Name: visualization of vascular ca2+ signaling triggered by paracrine derived ros
Uploaded: 2011-12-21T13:00:00
Description: Watch this Scientific Journal Video about Visualization of Vascular Ca2+ Signaling Triggered by Paracrine Derived ROS at JoVE.com

这项工作是支持批卫生研究院（R01 HL086699，HL086699 01A2S1，1S10RR027327 - 01）的MM。我们的文章部分是由卡尔蔡司显微成像公司的支持。

活性氧可测氧化敏感染料（1＆2）使用或使用质粒传感器（3＆4），共聚焦显微镜的活细胞。 1。 J774细胞胞质ROS的可视化<ol><li>生长在玻璃底部35 mm培养皿（哈佛大学仪器），48小时J774.1小鼠单核细胞衍生的巨噬细胞（10 6细胞/ ml） </li><li>挑战细胞TLR激动剂（2微克/毫升，磷壁酸锂TLR2激动剂;10μg/ml，保利（I：C）- TLR3激动剂; 1微克/ ml的LPS - TLR4的激动剂）为6小时37 ° C <...

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这里介绍的方法，允许快速和定量测量活性氧，在活细胞使用的氧化敏感的染料或质粒传感器。的Toll样受体激动剂（Toll样受体）的化合物，通过TLRs的细胞表面上的刺激细胞和触发下游信号通路15。在我们的协议中，我们使用三种不同的TLR激动剂，即，锂，磷壁酸TLR2激动剂;聚（I：C）。TLR3激动剂; LPS - TLR4的激动剂据报道，作为一个模型系统诱导ROS证明活性氧测量。使用氧化剂敏感染料的?...

<table border="1"><tbody><tr><td> 试剂 </td><td> 公司 </td><td> 目录编号 </td></tr><tr><td> Attofluor细胞室</td><td> Invitrogen公司</td><td> A7816 </td></tr><tr><td>抗霉素A </td><td>西格玛爱秩序</td><td> A8674 </td></tr><tr><td> DMEM低血糖液</td><td> Invitrogen公司</td><td> 10567-014 </td></tr><tr><td>内皮生长因子的补充（ECGS） </td><td>上州</td><td> 02-102 </td></tr><tr><td>胎牛血清</td><td> Invitrogen公司</td><td> 12662011 </td></tr><tr><td> G418 </td><td> Invitrogen公司</td><td> 10131-027 </td></tr><tr><td>明胶</td><td>西格玛爱秩序</td><td> G1393 </td></tr><tr><td> H 2 DCFDA </td><td> Invitrogen公司</td><td> D - 399 </td></tr><tr><td>脂多糖</td><td>西格玛爱秩序</td><td> L4516 </td></tr><tr><td>陆路交通管理局</td><td>西格玛爱秩序</td><td> L2515 </td></tr><tr><td> MitoSOX红</td><td> Invitrogen公司</td><td> M36008 </td></tr><tr><td> OPTI - MEM我降低血清培养基</td><td> Invitrogen公司</td><td> 51985091 </td></tr><tr><td> PEN /链球菌（10X） </td><td> Invitrogen公司</td><td> 15140163 </td></tr><tr><td> pHyPer - CYTO </td><td> Evrogen </td><td> FP941 </td></tr><tr><td> pHyPer - dMito </td><td> Evrogen </td><td> FP942 </td></tr><tr><td>保利（I：C） </td><td>西格玛爱秩序</td><td> P0913 </td></tr><tr><td> Prism软件5.0 </td><td> GraphPad软件公司</td><td></td></tr><tr><td> SigmaPlot 11.0 </td><td> SYSTAT软件公司</td><td></td></tr><tr><td>胰蛋白酶EDTA（10X） </td><td> Invitrogen公司</td><td> 15400054 </td></tr><tr><td> T - 25培养瓶</td><td>康宁</td><td> 430639 </td></tr><tr><td> T - 75培养瓶</td><td>屋宇署猎鹰</td><td> 353136 </td></tr><tr><td> 96孔TC微型孔板</td><td>屋宇署猎鹰</td><td> 353072 </td></tr><tr><td>禅2009软件</td><td>卡尔蔡司510元共聚焦显微镜</td><td></td></tr></tbody></table>

