Neutrophil-mediated Drug Delivery

项目总结： 血管是传递治疗性纳米颗粒的巨大障碍，然而现有的 解决这一问题的方法是不够的。中性粒细胞是最丰富的白细胞。 在血液中循环，并能够转运到组织中，以快速反应组织损伤和 感染。因此，中性粒细胞可以作为一种很好的载体来调节纳米颗粒在血管中的传输。 障碍。使用活体显微镜和急性肺炎症小鼠模型，我们已经证明 由白蛋白蛋白制成的纳米颗粒可以与激活的中性粒细胞特异性结合，从而导致 这些纳米颗粒通过中性粒细胞介导的转运穿过内皮屏障。因此，我们假设 中性粒细胞可以将装载在白蛋白纳米粒中的药物穿过血管屏障，并 将药物存放在病变组织中。此外，使用这种新的方法，我们可以防止急性 肺损伤是一种毁灭性的疾病，目前还没有有效的药物治疗。我们有 建议的三个目标： 目的1：确定靶向活化白蛋白所需的白蛋白纳米平台的性质 体内的中性粒细胞。我们将开发产生明亮的荧光标记白蛋白纳米颗粒的方法 用于追踪细胞和活老鼠体内的纳米颗粒。我们将讨论颗粒大小、密度、表面电荷的作用 以及白蛋白性质对纳米颗粒摄取的影响。我们将测定白蛋白的载药效率 用高效液相色谱/质谱仪测定纳米粒和药物在体外和体内的释放情况。 目的2：剖析中性粒细胞介导的纳米粒子跨血转运的机制。 血管屏障。利用活体显微镜下的小鼠生殖发育器组织，我们将实时可视化 中性粒细胞和携带白蛋白纳米粒的中性粒细胞对白蛋白纳米粒的摄取 血管内皮细胞并转移到炎症组织。在气管-内毒素诱导的急性肺炎症模型中， 我们将调查中性粒细胞摄取纳米颗粒是否会影响中性粒细胞的生物学功能(运输 使用共聚焦显微镜、生化方法和基因敲除小鼠模型。 目的3：确定我们的纳米颗粒平台治疗急性肺损伤的疗效。通过 检测小鼠肺内细胞因子、中性粒细胞浸润和血浆蛋白蓄积，以及组织化学， 我们将评估负载了NF-κB抑制剂的白蛋白纳米粒是否可以显著减轻肺损伤 炎症/损伤与免费药物相比。 在这些研究结束时，我们将开发新的策略，将治疗药物输送到靶点 关于中性粒细胞向病变部位的迁移，并创造了一种通过内皮运送纳米颗粒的方法 障碍。我们的研究将改变目前治疗急性和慢性炎症性疾病的策略 包括癌症在内的疾病。具体地说，我们的方法将对新的 治疗急性肺损伤和一种严重形式的急性呼吸窘迫综合征的药物治疗 (ARDS)，因为目前还没有有效的疗法来治疗这些死亡率高达40%的毁灭性疾病。 为了加强我们的研究计划，我们要求通过资助机会更新我们的活体显微镜。 《行政副刊》一书。

Project Summary: Blood vessels present a great barrier for delivering therapeutic nanoparticles, however the existing approaches are not sufficient to resolve this problem. Neutrophils are the most abundant white blood cells circulating in bloodstream, and are capable of transmigrating into tissues to rapidly respond tissue injury and infection. Therefore, neutrophils could be an excellent carrier to mediate nanoparticle transport across vessel barrier. Using intravital microscopy and an acute lung inflammation mouse model we have demonstrated that nanoparticles made from albumin protein can specifically bind activated neutrophils leading to the transport of these nanoparticles across endothelial barrier via neutrophil-mediated transmigration. Thus, we hypothesize that neutrophils can transport drugs loaded in albumin nanoparticles across a blood-vessel barrier and deposit the drugs in diseased tissues. Furthermore, using this novel approach we could prevent acute lung injury, a devastating disease currently without effective pharmacological therapies. We have proposed three aims: Aim 1: Define the properties of albumin nanoparticle platforms required for targeting of activated neutrophils in vivo. We will develop the methods to generate bright fluorescently-labeled albumin nanoparticles for tracking nanoparticles in cells and live mice. We will address the roles of particle size, density, surface charge and albumin properties in affecting nanoparticle uptake. We will determine drug loading efficiency in albumin nanoparticles and drug release profiles in vitro and in vivo using HPLC/mass spectroscopy. Aim 2: Dissect the mechanism of neutrophil-mediated nanoparticle transport across the blood- vasculature barrier. Using intravital microscopy of mouse cremaster tissues, we will real-time visualize the uptake of albumin nanoparticles by neutrophils and neutrophils carrying the nanoparticles transmigrate across endothelial vessels and move to inflammatory tissues. In a tracheal-LPS induced acute lung inflammation model, we will investigate whether nanoparticle uptake by neutrophils affects neutrophil biological functions (transport and activity) using confocal microscopy, biochemical approaches and knockout mouse models. Aim 3: Determine the efficacy of our nanoparticle platforms in treatment of acute lung injury. By measuring cytokines, neutrophil infiltration and plasma protein accumulation in mouse lungs, and histochemistry, we will evaluate whether albumin nanoparticles loaded with NF-κB inhibitors can dramatically mitigate lung inflammation/injury compared to the free drugs. At the end of these studies, we will develop the novel strategies of delivering therapeutic into targets based on neutrophil transmigration to diseased locations, and create a way to deliver nanoparticles across endothelial barrier. Our studies will change current strategies to treat inflammatory disorders underlying acute and chronic diseases including cancer. Specifically, our approach will significantly impact on the development of new pharmacological therapies to treat acute lung injury, and a severe form, acute respiratory distress syndrome (ARDS) because there are no effective therapies to treat these devastating diseases with 40% mortality. To enhance our research program, we request to update our intravital microscope via the funding opportunity of Administrative Supplement.