Harnessing the Power of Apoptosis to Create Regenerative Acellular Biologic Scaffolds

1605223-施密特通过使用天然衍生材料，特别是脱细胞基质，例如脱细胞人真皮和脱细胞人神经移植物，在再生医学方面取得了许多成功。这些脱细胞基质进一步发展的一个关键限制是在标准的脱细胞处理过程中失去基质微结构和生物活性，这通常始于一个导致细胞坏死(通过损伤死亡)的过程，释放整个基质中的细胞内成分，然后使用苛刻的化学物质清洗以去除细胞残留物，这也会导致不希望看到的基质破坏和有利的生物活性因子的去除。该项目寻求通过诱导细胞凋亡(程序性细胞死亡)而不是坏死来改变去细胞技术的范式。在细胞凋亡的过程中，细胞从基质中分离出来，形成小的凋亡小体，其中含有可以很容易去除的细胞成分。此外，在细胞凋亡期间，细胞会分泌细胞因子，这些细胞因子有助于剩余的细胞外基质中的组织再生。细胞凋亡技术将用于开发具有保存下来的微结构和生物活性因子的神经和肺组织特异性基质，这些基质可以与商业上可用的产品进行比较。神经组织基质将在大鼠坐骨神经缺陷模型中进行测试，并与商业产品进行比较。所获得的成果有望为医学研究界创造一个新的平台，用于生成具有增强的再生潜力的无细胞组织特异性基质，这可能会极大地改变再生医学的未来。通过培训一名研究生和几名本科生，开发有趣的互动课程和实验室，向中学生传授生物材料和生物医学工程知识，并提高年轻女性对STEM的兴趣，将达到教育和推广的效果。该项目的目标是利用细胞凋亡诱导细胞死亡的力量，创造具有保存的微结构和生物活性因子的组织特异性基质，能够促进组织和器官的再生。天然衍生材料，特别是脱细胞基质对于再生医学的成功至关重要。这些脱细胞基质进一步发展的一个关键限制是在标准脱细胞过程中失去精细的基质结构和生物活性。这项工作的假设是，通过凋亡诱导细胞死亡将使细胞和抗原更容易和更温和地去除，从而更好地保存基质微结构和生物活性因子。此外，凋亡细胞还会分泌促进植入后再生的因子。由组织特异性细胞外基质衍生的生物支架在组织工程和再生医学中是非常理想的，因为它们由植入部位天然的蛋白质组成，并为再生提供支架。然而，重要的是在植入前去除所有细胞成分，以防止免疫原性。最常用的商业去细胞方法是从导致细胞坏死的过程开始，释放整个基质中的细胞内成分，然后使用苛刻的化学物质清洗以去除细胞残留物，这也会导致不希望看到的基质破坏和有利的生物活性因子的去除。该项目寻求通过诱导细胞凋亡而不是坏死来改变脱细胞技术的范式。在细胞凋亡过程中，细胞从基质中分离出来，形成含有细胞成分的小的凋亡体，这些细胞成分很容易被去除。此外，在细胞凋亡期间，细胞分泌的细胞因子可以隔离在细胞外基质中，并被认为可以诱导增殖、干细胞募集和免疫调节，从而帮助组织再生。在特定的目标中，1利用具有代表性的组织和器官(神经和肺)中的细胞凋亡来创建具有保存的微结构和生物活性因子的新的组织特异性基质。经细胞凋亡优化的脱细胞移植物的生物活性和再生潜力将与商业上使用的脱细胞技术的移植物进行比较。特定目的2使用体内模型来分析经凋亡优化的无细胞移植物与由商业使用的脱细胞技术创建的移植物的免疫反应和再生潜力。所获得的成果有望为医学研究界创造一个新的平台，用于生成具有增强的再生潜力的无细胞组织特异性基质，这可能会极大地改变再生医学的未来。将通过培训一名研究生和几名本科生，开发有趣的互动课程和实验室，向中学生传授生物材料和生物医学工程知识，在S女孩？Place，Inc.为6-8年级的女孩开发课程，提高年轻女性S对STEM的兴趣，以及与盖恩斯维尔凯德博物馆建立新的合作伙伴关系，将课程扩展到全面的实验室，定期运行，鼓励儿童的创造力和发明，从而实现教育和外联的影响。

1605223 - SchmidtMany successes in regenerative medicine have been achieved through the use of naturally derived materials and particularly decellularized matrices, e.g., decellurized human dermis and decellurized human nerve graft. One key limitation to further advancement of these decellularized matrices is a loss of matrix microarchitecture and bioactivity during standard decellularization processing, which often starts with a process to induce cell necrosis (death through damage), releasing intracellular components throughout the matrix, followed by washes using harsh chemicals to remove cell remnants, which also results in undesired matrix destruction and removal of favorable bioactive factors. This project seeks to shift the paradigm in decellularization techniques by inducing apoptosis (programmed cell death) rather than necrosis. During apoptosis cells detach from the matrix and form small apoptotic bodies containing cellular components that can be readily removed. Furthermore, during apoptosis cells secrete cytokines that are known to aid in tissue regeneration in the remaining extracellular matrix. The apoptosis technique will be used to develop nerve and lung tissue specific matrices with preserved microarchitecture and bioactive factors that can be compared to commercially available products. The nerve tissue matrix will be tested in a rat sciatic nerve defect model, also in comparison with commercially available products. Results obtained are expected to contribute to the medical research community by creating a new platform for generation of acellular tissue specific matrices with enhanced regenerative potential, which could dramatically shift the future of regenerative medicine. Educational and outreach impact will be achieved through training of a graduate student and several undergraduate students and developing fun interactive