STEM CELL PRODUCTION CORE

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目及 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 骨髓间充质干细胞（Mesenchymal stem cells，MSC）是一种来源于骨髓的成体干细胞。这些细胞具有医学和治疗意义，因为它们已显示出分化成成骨细胞、脂肪细胞、软骨细胞、肌细胞、星形胶质细胞、少突胶质细胞和神经元。由于其固有的可塑性，这些细胞有可能用于治疗大量遗传疾病。邦内尔博士成功地确定了恒河猴间充质干细胞扩增和表征的要求。 干细胞生产核心设施（SCPC）专注于非人灵长类动物MSC的生成，维护和分布。我们常规地从恒河猴骨髓和脂肪组织样品中制备MSC。我们目前有一个由100多只不同年龄的恒河猴产生的MSC细胞系库，准备分发。SCPC目前不仅对基因治疗计划产生了巨大的影响，而且对TNPRC内的比较病理学等其他部门，以及杜兰健康科学中心和彭宁顿生物医学研究中心和其他研究实验室的部门和中心也产生了巨大的影响。 为合作研究项目生产恒河猴干细胞制剂。为此，我们计划使用在上一个资助期开发的技术来分离干细胞（MSC，NSPCs和ESCs），以扩大从不同年龄（胎儿，新生儿和成年人）和动物性别的动物中分离的各种干细胞类型的库。这些细胞将用于产生大型细胞库，供合作科学家在内部或外部使用。此外，我们将与有兴趣进行自体移植研究的科学家合作，从特定的动物中产生MSC银行，用于他们的研究计划。 继续对非人灵长类干细胞进行表征。我们在这些研究中的目标将是描绘在各种干细胞群的成熟和/或分化过程中发生的细胞和分子事件。该部门的科学家将继续在体外比较各种干细胞的生物学特性。将产生介导对恒河猴干细胞特异性的体外分化的测定，其将使MSC及其后代的分化和增殖最大化。体外表征的一个焦点将是所描述的所有干细胞沿着神经谱系的分化。为了完成克拉伯病的治疗，干细胞需要分化成少突胶质细胞。一个显着的努力，将朝着分化协议的开发和应用，以实现这一目标。 充分开发检测干细胞植入的检测方法。最重要的是，该分裂将在产生用于评估非人灵长类动物干细胞在鼠和非人灵长类动物模型中体内植入的测定中发挥重要作用。在上一个资助期间，该部门建立了使用人类核抗原抗体在小鼠模型中常规检测NHP细胞的技术，该技术可以轻松检测NHP细胞。该部门对BrdU进行了染色，并对EGFP进行了直接免疫荧光评估。在下一个资助期内，该部门将需要重点关注NHP组织中NHP细胞的检测，使用Y染色体原位杂交或使用单核苷酸多态性（SNP）检测细胞等技术。这将为我们提供一个机会，与遗传学和基因组库核心在兽医部门和DNA微阵列和表达核心在细菌学和寄生虫学上的SNP用于干细胞检测的应用合作，因为这些核心将开发SNP技术用于检测亲本谱系。 结合干细胞和基因转移技术用于恒河猴Krabbe病模型。 我们的目标是在体外和体内表征干细胞及其分化后代中有效基因递送和表达的要求。我们的各种载体系统的非人灵长类干细胞的基因转移效率的特点。这将使我们有机会产生标记有报告基因元件、EGFP或β-半乳糖苷酶的干细胞库。更重要的是，我们将评估的要求，提供高水平和长期的基因表达在恒河猴骨髓间充质干细胞在体外和体内移植后。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Mesenchymal stem cells or marrow stromal cells (MSCs) are a subset of adult stem cells from bone marrow. These cells are of medical and therapeutic interest because they have been shown to differentiate into osteoblasts, adipocytes, chondrocytes, myocytes, astrocytes, oligodendrocytes and neurons. Due to their inherent plasticity, these cells have the potential to be useful for the treatment of a large number of genetic diseases. Dr. Bunnell has successfully defined the requirements for the expansion and characterization of rhesus macaque MSCs. The Stem Cell Production Core Facility (SCPC) focuses on generation, maintenance and distribution of nonhuman primate MSCs. We routinely prepare MSCs from rhesus macaque bone marrow and adipose tissue samples. We presently have a bank pf MSC cell lines generated from over 100 rhesus macaques of varying age prepared and ready for distribution. The SCPC presently has a large impact not only on the Gene Therapy Program, but on other divisions such as Comparative Pathology within the TNPRC, but also Departments and Centers within the Tulane Health Sciences Center and the Pennington Biomedical Research Center and other research labs nationally. Produce rhesus macaque stem cell preparations for collaborative research projects. For this we plan to use the techniques developed over the last funding period to isolate stem cells (MSCs, NSPCs and ESCs) to expand the bank of various stem cell types isolated from animals of various ages (fetal, newborn and adults) and sexes of animals. The cells will be used to generate large cell banks for use internally or externally by collaborating scientists. Additionally, we will work with scientists interested in performing autologous transplantation studies by generating MSC banks from specific animals for their research programs. Continue the characterization of nonhuman primate stem cells. Our goal in these studies will be to delineate the cellular and molecular events that take place during maturation and/or differentiation of the various stem cell populations. Scientists in the division will continue to compare the biologic properties of the various stem cells in vitro. Assays mediating in vitro differentiation specific to rhesus stem cells that will maximize the differentiation and proliferation of MSCs and their progeny will be generated. One focus of the in vitro characterization will be the differentiation along neural lineages for all of the stem cells described. In order to accomplish the treatment of Krabbe's disease, stem cells will need to undergo differentiation into oligodendrocytes. A significant effort will be made towards the development and application of differentiation protocols to accomplish this goal. Fully develop assays for the detection of the engraftment of stem cells. Most importantly, the division will play a significant role in the generation of assays for the assessment of engraftment of nonhuman primate stem cells in vivo in both murine and nonhuman primate models. Over the last funding period, the division established technologies for the routine detection of NHP cells in murine models using antibodies to human nuclear antigen, which readily permits the detection of NHP cells. The division has in place staining for BrdU and direct immunofluorescence assessment of EGFP. Over the next funding period, the division will need to focus on the detection of NHP cells in NHP tissues, using technologies such as in situ hybridization for the Y chromosome or detection of cells using single nucleotide polymorphisms (SNPs). This will provide us an opportunity to collaborate with the Genetics and Genome Banking core in the Division of Veterinary Medicine and the DNA Microarray and Expression Core in Bacteriology and Parasitology on the application of SNPs for stem cells detection, as these cores will be developing SNP technologies for the testing of parental lineages. Combining stem cell and gene transfer technologies for use in the rhesus Krabbe's disease model. Our goal is to characterize the requirements for effective gene delivery and expression in stem cells and their differentiated progeny both in vitro and in vivo. The gene transfer efficiency of our various vector systems for nonhuman primate stem cells will be characterized. This will permit us the opportunity to generate banks of stem cells labeled with reporter gene elements, EGFP or beta-galactosidase. More importantly, we will assess the requirements that provide for high levels and protracted duration of gene expression in rhesus MSCs both in vitro and in vivo following transplantation.