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.

造血干细胞在后血压下驻留在骨髓内，为宿主的生命提供了所有造血谱系，并且它们在过去几十年中的广泛特征导致了临床应用，包括骨髓移植的良性和恶性疾病，影响造血室。 基于CD34抗原富集的原始区室的表达，骨髓细胞分离，分离CD34阳性种群的技术通常用于临床实践中。 真正的造血干细胞的测定需要移植和再填充，因此很难研究在人类细胞中查询这种潜力的研究。 因此，我们已经开发了一种具有强大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。 该测定法可以使我们能够在相关模型中首次在植入的人类细胞中测试球蛋白媒介。 此外，可以利用造血干细胞的离体分化沿红细胞谱系的造血干细胞在转移人β球蛋白基因后指定病毒载体的定向β-珠蛋白表达。 事实证明，这些系统可用于开发用于临床应用的病毒矢量系统，包括由前向蛋白载体开发的，该载体已由我们的组开发，并证明了10倍较高的矢量滴度以及长期重现细胞的10倍较高的转导效率。