lessons and labs to teach middle school students about biomaterials and biomedical engineering and to increase young women's interest in STEM.The goal of this project is to harness the power of apoptosis induced cell death to create tissue specific matrices with preserved microarchitecture and bioactive factors capable of enhancing regeneration in tissues and organs. Naturally derived materials and particularly decellularized matrices have been essential to success in regenerative medicine. One key limitation to further advancement of these decellularized matrices is a loss of fine matrix structure and bioactivity during standard decellularization processing. The hypothesis of this work is that induction of cell death by apoptosis will enable easier and gentler cell and antigen removal, thus better preserving matrix microarchitecture and bioactive factors. Furthermore, apoptotic cells secrete factors to enhance regeneration after implantation. Biological scaffolds derived from tissue specific extracellular matrix are highly desirable for tissue engineering and regenerative medicine because they are composed of proteins native to the implant site and provide a scaffold for regeneration. However, it is important to remove all cellular components prior to implantation to prevent immunogenicity. Most commonly used commercial decellularization start with a process to induce cell necrosis, releasing intracellular components throughout the matrix, followed by washes using harsh chemicals to remove cell remnants, which also results in undesired matrix destruction and removal of favorable bioactive factors. This project seeks to shift the paradigm in decellularization techniques by inducing apoptosis rather than necrosis. During apoptosis cells detach from the matrix and form small apoptotic bodies containing cellular components, which can be readily removed. In addition, during apoptosis cells secrete cytokines that can be sequestered in the ECM, and are known to induce proliferation, stem cell recruitment, and immunomodulation, thereby aiding in tissue regeneration. In Specific Aim 1 exploits cell apoptosis in representative tissues and organs (nerve and lung) to create novel tissue specific matrices with preserved microarchitecture and bioactive factors. Bioactivity and regenerative potential of apoptosis-optimized acellular grafts will be compared to grafts from a commercially used decellularization technique. Specific Aim 2 uses in vivo models to analyze immune response and regenerative potential of apoptosis-optimized acellular grafts versus grafts created from a commercially used decellularization technique. Results obtained are expected to contribute to the medical research community by creating a new platform for generation of acellular tissue specific matrices with enhanced regenerative potential, which could dramatically shift the future of regenerative medicine. Educational and outreach impact will be achieved through training of a graduate student and several undergraduate students, developing fun interactive lessons and labs to teach middle school students about biomaterials and biomedical engineering, developing lessons for 6th-8th grade girls at Girl?s Place, Inc. to increase young women?s interest in STEM and forging a new collaboration with the Cade Museum of Gainesville to expand lessons into full scale labs that will be run regularly to encourage creativity and invention in children.