Ice-free vitrification and nano warming technology for banking of cardiovascular structures.

用于心血管结构银行的无冰玻璃化和纳米加温技术。

• 批准号: 10026454
• 负责人: Kelvin G.M. Brockbank
• 金额: $ 83.78万
• 依托单位: TISSUE TESTING TECHNOLOGIES, LLC
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-11-01 至 2023-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10026454
• 关键词: Achievement; Animal Model; Animals; Aorta; Area; Arteries; Bathing; Biocompatible Materials; Biological; Biological Assay; Biomechanics; Blood Preservation; Blood Vessels; Blood Volume; Cardiovascular system; Carotid Arteries; Cations; Cell Survival; Cells; Chemistry; Clinical; Controlled Environment; Convection; Coronary Artery Bypass; Cryopreservation; Cryopreserved Tissue; Crystallization; Detection; Diagnostic; Dialysis procedure; Diffuse; Disaccharides; Dry Ice; Electromagnetics; Endothelium; Engineering; Evaluation; Excision; Exposure to; Extracellular Matrix; Family suidae; Formulation; Fracture; Freezing; Fresh Tissue; Future; Generations; Glass; Heating; Histopathology; Human; Hyperplasia; Ice; In Vitro; Inflammation; Laser Scanning Microscopy; Lead; Liquid substance; Logistics; Lung; Magnetic Resonance Imaging; Magnetic nanoparticles; Market Research; Measurement; Mechanics; Medial; Medicine; Metals; Methods; Microscopic; Modeling; Nitrogen; North America; Organ; Outcome; Patients; Peripheral; Permeability; Phase; Physiological; Property; Protocols documentation; Pulmonary artery structure; Raman Spectrum Analysis; Regenerative Medicine; Reproductive Medicine; Research; Residual state; Rewarming; Sample Size; Sampling; Scanning Electron Microscopy; Shipping; Small Business Innovation Research Grant; Solid; Specimen; Structure; Surface; System; Techniques; Technology; Temperature; Testing; Thick; Thinness; Time; Tissue Engineering; Tissue Preservation; Tissue Viability; Tissues; Transition Temperature; Translations; Transplantation; Vascular Graft; Work; base; blood vessel transplantation; cellular engineering; clinical application; clinically relevant; cryogenics; crystallinity; cytotoxicity; experience; experimental study; femoral artery; human tissue; in vivo; innovation; method development; microwave electromagnetic radiation; nano; nanoparticle; nanowarming; nonhuman primate; novel; novel strategies; packaging material; phase change; physical state; practical application; pre-clinical; preclinical evaluation; preservation; pressure; prevent; radio frequency; resazurin; response; second harmonic; technology development; transplant model; trauma care; vapor

ABSTRACT This proposal focuses on translation of ice-free cryopreservation by vitrification employing a novel approach of volumetric heating by nanowarming using Fe nanoparticles in an alternating electromagnetic ?eld. Vitrification, sub-zero storage below the glass transition temperature in a “glassy” rather than a crystalline frozen phase, is a form of cryopreservation that avoids ice formation. Vitri?cation can be achieved by quickly cooling the material to cryogenic storage temperatures, where ice cannot form. Vitri?cation can be maintained at the end of the cryogenic protocol by quickly rewarming the tissue to temperatures above the temperatures where ice nucleation may occur. The magnitude of the rewarming rates necessary to maintain vitri?cation is much higher than the magnitude of the cooling rates that are required to achieve it in the ?rst place. The most common approach to achieve the required cooling and rewarming rates is by convection based boundary warming in which the the specimen's surface is exposed to a temperature controlled environment, such as a fluid bath. Due to the underlying principles of heat transfer, there is a size limit in the case of surface boundary heating beyond which crystallization cannot be prevented at the center of the specimen. Furthermore, due to the underlying principles of solid mechanics, there is also a size limit beyond which thermal expansion in the specimen can lead to structural damage and fractures. Volumetric heating by nanowarming during the rewarming phase of the cryogenic protocol can alleviate these size limitations. Vitrification is already an important enabling approach for reproductive medicine with the potential to permit storage and transport of cells, tissues and organs for a great variety of biomedical uses. Unfortunately, practical application of vitrification has been limited to smaller systems such as cells and thin tissues due to diffusive and phase change limitations that preclude use for blood vessels, larger tissues and organs. To circumvent this problem we demonstrated that nanowarming effectively rewarms blood vessels in our preliminary research. Our experiments demonstrated that this innovative rewarming technique rewarmed vitrified femoral and carotid arteries in volumes ranging from 1 to 50mL with retention of cell viability and physiologic function. However, warming of thick arteries was suboptimal. We propose using large animal blood vessel, models for further optimization and evaluation of nanowarmed vessels using a combination of in vitro and in vivo studies. In Phase 1 in a single specific aim we will optimize ice-free vitrification of thick walled arteries, aorta and pulmonary, with a go/no go objective of achieving > 90% viability for progression to Phase 2. In Phase 2 specific aims, we propose using porcine vascular models in a combination of ex vivo and in vivo studies. The magnetic nanoparticles will be distributed around and within the internal spaces of vessels. The large vessel lumen space makes them a good choice for optimization of vitrification and nanowarming. In Aim 1 we will evaluate cryopreserved arteries after real time shipping, comparing methods and validating the transport conditions that are finally approved based upon absence of tissue cracking. In Aim 2 we will characterize the post-ice-free cryopreservation state of arteries preserved for at least 2 years. In addition, during this aim we will characterize the chemistry and biomaterial properties of ice-free cryopreserved blood vessels. Effective vitrification will be evaluated using cryomacroscopy to detect ice formation and cryoprotectant residuals by Raman spectroscopy. In Aim 3 we will perform short-term transplant studies (28 days) in two porcine vascular models (femoral and pulmonary artery into the carotid and pulmonary, respectively) in order to validate our technology for a future Phase IIb SBIR proposal using clinically relevant preclinical non-human primate models and human tissues.

