lE3u1lx,&amp;5i0.I.0}dlu】lf【山生理46卷看出,在所测组织中均有NOSmRNA分布mRNA的含量约为肾脏和心脏的5倍.其中以小脑为最高,肝脏最低;其小脑内NOS取107的平滑肌细胞和内皮细胞,提取总RNA,经RT—PCT测定NOSmRNA,其细胞总RNA为2g.结果如图4所示.由图可以看出,NOSmRNA不仅存在于内皮细胞,亦分布于平滑肌细胞中,内皮细胞NOSmRNA的含量较高于平滑肌细胞图4NOS基因在正常大鼠不同组织和培养细胞中的表选Fig.4ExpressionofNOSgeneinvarioustissuesandculturedcellsofnormalratby RT—PCRM:HX174/HaeI}H:Heart;B}Brain;P;Lung;E,EC:Endothelialcall;K:KJdney~L!Uver;S,SMC:Smoothmusclec~11.2.3自发性高血压大鼠NosmRNA的变化分别取SHR和WKY大鼠的小脑,肾,肝组织,提取总RNA,依前法测定NOSmRNA .其所测样品总RNA用量均为2.结果如图5所示,由圈可以看出,在所检测的组织中,SHR大鼠NOSmRNA均低于WKY正常大鼠,其中在小脑降低了39.在肾脏降低了55,在肝脏降低了88.分别取10T个培养的WKY和SHR主动脉平精肌细胞,提取总RNA,各取2,同样用RT-PCR检测NOSmRNA,结果如图5所示.由图亦可以看出,SHR大鼠平精肌细胞中NOSmRNA含量明显降低,约较WKY大鼠降低了62.由以上结果可以看出,在SHR大鼠,无论是平滑肌细胞还是小脑,肾脏,其NOS基因的转录均明显降低.2.4自发性高血压大鼠内皮依赖性血管舒张反应为了进一步鉴定SHR大鼠血管EDRF/NO的反应性.取6只SHR大鼠主动脉环进行离giJc05u'j.三nlJ|.;4期一氧化氨合成酶(NOS)基因表达的半定量检测及其运用35J团SHR口WKY图5NOS基因在sHR和WKY不同组织和细胞中的表达Fig.5ExpressionofNOSgeneinvarioustissuesandculturedcellsinSHRandWKYn一3,土sE;P&lt;0.05,.'P&lt;0.01.M:174/HaeISMC:SmoothmusclecellSc:SHRSMC}SL:SHRLung;SK{SHRKidney;SB:SHRBrain;We!WKYSMClWL:WKYLung;WK:WKYKidney'WB:WKYBrain-圈6乙酰胆碱诱导的SHR和WKY主动脉内皮依赖性舒张反应≤Fig-6Endothelium—dependentrelaxationevkedbyAChinaortafromSHRandWKY.n=6.~4-SE,P&lt;0.05,"P&lt;0.01.j0——0WKygroupI●——●跚Rgroup1——L-ARG+SHR.囊~体灌流实验,观察乙酰胆碱(ACh)内皮依赖性血管舒张反应,结果如图6所示.由图可以看出.SHR大鼠主动脉ACh所引起的血管内皮依赖性舒张反应明显低于WKy 大鼠,其㈣uIdI.iirJ0j△2EⅢl;=E生理46卷最大的舒张反应亦只有WKY大鼠的I/2.为了进一步验证SHR大鼠ACh内皮依赖性血管舒张反应降低的机制,本实验还将10mol/LL-ARG预先加入SHR大鼠主动脉环灌流槽中,以补充EDRF/NO的前体,10min后,再进行ACh舒张反应实验,结果如图6所示.由图可以看出,补充Ⅱ)F,NO的前体,可使SHR主动脉ACh内皮依赖性舒张反应恢复.这些实验结果说明,自发性高血压大鼠NOS基因转录和表达下降,内皮依赖性血管舒张反应降低.3讨论3.1mRNA的测定mRNA的测定是分子生物学研究中一个常用的方法,主要有斑点杂交,NorthernBlotting等几种方法].但是由于mRNA含量少,又极易降解,因此对于其含有极小量特异mRNA的组织和样品,测定较为困难为此,近年来发展了RNA—protoc-fiveassay和含内标化的RT—PCR定量测定mRNA的方法.RNA—protectiveassay 虽然灵敏,准确,但操作复杂,价格昂贵,不易普及].定量RT—PCR虽然简便,快速并能定量,但因含有一个"内标准"而易产生"管效应",从而抑制特异性RT—PCR反应,干扰mRNA 的测定].本工作参照Martin(1993)~所建立的RT-PCR定量测定p-肾上腺素能受体mRNA 的方法,建立了NOSmRNA的测定方法.这种方法不另设内标准,排除了"管效应",而应用["P3dC'FP直接掺入PCR扩增反应,这不仅可使小量mRNA通过RT—PCR得以扩增和放大,而且可以通过掺入的放射性强度,直接进行rnRNA的定量测定.这种方法具有明显的荆量一效应关系和良好的重复性.