Modeling immunity with a hybrid lymph node tissue-chip

项目摘要/摘要 免疫系统中的局部相互作用决定了免疫反应是保护性的还是 破坏性的、抗感染的或启动自身免疫的。获得性免疫始于淋巴(LN)， 高度组织化和充满活力的组织。目前，很难分析炎症介质或 合理设计动脉粥样硬化、类风湿性关节炎等慢性炎症性疾病的治疗方案 多发性硬化症，这些疾病加起来影响了5%-7%的西方人口。我们假设分析本地数据 在完整组织中的体外反应将提供从当前方法(体外/体内)不易获得的信息 Vivo)。这样的实验需要新的工具来分析免疫系统中的动力学，我们通过 结合了生物分析化学、微流体学和免疫学的专业知识。 在这个项目中，我们将开发一种新的体外免疫模型，使用微流控的混合 培养和LN切片。在目标1中，我们将建立长期的文化与分析方法相结合的生活 小鼠和人的LN切片。切片培养提供了保存细胞外的优势 微环境和任何与矩阵结合的信号。我们将优化小鼠LN的长期培养(7-21天) 切片和人类扁桃体切片，以保持高活性，低细胞激活标记静息，并能力 对炎症性和抗原特异性刺激做出反应。在目标2中，我们将开发一种新的微流控系统 按需局部刺激LN切片。我们将提高我们以前开发的空间分辨率 设备，使用短期和长期目标，以直径2-10个细胞群(横向分辨率20-100μm)为目标 刺激。我们还将通过使用移动端口按需选择投递区，使 只需最少的操作即可接触到整个组织。在目标3中，我们将验证混合微流控组织切片 用于分析炎症反应和抗炎治疗的系统。我们将比较 切片与细胞培养和体内对促炎细胞因子肿瘤坏死因子-α的炎症反应 系统。最后，我们将测试该模型在多大程度上提供了指导免疫治疗的新信息， 通过使用小鼠和人类组织的混合组织芯片系统来比较竞争的效果 肿瘤坏死因子-α抑制剂(抗肿瘤坏死因子-α单克隆体或可溶性肿瘤坏死因子-α受体)。 将局部微流控刺激与组织切片技术相结合，产生了第一个实验 用于分析活体LN组织中空间组织的信号和细胞-细胞相互作用的平台。这是一项创新 平台将促进基础和转化性生物医学研究：当地提供的细胞因子将作为 一个急需的急性或慢性炎症模型，以及当地提供的免疫疗法将指导 靶向载药纳米粒的设计。这项技术广泛适用于多种炎症性疾病 疾病，包括类风湿性关节炎、慢性疾病、多发性硬化症、阿尔茨海默病和癌症。

PROJECT SUMMARY/ABSTRACT Local interactions in the immune system determine whether an immune response is protective or destructive, fighting infection or initiating autoimmunity. Adaptive immunity begins in the lymph node (LN), a highly organized and dynamic tissue. Currently, it is difficult to parse the role of inflammatory mediators or rationally design therapies for chronic inflammatory disease such as artherosclerosis, rheumatoid arthritis and multiple sclerosis, which together affect 5 – 7% of the Western population. We hypothesize that analyzing local responses ex vivo in intact tissue will provide information not easily obtained from current methods (in vitro/in vivo). Such experiments require new tools to analyze dynamics in the immune system, which we develop by combining expertise in bioanalytical chemistry, microfluidics, and immunology. In this project, we will develop a novel ex vivo model of immunity, using a hybrid of microfluidic culture and LN slices. In Aim 1, we will establish long-term culture coupled with analysis methods for live murine and human LN slices. Slice culture offers the advantage of preservation of the extracellular microenvironment and any matrix-bound signals. We will optimize long-term culture (7-21 days) for murine LN slices and human tonsil slices to maintain high viability, low cellular activation markers at rest, and ability to respond to inflammatory and antigen-specific stimuli. In Aim 2, we will develop a novel microfluidic system for on-demand local stimulation of LN slices. We will improve the spatial resolution of our previously developed device, to target clusters 2 – 10 cells in diameter (20 – 100 μm lateral resolution) using short- and long-term stimulation. We will also enable on-demand selection of delivery zone by using a mobile port, making the whole tissue accessible with minimal handling. In Aim 3, we will validate the hybrid microfluidic-tissue slice system for analysis of inflammatory responses and anti-inflammatory therapies. We will compare the inflammatory response to a pro-inflammatory cytokine, TNF-α, in slices versus cell cultures and in vivo systems. Finally, we will test the extent to which the model provides new information to guide immunotherapy, by using the hybrid tissue-chip system with mouse and human tissue to compare the effects of competing TNF-α inhibitors (anti-TNF-α monoclonals or soluble TNF-α receptor). Combining local microfluidic stimulation with tissue slice technology produces the first experimental platform for analysis of spatially organized signaling and cell-cell interactions in live LN tissue. This innovative platform will advance both basic and translational biomedical research: locally delivered cytokines will serve as a much-needed model of acute or chronic inflammation, and locally delivered immunotherapies will guide the design of targeted drug-loaded nanoparticles. This technology is broadly applicable for a host of inflammatory diseases, including rheumatoid arthritis, Chron’s disease, multiple sclerosis, Alzheimer’s disease, and cancer.