visualization of vascular ca2+ signaling triggered by paracrine derived ros

氧化应激有牵连的一些包括缺血/再灌注损伤和败血症的病理条件。氧化应激的概念是指异常形成的活性氧（活性氧），其中包括 Ø 2• - ，H 2 O 2，和羟基自由基。活性氧的影响多种细胞过程，包括信号转导，细胞增殖和细胞死亡 1-6 。活性氧有可能直接损伤血管和器官的细胞，并能启动中学的化学反应和遗传变异，最终导致初始ROS介导的组织损伤的放大。放大级联，加剧了不可逆的组织损伤的一个关键组成部分是招募和循环炎性细胞的激活。在炎症，炎性细胞产生细胞因子如肿瘤坏死因子-α（TNFα）和IL - 1，激活内皮细胞（EC）和上皮细胞，并进一步增强炎症反应 7 。血管内皮功能障碍是一种急性炎症的建立功能。巨噬细胞促进血管内皮功能障碍，在炎症的机制仍不清楚。 • -和各种促炎性细胞因子，从而触发在相邻细胞的病理信号 8激活巨噬细胞外释放的O 2的结果。 NADPH氧化酶是ROS的主要的主要来源，在大多数的细胞类型。最近，它是我们和其他9,10所示，由NADPH氧化酶产生的活性氧诱导线粒体ROS在许多病理生理条件的生产。因此，测量线粒体ROS产生同样重要的是，除了测量胞质ROS的。巨噬细胞产生ROS的黄素酶NADPH氧化酶在炎症反应中起着主要作用。一旦被激活，吞噬细胞NADPH氧化酶产生丰富的O 2• - 11,12在宿主防御机制的重要。虽然旁分泌衍生Ø 2• -扮演重要的作用，在血管疾病的发病机制，包括CA 2旁分泌ROS诱导的细胞内信号可视化 +动员仍然是假说。我们已经开发出了激活巨噬细胞作为氧源•使用模型-信号转导的一个相邻的内皮细胞。使用这个模型，我们表明，巨噬细胞源性Ø 2• -在邻近的内皮细胞钙信号。

Watch this Scientific Journal Video about Visualization of Vascular Ca2+ Signaling Triggered by Paracrine Derived ROS at JoVE.com

Visualization of Vascular Ca2+ Signaling Triggered by Paracrine Derived ROS

一种有效的方法获得可视化的旁分泌衍生活性氧诱导内皮细胞Ca2 +信号的见解的描述。这种方法测量旁分泌衍生活性氧引发共培养模型在血管内皮细胞内Ca2 +动员的优势。

血管钙的可视化 2 +信号旁分泌源性活性氧引发

Oxidative stress has been implicated in a number of pathologic conditions including ischemia/reperfusion damage and sepsis. The concept of oxidative ...

Research

JoVE Journal

Biology

Visualization of Vascular Ca2+ Signaling Triggered by Paracrine Derived ROS

Techniques for Imaging Ca2+ Signaling in Human Sperm

Techniques for Imaging Ca2+ Signaling in Human Sperm

技术成像钙 2 +信号在人类精子

Fluorescence microscopy of cells loaded with fluorescent, Ca2+-sensitive dyes is used for measurement of spatial and temporal aspects of Ca2+ signaling in ...

科研

生物学

Visualization of ATP Synthase Dimers in Mitochondria by Electron Cryo-tomography

ATP合酶二聚体线粒体通过冷冻电子断层扫描可视化

Electron cryo-tomography is a powerful tool in structural biology, capable of visualizing the three-dimensional structure of biological samples, such as ...

Automated Analysis of Dynamic Ca2+ Signals in Image Sequences

Automated Analysis of Dynamic Ca2+ Signals in Image Sequences

动态钙的自动分析<sup&gt; 2 +</sup&gt;在图像序列信号

Intracellular Ca2+ signals are commonly studied with fluorescent Ca2+ indicator dyes and microscopy techniques. However, quantitative analysis of Ca2+ ...

Fecal Glucocorticoid Analysis: Non-invasive Adrenal Monitoring in Equids

糖皮质激素粪分析：在马属动物无创监测肾上腺

Adrenal activity can be assessed in the equine species by analysis of feces for corticosterone metabolites. During a potentially aversive situation, ...

DiI Perfusion as a Method for Vascular Visualization in Ambystoma mexicanum

DiI Perfusion as a Method for Vascular Visualization in Ambystoma mexicanum

DiI灌注作为血管可视化的方法<em&gt;墨西哥恶性肿瘤</em

Perfusion techniques have been used for centuries to visualize the circulation of tissues. Axolotl (Ambystoma mexicanum) is a species of salamander that ...

Visualization of DNA Compaction in Cyanobacteria by High-voltage Cryo-electron Tomography

高压低温电子层析成像对蓝藻 DNA 压实的可视化研究

This protocol describes how to visualize the transient DNA compaction in cyanobacteria. DNA compaction is a dramatic cytoplasmic event recently found to ...

Retinal Cryo-sections, Whole-Mounts, and Hypotonic Isolated Vasculature Preparations for Immunohistochemical Visualization of Microvascular Pericytes

视网膜低温切片、全支架和低渗隔离血管毛细血管微血管组织免疫组化的制备

Retinal pericytes play an important role in many diseases of the eye. Immunohistochemical staining techniques of retinal vessels and microvascular ...

A Proximal Culture Method to Study Paracrine Signaling Between Cells

一种研究细胞间分泌信号的近距离培养方法

Intercellular interactions play an important role in many biological processes, including tumor progression, immune responses, angiogenesis, and ...

Myeloid Innate Signaling Pathway Regulation by MALT1 Paracaspase Activity

palt1 副酶活性对髓系先天信号通路的调控

Besides its function in lymphoid cells, which has been addressed by numerous studies, the paracaspase MALT1 also plays an important role in innate cells ...

Applications of Spatio-temporal Mapping and Particle Analysis Techniques to Quantify Intracellular Ca2+ Signaling In Situ

Applications of Spatio-temporal Mapping and Particle Analysis Techniques to Quantify Intracellular Ca2+ Signaling In Situ

时空映射和粒子分析技术在细胞内 ca2 +原位信令定量中的应用

Ca2+ imaging of isolated cells or specific types of cells within intact tissues often reveals complex patterns of Ca2+ signaling. This activity requires ...