Enhancing Bcl-2 Expression for Bone Regeneration.

增强 Bcl-2 表达促进骨再生。

基本信息

批准号：
8676512
负责人：
MICHAEL T LONGAKER
金额：
$ 24.16万
依托单位：
STANFORD UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-05-22 至 2016-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8676512
关键词：
Address Adipose tissue Apoptosis Apoptosis Regulator BCL2 gene Back Bacterial DNA Bone Regeneration Bone Transplantation Calvaria Caspase Cell Proliferation Cell Survival Cell Transplants Cells Complex DNA DNA Sequence Defect Development Encapsulated Endocytosis Ensure Environment Etiology Excision Fluorescence-Activated Cell Sorting Gelatin Gene Expression Genes Glycolates HA coating Harvest Healed Hour Human Hydrogels Hypoxia Image Immunocompromised Host Implant In Situ In Vitro Inflammation Inflammation Mediators Investigation Jet Injections Light Luciferases Malignant Neoplasms Mediating Mediator of activation protein Mesenchymal Modality Morbidity - disease rate Mus Pathway interactions Patients Process Proteins Regenerative Medicine Replication Origin Reporter Research Personnel Risk Site Staurosporine Stem cells Stromal Cells Techniques Therapeutic Tissues Transcript Transfection Trauma Up-Regulation X-Ray Computed Tomography angiogenesis base bone clinical application congenital anomaly craniofacial gene therapy healing implantation improved in vivo innovation iron oxide nanoparticle novel strategies public health relevance reconstruction regenerative repaired research study response restoration scaffold skeletal tissue regeneration tissue repair tumor tumorigenic vector wound

项目摘要

DESCRIPTION (provided by applicant): Reconstruction of craniofacial skeletal defects from both congenital and acquired etiologies can often present a daunting challenge. Contemporary strategies employ bone grafting and free tissue transfer, but donor site morbidity, as well as suboptimal restoration of form and function, continues to drive the development of novel approaches. The field of regenerative medicine holds significant promise to address this need, employing cellular-based strategies to replace damaged or deficient tissues. Despite tremendous advances, however, several complex issues remain. Clinical application of stem cells often involve placement into hostile wounds that are hypoxic and possess dramatically upregulated inflammatory mediators. Within such a harsh environment, cellular survival is uncertain, and the utility of implanted cells can be severely compromised. In light of this, there s growing recognition that simple placement of stem cells into a wound is not sufficient to ensure maximal tissue regeneration. While investigators have focused on promoting angiogenesis or limiting inflammation, a novel approach would be to enhance pro-survival pathways within the transplanted cells themselves through upregulation of Bcl-2, a regulator of apoptosis, for bone regeneration. Importantly, increased expression of Bcl-2 has been shown to reduce apoptosis without limiting subsequent differentiation capacity in mesenchymal cells. However, limited expression of Bcl-2 is necessary to avoid any tumorigenic risk. We will therefore employ a non-integrating, non-viral minicircle vector to transfect human adipose-derived stromal cells and evaluate their subsequent viability and bone forming capacity. Furthermore, to maximize the translational potential of our approach, we will develop an innovative scaffold incorporating polyethylenimine- encapsulated superparamagnetic iron oxide nanoparticles complexed to our Bcl-2 expressing minicircle. This will allow for an "off the shelf" approach where freshly harvested cells can be directly seeded onto scaffolds and placed back into the patient for in situ transfection. We will evaluate the ability for this novel strategy to enhance survival and bone regenerative ability of human adipose-derived stromal cells implanted into non- healing critical-sized calvarial defects in immunocompromised mice. Collectively, the experiments proposed will determine the ability for Bcl-2 upregulation to promote bone repair and facilitate development of new clinical applications to repair large skeletal defects.

描述（由申请人提供）：先天性和后天性病因导致的颅面骨骼缺陷的重建通常会带来艰巨的挑战。当代策略采用骨移植和游离组织移植，但供体部位发病率以及形态和功能恢复不理想，继续推动新方法的发展。再生医学领域有望解决这一需求，采用基于细胞的策略来替代受损或缺陷的组织。然而，尽管取得了巨大进步，但仍然存在一些复杂的问题。干细胞的临床应用通常涉及将其置于缺氧且炎症介质显着上调的敌对伤口中。在如此恶劣的环境中，细胞的存活是不确定的，植入细胞的效用可能会受到严重影响。有鉴于此，人们越来越认识到，简单地将干细胞放入伤口中不足以确保最大限度的组织再生。虽然研究人员专注于促进血管生成或限制炎症，但一种新方法是通过上调 Bcl-2（细胞凋亡调节剂）来增强移植细胞本身的促生存途径，以促进骨再生。重要的是，Bcl-2 表达的增加已被证明可以减少细胞凋亡，而不限制间充质细胞随后的分化能力。然而，Bcl-2 的有限表达对于避免任何致瘤风险是必要的。因此，我们将采用非整合、非病毒小环载体转染人脂肪源性基质细胞，并评估其随后的活力和骨形成能力。此外，为了最大限度地发挥我们方法的转化潜力，我们将开发一种创新支架，将聚乙烯亚胺封装的超顺磁性氧化铁纳米颗粒与我们的 Bcl-2 表达小环复合。这将允许采用“现成的”方法，将新鲜收获的细胞直接接种到支架上并放回患者体内进行原位转染。我们将评估这种新策略的能力，以提高植入免疫功能低下小鼠的非愈合临界尺寸颅骨缺损中的人类脂肪源性基质细胞的存活率和骨再生能力。总的来说，所提出的实验将确定 Bcl-2 上调促进骨修复的能力，并促进修复大型骨骼缺陷的新临床应用的开发。