Isolation, characterization, and transplantation of candidate stem cells

候选干细胞的分离、表征和移植

• 批准号: 9157366
• 负责人: John Tisdale
• 金额: $ 56.41万
• 依托单位: NATIONAL HEART, LUNG, AND BLOOD INSTITUTE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/9157366
• 关键词: Affect; Benign; Biological Assay; Blood; Bone Marrow; Bone Marrow Cells; Bone Marrow Transplantation; CD34 Antigens; CD34 gene; Cell Lineage; Cells; Clinic; Disease; Engraftment; Erythroid; Genes; Globin; Hematopoiesis; Hematopoietic; Hematopoietic stem cells; Hereditary Disease; Human; Knowledge; Life; Malignant - descriptor; Methodology; Modeling; Natural regeneration; Organ; Population; Source; Speed; Stem cells; Stress; System; Techniques; Testing; Time; Transplantation; Viral Vector; Xenograft Model; base; beta Globin; clinical application; clinical practice; human disease; insight; repaired; transduction efficiency; vector

Hematopoietic stem cells reside within the bone marrow post-natally, providing all hematopoietic lineages for the life of the host, and their extensive characterization over the last several decades has led to clinical application including bone marrow transplantation for benign and malignant disorders affecting the hematopoietic compartment. Isolation of bone marrow cells based upon expression of the CD34 antigen enriches for the primitive compartment, and techniques to isolate the CD34-positive population are commonly used in clinical practice. Assays for true hematopoietic stem cells require transplantation and repopulation, and thus studies querying such potential among human cells are difficult. We have thus developed a xenograft model with robust human hematopoietic stem cell engraftment, and have demonstrated engraftment of all cell lineages. This assay will allow us to test globin vectors for the first time among engrafted human cells in a relevant model. Furthermore, ex vivo differentiation of hematopoietic stem cells along the erythroid lineage can be utilized to assay viral vector directed beta-globin expression after transfer of the human beta globin gene. These systems have proven useful in developing viral vector systems for clinical application including a forward oriented globin vector which has been developed by our group and demonstrates 10 fold higher vector titer as well as 10 fold higher transduction efficiency for long term repopulating cells.

造血干细胞出生后存在于骨髓中，为宿主的一生提供所有造血谱系，过去几十年来，它们的广泛表征已导致临床应用，包括骨髓移植治疗影响造血室的良性和恶性疾病。 基于 CD34 抗原表达的骨髓细胞分离可富集原始区室，并且分离 CD34 阳性群体的技术在临床实践中常用。 对真正的造血干细胞的分析需要移植和再增殖，因此探究人类细胞的这种潜力的研究很困难。 因此，我们开发了具有强大的人类造血干细胞植入的异种移植模型，并证明了所有细胞谱系的植入。 该测定将使我们能够首次在相关模型中的移植人类细胞中测试珠蛋白载体。 此外，造血干细胞沿红系谱系的离体分化可用于测定转移人β珠蛋白基因后病毒载体定向的β-珠蛋白表达。 这些系统已被证明可用于开发用于临床应用的病毒载体系统，包括由我们小组开发的正向珠蛋白载体，并表现出高10倍的载体滴度以及高10倍的长期再增殖细胞的转导效率。