我们曾经应用NorthernBlotting方法,测定不同组织中NOSmRNA含量,结果发现需要4O一6O嵋总RNA,才可检测到太鼠小脑和肾中NOSmR-NA,在肝脏,肺,平滑肌细胞中即使应用再太剂量总RNA亦难以测定;而应用本文这种方法,只需0.25—2g总RNA,即可测出组织中所含的NOSmRNA.应该指出.应用这种方法时,对提取的总RNA定量必须十分准确.ODOD的比值必须在18以上,总RNA定量应多次测量.3.2NOSmRNA在体内分布关于NOSmRNA在组织中的分布已有报道Brcdt等(1991)应用大鼠脑NOSeDNA探针和NorthernBlotting分析,发现NOSmRNA只存在于脑内rIWilliam等(1992)应用牛内皮细胞NOSeDNA为探针和NorthernBlotting分析,发现NOS只分布于内皮细胞中0}从而提出NOSmRNA不是普遍存在的.在许多组织中很难测到NOSmRNA的存在,但是不同组织的NOSe.DNA具有高度同源性[JI应用NOS 生物测定的方法证明,N0s亦广泛分布于不同组织中口".因此我们设想,在一些组织中难以测定出NOSmRNA,主要是因为组织中NOS特异性的mRNA在总RNA中所占比例过少,测定方法不够灵敏所致.本工作应用快速,灵敏的定量RT-PCR方法,不仅测出了脑内和内皮细胞中NOSmRNA,亦测出平滑肌细胞和心脏,肾脏,肺,肝脏组织中NOSmRNA.提示:NOSmRNA在体内不同组织中是广泛存在的.这与生物检测的结果相一致.过去认为NOSmR—NA主要存在于内皮细胞和巨噬细胞,本工作证明了在平滑肌细胞中亦有NOS的高表达}最近Nunokawa等(】993)从平滑肌细胞中克隆到NOSeDNA:Ⅲ,说明平滑肌细胞和内皮细胞4期一氧化氮合成酶(NOS)基因表达的半定量检测及其运用353一样,亦是NOS基因转录和表达的场所之一.近年来的研究表明,NOS可分为结构型和诱导型两类,而大鼠脑NOS即属于结构型[t3;我们应用依据大鼠脑NOScDNA序列设计合成的PCR引物,检测出在脑,心,肾,肺,肝,血管内皮细胞和平滑肌细胞中都有NOSmRNA的表达提示,结构型NOS可能是不同组织中所共有的.关于NOS结构型与诱导型之间的相互关系及其组织特异性,还需进一步研究3.3NOS在高血压发病中的作用既往的研究证明,内皮细胞损伤,Ea~RF/NO生成障碍是高血压发病的一个重要因素口.我们的实验亦证明,SHR大鼠血管内皮依赖性舒张反应明显下降.但是,这些研究都是基于药理方面的实验或是间接测定NOS的结果.关于高血压时,NOS基因转录和表达的变化,迄今尚未见报道.我们首次测定了SHR 大鼠不同组织和平滑肌细胞中NOSmRNA的变化,发现SHR大鼠NOS基因转录和表达明显低于WKY大鼠NOS是EDRF/No生成的关键因素,NOS基因表达下降,可致EDRF/NO 生成下降,这可能是高血压发生和发展的又一个重要因素.关于高血压时NOS基因转录下降的机制目前尚不了解.现有一些实验证明,NOS 基因表达受许多生长园子,细胞园子的调节Its].因此SHR大鼠NOSmRNA下降,亦可能是继发于生长因子,细胞因子的变化,亦可能是基因结构的改变所致.我们的初步实验证明,应用NOScDNA探针对SHR大鼠NOS基因进行限制性长度多态性分析,发现SHR 大鼠NOS基因可能有多态性变化.参考文献[1]FrinC~-R.GandOanso~,nL(1993).Pdsinginterest_mnitricoxidesyntha~.T/B8,18(2), 35—36.[2]William,C.S.,Jeffrey,K.H.andCynil~,M.B.(1992).Molecularcloningandexpressionof aeDNAencodingendot~lialcellnitricoxidesymha~.,J.嘲.0.,2I1,15274—15276.[3]Brech-nS.-Hwang,P.andSnyde,S.H.(1991).Clonedandexpre~ednitricoxidete$ntlm~s ttu~-rurallyre穹embl鹪~ytoeaxrocnoP-450reductase.神,351,7l4—718.[4]Herman.W.andGooSer,D.(1977).hmmn0fIucenoeintheidentificationofdtfferer*ttattnamoothmugcleoe1lsinculture.脚.D妇H.朋啪H∞-18,52—54.[5]Booyse,F.M.,Sedlak,B.J.andRatelson,E.(1975).Ctlltttre0farterialend劬e1iaIcellslcharacterl-zat~onandgrowthofbovineaofticendothelJalceils.M帕.