Corneal Endothelial Cell Transplantation

描述(申请人提供)：角膜内皮细胞功能障碍导致视力下降，只能通过角膜移植来取代体内不能增殖的人角膜内皮细胞(HCECs)。在美国，每年因内皮细胞功能障碍而进行的角膜移植手术超过15,000例，但随着组织需求的增加和组织供应的威胁，供体组织危机迫在眉睫。为了满足未来的需求，供体池可以通过移植培养的内皮细胞来扩大，但要想成功地进行细胞移植，还需要新的方法将培养的细胞输送到眼睛。我们的研究计划的目标是开发一种不需要在膜性细胞载体上转移细胞的方法，将HCEC移植到角膜后基质中。我们推测，如果细胞与角膜基质直接接触，将有助于将HCECs移植到角膜后基质中。在初步研究中，我们成功地将超顺磁性微球(SPM)掺入体外培养的人脐静脉内皮细胞中，并证明了细胞向磁源迁移。我们将通过施加外加磁场，将含有SPM的HCECs定位到角膜基质上，从而消除对膜细胞载体的需求。这种方法的主要好处将是急需扩大供体池，但此外，拟议的技术在体内应用于人类时将是微创和无缝合的，促进细胞直接附着到角膜基质将使疾病的治疗成为可能，例如福克斯内皮营养不良症，除了内皮细胞外，还需要去除Descemet膜。我们将通过单层培养人脐静脉内皮细胞，并将其移植到体外人眼前节灌流器官培养系统中的人角膜上来验证我们的假设。从这些研究中获得的数据将成为未来在动物身上进行内皮细胞移植研究的基础，最终将在人类身上进行。为了验证我们的假设，这项提议将研究三个特定的目标：“确定SPM的类型、大小和浓度对体外培养的人脐静脉内皮细胞的活力、磁引力和透光率的影响。”确定将含有SPM的悬浮培养的人脐静脉内皮细胞移植到人眼前节灌流器官培养模型中的解剖学和生理学结果。“确定HCEC附着在角膜基质上的机制。与公共健康相关在这项提案中，我们将优化一种移植人角膜内皮细胞的新技术，这可能会成为目前需要角膜组织移植的许多疾病的治疗方法。内皮细胞移植的主要优势将是减少甚至可能消除角膜供体组织短缺，因为移植的细胞可以在实验室培养并用于治疗多名患者，而不是只治疗一名患者。

DESCRIPTION (provided by applicant): Corneal endothelial cell dysfunction causes decreased vision and can only be treated by corneal transplantation to replace human corneal endothelial cells (HCECs), which do not proliferate in vivo. Over 15,000 corneal transplants are performed every year in the United States for endothelial cell dysfunction, but with increasing tissue demand and threats to the tissue supply, a donor tissue crisis is imminent. To meet future demand, the donor pool can be expanded by transplanting cultured endothelial cells, but novel methods of delivering cultured cells to the eye are needed for cell transplantation to be successful. The goal of our research program is to develop a method of HCEC transplantation to posterior corneal stroma without transferring cells on a membranous cell carrier. We hypothesize that transplantation of HCECs to posterior corneal stroma will be facilitated if cells are drawn into direct contact with the stroma. In preliminary studies, we have successfully incorporated superparamagnetic microspheres (SPMs) into cultured HCECs in vitro and demonstrated cell migration toward a magnetic source. We will localize HCECs with incorporated SPMs to the corneal stroma by applying an external magnetic field, eliminating the need for a membranous cell carrier. The major benefit of this approach will be the much needed expansion of the donor pool, but in addition, the proposed technique will be minimally invasive and sutureless when applied to humans in vivo, and promoting cell attachment directly to corneal stroma will enable treatment of diseases, such as Fuchs' endothelial dystrophy, that require removal of Descemet's membrane in addition to endothelial cells. We will investigate our hypothesis by culturing HCECs in monolayer and transplanting them to human corneas in a perfusion organ culture system of human anterior segments ex vivo. The data obtained from these studies will become the foundation for future endothelial cell transplantation studies in animals, and ultimately in humans. Three specific aims will be investigated in this proposal to test our hypothesis: " Determine the effect of the type, size, and concentration of SPMs on the viability, magnetic attraction, and light transmittance of cultured HCECs in vitro. " Determine the anatomical and physiological outcomes of transferring suspended cultured HCECs with incorporated SPMs to the human perfusion organ culture model of anterior segments. " Determine the mechanism of HCEC attachment to corneal stroma. PUBLIC HEALTH RELEVANCE In this proposal, we will optimize a novel technique of transplanting human corneal endothelial cells, which might become a treatment for many diseases that presently require a corneal tissue transplant. The major advantage of endothelial cell transplantation will be to reduce, and possibly eliminate, corneal donor tissue shortages because the transplanted cells can be grown in a laboratory and used to treat several patients instead of only one patient.