Corneal Endothelial Cell Transplantation

描述（由申请人提供）：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替代人角膜内皮细胞（HCEC）进行治疗，HCEC在体内不会增殖。在美国，每年有超过15，000例角膜移植用于内皮细胞功能障碍，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，可以通过移植培养的内皮细胞来扩大供体库，但是需要将培养的细胞输送到眼睛的新方法才能使细胞移植成功。本研究的目的是建立一种不需要将细胞转移到膜状细胞载体上的角膜内皮细胞移植到角膜后基质的方法。我们推测，如果细胞与基质直接接触，将促进HCEC移植到角膜后基质。在初步研究中，我们已经成功地将超顺磁性微球（SPM）到体外培养的HCECs，并证明了细胞向磁源迁移。我们将通过施加外部磁场将具有整合的SPM的HCEC定位于角膜基质，从而消除对膜细胞载体的需要。这种方法的主要益处将是急需的供体库的扩增，但此外，所提出的技术在体内应用于人类时将是微创的和无缝合的，并且促进细胞直接附着于角膜基质将使得能够治疗除了内皮细胞之外还需要去除后弹力膜的疾病，例如Fuchs内皮营养不良。我们将探讨我们的假设，培养HCECs在单层和移植到人角膜的灌注器官培养系统的人眼前节离体。从这些研究中获得的数据将成为未来动物内皮细胞移植研究的基础，并最终在人类。本提案将研究三个具体目标以验证我们的假设：“确定SPM的类型、大小和浓度对体外培养的HCEC的活力、磁吸引力和透光率的影响。“确定将悬浮培养的HCECs与结合的SPM转移到前段的人类灌注器官培养模型的解剖学和生理学结果。“确定HCEC附着于角膜基质的机制。在这项提案中，我们将优化一种新的移植人类角膜内皮细胞的技术，这可能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点将是减少并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中生长并用于治疗多个患者而不是仅一个患者。

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.