姗商靖∞..¨.925—939. [6]aI∞mii1|H,Rand鼬ochl,N.(1987).Sin~e-stepmethod0P2qAIscdailonbyacidguantdiniumcain- extraction.肺c_Blsd~a..162.156—159.[7]Mar吐n,U..Michael,B.andJohn,S.E.(1993).Altered蜘酋onB-sdr能舡g【te憷呻rkimuleand~-adeene.zglc胁D佃reinthetatartshumanheart.帕'曲_,87,454—463.[8]Frederick-M.A.,Roge~,h-Robert,E.K.,Dav~l,nM.,S~idman,J.G.,dehn+A.S.and Kcvln,S.(1992).丽州丹咄帅缸脚咖.PlIb1hedbyGreenePublishingAssociatesandJohn Wiley&amp;Sons.,2rid.-呻.4-18~4-23.NewY ork.[93A1ice,M.W.,M~baeJ,V.D.andDavid,F.址(1989).QI衄乜吐onmRNAbymeIxdymeras~chainteac~on.Neff..删...86.9717—9721.[10]Natlum,C.(1992).Njn缸oxideasasecretorypr~luetofmammaliancells.FA~BBJ.,8,3051—3064.[¨]Hchalet"son,A.H.(1991).FMldothelinminCOllt/e1..^J..B5,ll6—125.[12]Nunol~wa?Y.,Islti~a-N.andTana[~,&amp;(1993).Cloningofinducibleninjcoxidesy nthas~r砒v-~lscu—lar~atooth/~rluseleo~ils..咖.脑."矾嘲.,191-89—94,生理46卷[13]David,A.G.,Andreas,K.N.andMauriclo,D.S.(1993).Cytokines,endotoxin,andgluco corticoidsregulatetheexpressionofinduciblenixieoxidesyntha~inhepatocytes.Proc.NatI_删?U3A.?90,施新猷(1989).医学实验动物学PP.39—40.陕西科学出版社,西安. ActaPhysio~ogicnSinlca1994,4g(4),347—354 SEMIQUANTITATIVEDETECTIONANDAPPLICATIONABOUT THEEXPRESS10NOFNIT砒COXIDESYNTHASEGENEZHAN6CHEN-HUI,LIQlANHONG,ZHAN6JIFNG,ZHOUHONG, ZHANGXn~G-BOANDTANGJIAN(,Mdeen~丑乩鲫,Card~sedar8硒由,&amp;咖Ma~a/,&amp;i谛10083)A±I'RAClUsingtheReverseTranscription(RT)一PolymeraseChainReaction(PER)method ofMartin(1993)forsemiquantitationdeterminatingofNOSgene,itwasfoundthat NOSmRNAisnotonlyexistedinbrain.butalsodistributedextensivelyinheart,kid- hey,lungandliver.Amongofthem,NOSmRNAlevelswerehighestinthebrain'fol- lowedindescendingorderbykidney,heart.Inadditiontoendothelialcell,NOSgene wasalsohighlyexpressedinsmoothmusclecells,suggestingthattheymaybeaDimpor- tantsiteofNOSinorganism.Furthermore.NOSmRNAlevelswerefoundtodecrease significatelyinbrain,kidney,liverandsmoothmusclecellinspontaneoushypertensive rat.Thesedatasuggestthatpathogenyofbypertentionmayberelatedtolowexpression 0fNOSgeneinthesetissues.KeywordstnitricoxideJv/nthm}8eneexpre*tton{polymerase~[111111reaction。

