Transplantation of Genetically Modified, Immune-privileged Steroli Cells

描述（由申请人提供）：支持细胞是免疫特权细胞，在跨异体或异种屏障移植时，具有长期存活能力，无需使用免疫抑制药物。当大多数其他细胞被免疫排斥时，Sertoli细胞在移植中存活的能力表明，Sertoli细胞可以被设计为基因治疗的载体。我研究的长期目标是利用基因工程、具有免疫特权的Sertoli细胞作为无限来源的组织，可以作为一种载体，传递治疗相关的蛋白质，作为治疗多种疾病的一种手段。因此，本应用的目的是评估使用Sertoli细胞产生功能水平的胰岛素作为糖尿病小鼠有效的长期治疗策略的可行性。假设具有免疫特权的Sertoli细胞可以通过基因工程稳定表达基础水平的胰岛素，并且这些细胞可以作为异体或异种移植在糖尿病小鼠中存活而不需要免疫抑制。这一假设是基于我们的初步数据，这些数据表明未修饰的Sertoli细胞在不使用免疫抑制药物的情况下作为同种异体移植和异种移植物移植时长期存活，并且稳定表达GFP的Sertoli细胞在同种异体移植中存活，同时继续表达GFP。具体目标是为了验证这一假设。在Aim 1中，胰岛素表达的支持细胞的功能将被检查。小鼠和猪稳定表达胰岛素的支持细胞将在体外和体内移植到糖尿病免疫缺陷SCID小鼠体内后，检测胰岛素的产生和分泌功能水平。在Aim 2中，这些表达胰岛素的Sertoli细胞在跨越免疫屏障移植后存活（保持免疫特权）的能力将被检查。表达胰岛素的支持细胞将作为同种异体移植（小鼠支持细胞）或异种移植（猪支持细胞）移植到具有免疫能力的糖尿病C3H小鼠体内。功能性胰岛素的产生和支持细胞的存活将被检查。预计经过工程改造以稳定传递基础水平胰岛素的支持细胞能够在移植到糖尿病小鼠体内后长期降低血糖水平。这一贡献意义重大，因为它将提供证据，证明基因工程、具有免疫特权的Sertoli细胞可以作为移植组织的无限供应。

DESCRIPTION (provided by applicant): Sertoli cells are immune-privileged cells that have the ability to survive long-term without the use of immunosuppressive drugs when transplanted across allogeneic or xenogeneic barriers. The ability of Sertoli cells to survive transplantation when most other cells are immunologically rejected suggests Sertoli cells could be engineered as a vehicle for gene therapy. The long-term goal of my research is to use genetically engineered, immune-privileged Sertoli cells as an unlimited source of tissue that can be used as a vehicle to deliver therapeutically relevant proteins as a means to treat multiple disorders. Therefore, the objective of this application is to evaluate the feasibility of using Sertoli cells engineered to produce functional levels of insulin as an effective long-term, therapeutic strategy in diabetic mice. It is hypothesized that immune- privileged Sertoli cells can be genetically engineered to stably express basal levels of insulin, and that these cells can survive transplantation as allografts or xenografts in diabetic mice without the need for immunosuppression. This hypothesis is based on our preliminary data, which demonstrated unmodified Sertoli cells survive long-term when transplanted as allografts and xenografts without the use of immunosuppressive drugs and that Sertoli cells stably expressing GFP survive allogeneic transplantation while continuing to express GFP. The specific aims are designed to test this hypothesis. In Aim 1, the function of insulin expressing Sertoli cells will be examined. Mouse and porcine Sertoli cells stably expressing insulin will be examined for production and secretion of functional levels of insulin in vitro and after transplantation into diabetic immunodeficient SCID mice in vivo. In Aim 2, the ability of these insulin-expressing Sertoli cells to survive (remain immune-privileged) when transplanted across immunological barriers will be examined. Insulin-expressing Sertoli cells will be transplanted into diabetic, immune-competent C3H mice as allografts (mouse Sertoli cells) or xenografts (porcine Sertoli cells). The production of functional insulin and Sertoli cell survival will be examined. It is expected that Sertoli cells engineered to stably deliver basal levels of insulin are capable of lowering blood glucose levels long-term after transplantation into diabetic mice. This contribution is significant because it will provide evidence that genetically engineered, immune-privileged Sertoli cells can be used as an unlimited supply of tissue for transplantation. PUBLIC HEALTH RELEVANCE: There are many disorders that have the potential to be treated by gene therapy; for example, delivery of factor VIII for the treatment of hemophilia. However, an efficient and safe delivery system has not yet been developed. We will explore the novel and innovative concept that immune-privileged Sertoli cells can be engineered to deliver therapeutic proteins. In the long-term these cells could provide an unlimited supply of genetically modified tissue that could be transplanted without the use of chronic immunosuppression as a means of treatment for multiple disorders.