抽象的 该提案的重点是通过使用一种新颖的方法来翻译无冰冷冻保存 通过纳米武器使用fe纳米颗粒在替代电磁？ELD中使用纳米颗粒来加热。玻璃， 玻璃过渡温度以下以“玻璃”而不是结晶冷冻相的玻璃过渡温度的零储存量为 一种避免冰形成的冷冻保存形式。通过快速冷却可以实现vi​​tri？阳离子 冰储存温度的材料，冰无法形成。可以在末端维持Vitri？阳离子 通过快速将组织转换为高于温度的温度，使得低温方案 可能会发生成核。维持VITRI所需的重新传输速率的大小要高得多 比在第一个地方实现的冷却速率的大小。最常见的 实现所需的冷却和重新加热速率的方法是基于施工的边界变暖 样品表面暴露于温度控制的环境（例如流体浴）。 由于传热的基本原理，在表面边界加热的情况下，尺寸极限 除此之外，不能在样品的中心预防结晶。此外，由于 固体力学的基本原理，还有一个尺寸限制，超出了其中的热膨胀 标本会导致结构损伤和断裂。纳米武器在 低温方案的重新加热阶段可以减轻这些尺寸限制。玻璃化已经是 重要的促进方法的生殖医学方法，有可能允许存储和运输 细胞，组织和器官用于多种生物医学用途。不幸的是，实际应用 由于扩散和相，玻璃化限于较小的系统，例如细胞和薄组织 改变限制，排除用于血管，较大组织和器官的使用。解决这个问题 我们证明，在我们的初步研究中，纳米武术有效地重新加热了血管。我们的 实验表明，这种创新的重新加热技术重新加热了股股和颈动脉 体积的动脉范围为1至50毫升，并保留细胞活力和生理功能。 但是，厚动脉的变暖是次优的。我们建议使用大型动物血管，模型 为了进一步优化和评估纳米臂血管，使用体外和体内的组合 研究。在单个特定目标中，我们将优化厚壁动脉的无冰玻璃化， 主动脉和肺部，具有GO/NO的目标，可以实现> 90％的生存能力，以进展到第2阶段。 第2阶段的特定目的，我们建议在离体和体内结合使用猪血管模型 研究。磁性纳米颗粒将分布在容器的内部空间周围和内部。 大容器腔空间使它们成为优化玻璃化和 纳米武装。在AIM 1中，我们将在实时运输后评估冷冻保存的动脉，比较方法 并验证根据没有组织裂纹的最终批准的运输条件。目标 2我们将表征至少2年的无冰后冷冻保存状态。在 此外，在此目标中，我们将表征无冰的化学和生物材料 冷冻保存的血管。有效的玻璃化将使用冷act镜检查评估以检测冰 拉曼光谱法的形成和冷冻保护剂残差。在AIM 3中，我们将短期执行 两种猪血管模型（股动脉和肺动脉纳入颈动脉）的移植研究（28天） 和肺部分别）为了验证我们的技术，以实现未来的IIB SBIR建议 临床相关的临床前非人类灵长类动物模型和人体组织。