胃泌素释放肽前体在小细胞肺癌诊断及治疗中应用探讨

胃泌素释放肽前体在小细胞肺癌诊断及治疗中应用探讨

Journal of Mathematical Medicine Vol.34No.32021文章编号:1004-4337(2021)03-0328-03中图分类号:R7342文献标识码:A•临床科研分析•胃泌素释放肽前体在小细胞肺癌诊断及治疗中应用探讨卢佳荣(汕头市澄海区人民医院汕头515800)摘要:目的:探究胃泌素释放肽前体在小细胞肺癌诊断及治疗中应用。

方法:选取2018年12月〜2019年11月某院收治的小细胞肺癌和非小细胞肺癌患者各65例,分为小细胞肺癌患者组和非小细胞肺癌患者组。

结果:小细胞肺癌组的Pr()GRP(胃泌素释放肽前体)CEA(癌胚抗原)以及NSE(神经元特异度烯醇化酶)等血清指标高于非小细胞肺癌组(P<0.05)且小细胞肺癌组的CY-FRA21-1(细胞角蛋白片段浓度)等血清指标明显低于非小细胞肺癌组(P<0.05);小细胞肺癌组中,Pr()GRP的敏感度和特异度均高于CEA、NSE和CYFRA2-1等血清指标(P<0.05)结论:PoGRP可以为小细胞肺癌的临床诊断和治疗提供依据,值得推广。

关键词:胃泌素释放肽前体;小细胞肺癌;诊断;治疗doi:10.3969/j.issn.1004-4337.2021.03.005现阶段我国空气污染问题日益加重,肺癌患者的数量正在逐年增加,成为恶性肿瘤中的首位。

肺癌在临床上按照性质分为小细胞肺癌(SCLC)以及非小细胞肺癌(NSCLC),小细胞肺癌在肺癌患者数量中占有1/4,属于一类恶化度高,具有较快的侵袭性生长,极易形成具备广泛性坏死以及淋巴结转移等特征的恶性肿瘤。

经临床实践可知,大多数患者在诊断和治疗时已经为晚期,预后水平比较低,具有极低的5年生存率。

对两者进行明确区分是十分必要的,胃泌素释放肽前体(PoGRP)是现阶段应用的肿瘤标记物本文研究胃泌素释放肽前体在小细胞肺癌诊断及治疗中应用,现报告如下。

1资料与方法1.1一般资料选取2018年12月〜2019年11月我院收治的小细胞肺癌和非小细胞肺癌患者各65例,主要分为小细胞肺癌患者组和非小细胞肺癌患者组。

1 UV-vis-2013

1 UV-vis-2013
分子中电子经紫外可见光照射时电子从基态低能级跃迁到激发态高能级此时电子就吸收了相应波长的光波就形成了紫外可见光谱p4引起分子中电子能级跃迁的光波范围分为三个区域faruv10200nmvacuumuvregionnearuv
Spectroscopy

波 谱 学
中药化学教研室
梁 侨 丽
Office : B1-110
CH 3 CH 3
Solvent effect (cont)
在不同溶剂中谱带产生的位移称为溶剂效应。 是由于不同极性溶剂对基态和激发态样品分子的生色团稳 定化程度不同所致
溶剂极性增大使π—π*红移,n —π*跃迁蓝移
Solvent effect
(cont)
• This is caused by attractive polarisation forces between the solvent and the absorber, which lower the energy levels of both the excited and ground states in p -p* transitions . This effect is greater for the excited state, and so the energy difference between the excited and ground states is slightly reduced - resulting in a small red shift. • This effect also influences n -p* transitions but is overshadowed by the blue shift resulting from solvation of lone pairs(未成对电子).

蛋白质结构测定

蛋白质结构测定

In general
In particular, hydrophobic amino acids can be involved in binding/recognition of hydrophobic ligands such as lipids. Aromatic residues can also be involved in interactions with non-protein ligands that themselves contain aromatic groups via stacking interactions.
紫外与荧光光谱在蛋白质结构研究 中的应用
1 紫外差光谱法 (UV difference spectrum)
利用分光光度法测定大分子溶液状态下的构象,其原理
是利用环境对生物大分子生色基团的微扰而使吸收峰值的
位置、强度和带宽发生变化,如利用溶剂、pH、温度、 浓度等的微扰,使蛋白分子的酪氨酸的吸收峰发生变化, 从而推断这些生色基团在大分子中的位置和状态。
Fluorescence spectral shifts can be very large
Tryptophan in various solvents: hexane, trehalose glass(玻璃海藻糖), glycerol (丙三醇), water Ca-parvalbumin(小清蛋白): tryptophan is buried, 305 nm Ca-free parvalbumin: tryptophan is exposed to solvent
Tyr fluorescence

若蛋白质只含Tyr,不含Trp,
则蛋白质变性后,荧光强度 明显增强。

Polo样蛋白激酶1参与有丝分裂调控的研究进展

Polo样蛋白激酶1参与有丝分裂调控的研究进展

Polo样蛋白激酶1参与有丝分裂调控的研究进展赫玮(综述);高丰厚(审校)【摘要】As a crucial part of the cell cycle,the precise regulation of mitosis is precisely and strictly regulated,and along with the exploration in the regulation of mitosis,the understanding of life has deepened gradually as well.Polo-like kinase 1(PIK1) is involved in different processes of mitosis,and here is to sum-marize the functions,such as the activation of CDK1-Cyclin B complex,formation of spindle,segregation of chromosome and cytokinesis,and depict PLK-1′s significance for mitosis and put forward the possible direc-tions of further studies.%作为细胞周期的关键环节,有丝分裂过程受到严格而精细的调控,随着对有丝分裂调控的探讨与拓展,也逐渐加深了人们对生命本质的理解。

研究发现Polo 样蛋白激酶1( PLK1)参与细胞有丝分裂调控的各环节,该文拟归纳总结 Plk1在有丝分裂中诸如 CDK1-Cyclin B 复合物的激活、纺锤体形成、染色体分离和胞质分裂这些过程中的研究进展,并描绘PLK-1在有丝分裂调控中的作用与意义,为进一步深入探讨PLK-1与有丝分裂调控指出可能的发展方向。

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a rXiv:as tr o-ph/21362v116Oct22VLT spectroscopy of NGC 3115globular clusters Harald Kuntschner 1,Bodo L.Ziegler 2,3,R.M.Sharples 4,Guy Worthey 5,and Klaus J.Fricke 31European Southern Observatory,Karl-Schwarzschild-Str.2,85748Garching bei M¨u nchen,Germany 2Academy of Sciences,Theaterstr.7,37073G¨o ttingen,Germany 3Universit¨a tssternwarte,Geismarlandstrasse 11,37083G¨o ttingen,Germany 4Department of Physics,University of Durham,Durham DH13LE,UK 5Department of Physics,Washington State University,1245Webster Hall,Pullman,WA 99164-2814,USA Abstract.We present results derived from VLT–FORS2spectra of 17globular clus-ters associated with the nearby lenticular galaxy NGC paring line-strength indices to new stellar population models by Thomas et al.we determine ages,metal-licities and element abundance ratios.Our data are also compared with the Lick/IDS observations of Milky Way and M 31globular clusters.Our best age estimates show that the observed clusters which sample the bimodal colour distribution of NGC 3115glob-ular clusters are coeval within our observational errors (2–3Gyr).Our best calibrated age/metallicity diagnostic diagram (H βvs [MgFe])indicates an absolute age of 11–12Gyr consistent with the luminosity weighted age for the central part of NGC 3115.We confirm with our accurate line-strength measurements that the (V −I )colour is a good metallicity indicator within the probed metallicity range (−1.5<[Fe /H]<0.0).The abundance ratios for globular clusters in NGC 3115give an inhomogeneous picture.We find a range from solar to super-solar ratios for both blue and red clusters.This is similar to the data for M 31while the Milky Way seems to harbour clusters which are mainly consistent with [α/Fe]≃0.3.1The Sample and observations The candidate GCs were selected from the HST/WFPC2investigation of Kundu &Whitmore (1998)who detected 144globular cluster (herafter GC)candidates in the central region of NGC 3115.In order to keep integration times reasonablyshort only clusters with V <22(the peak of the GC luminosity function is at V =22.37±0.05)were selected while keeping a balance between red (V −I <1.06)and blue (V −I ≥1.06)clusters.The overall GC population of NGC 3115shows a clear bimodal colour distribution with mean metallicities at [Fe /H]≃−0.37and [Fe /H]≃−1.36as estimated from the V −I colours.In order to utilize the full field-of-view of FORS2we supplemented this list with GC candidates from a low-resolution spectroscopy survey reported in Kave-laars (1998)and also placed some slits on promising objects without prior infor-mation.In total 29spectra were obtained.Applying a cut in recession velocity and S/N (S/N ≥12per pixel)yielded a final sample of 17GCs.2Harald Kuntschner et al.The observations were carried out with FORS2on the VLT with the blue 600l/mm grism and1′′wide MXU slitlets giving a resolution of∼5˚A(FWHM). The total exposure time was12440s.2Abundance RatiosTheory of chemical enrichment predicts that stellar populations created in a short burst of star formation show elevated magnesium-to-iron abundance ratios while extended periods of star formation lead to roughly solar abundance ratios. Abundance ratios for integrated stellar populations can be best explored in a Mg b vs Fe index diagram( Fe =(Fe5270+Fe5335)/2).Figure1shows this diagnostic line-strength index diagram for GCs in NGC3115,the Milky Way and M31.Wefind that GCs in NGC3115show a range in abundance ratios similar to M31from[Mg/Fe]=0.0to0.5by comparing with models of Thomas et al. (2002).This applies to both the red and blue GCs in NGC3115.Fig.1.Probing the[Mg/Fe]ratios of globular clusters in a Mg b vs Fe diagram.(a) Our sample of globular clusters in NGC3115is shown as triangles(blue clusters)and circles(red clusters).The largefilled square represents the centre of NGC3115taken from Trager et al.(1998)and the smallfilled squares represent the data of Fisher,Franx &Illingworth(1996)which cover radii up to40′′along the major axis.(b)Milky Way globular clusters observed with the Lick/IDS instrumentation.(c)Globular clusters in M31observed with the Lick/IDS instrumentation.Thefilled triangle represents the centre of M31taken from Trager et al.(1998).Overplotted in all panels are models by Thomas et al.(2002)with abundance ratios of[Mg/Fe]=0.0.,0.3,0.5as indicated in the left panel.The models span a range in age(3–12Gyr)and metallicity([Fe/H]=−2.35to+0.3)VLT spectroscopy of NGC3115globular clusters3 3Age and metallicity estimatesIn Figure2we show age/metallicity diagnostic diagrams using the HβBalmer ab-sorption and the mean metallicity([Fe/H])indicator[MgFe].The error weighted mean ages of the blue and red GC populations are very similar to within2–3Gyr and favour absolute model ages of11–12Gyr.There is a clear correlation between the(V−I)colour and the metallicity as measured from metal absorption lines (see also Figure3).Fig.2.Age and metallicity diagnostic diagrams using as metallicity indicator[MgFe] ([MgFe]=4Harald Kuntschner et al.•We present a comparison of photometric and spectroscopic metallicity de-terminations andfind a good linear relation in the metallicity range probed by our sample of NGC3115clusters(−1.5<[Fe/H]<0.0).The photo-metric estimates are systematically lower(≃−0.26)in comparison with our spectroscopic measurements.•The existence of solar as well as elevated Mg-to-Fe ratios at a given metallic-ity for GCs in NGC3115indicates that a simple scenario of two distinct star-formation episodes is not sufficient to explain the formation of this galaxy.Probably a realistic model needs to incorporate more than two distinct star-formation events.For a detailed presentation of this work we refer the reader to our paper Kuntschner et al.,A&A acc.(astro-ph/0209129).References1.Fisher,D.,Franx,M.,Illingworth,G.1996,ApJ,459,1102.Kavelaars,J.J.1998,Ph.D.Thesis3.Kundu,A.,Whitemore,B.C.1998,AJ,116,28414.Thomas, D.,Maraston, C.,Bender,R.2002,submitted to MNRAS(astro-ph/0209250)5.Trager,S.C.,Worthey,G.,Faber,S.M.,Burstein,D.,Gonzalez,J.J.1998,ApJS,116,1Acknowledgements:Based on observations collected by Dr.J.Heidt(Heidel-berg)at the European Southern Observatory,Cerro Paranal,Chile(ESO No.66.B-0131).Part of this work was supported by the Volkswagen Foundation (I/76520).parison of photometric and spectroscopic metallicity estimates.(a)shows a direct comparison of observed quantities.In(b)we show a comparison between an empirical calibration of(V−I)colour against metallicity(dashed line)and predictions from stellar population models(solid line).(c)Photometric metallicities are estimated from the(V−I)colour,while spectroscopic metallicities are derived from the[MgFe] index assuming a constant age of12Gyr.The dotted line is the line of unity while the solid line is a linearfit to the data.。

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