Mesenchymal Stem Cells can Restore and Maintain Corneal Endothelial Function

项目摘要 在本项目中，我们建议研究间充质干细胞(MSCs)分化为 功能活跃的角膜内皮细胞(CECs)将被移植到患病的眼睛。作为MSCs的来源， 骨髓MSCs(BM-MSCs)、角膜缘MSCs(L-MSCs)和脂肪来源MSCs(A-MSCs)将 已评估。我们将(1)通过改变众多参数来确定不同MSCs的分化潜能 并评估参与它们分化的不同分子途径，以及(2)确定 分化的细胞在疾病条件下再生功能活跃的内皮细胞。差异化， 在此通过体外、体外和体内进行的表征、生物相容性和功能化研究。 活体研究将提供生物整合和再生程度的定性和定量信息 分化的细胞与宿主S角膜细胞和细胞外基质接触的潜力。我们预计 从MSCs分化而来的CECs在移植后可以在体内增殖，保持最佳的水化状态 角膜基质。自体或异体间充质干细胞能否分化为CEC并移植到 内皮疾病患者恢复正常的内皮功能，不会引起任何免疫反应 或炎症反应，而不使用整个供体角膜(DC)，这可以简化治疗 角膜内皮疾病和增加树突状细胞可用于其他类型的角膜移植。因此，我认为，这 拟议的研究计划有可能给角膜疾病的治疗带来革命性的变化，而不仅仅是 不仅影响角膜内皮，还影响其他角膜层。研究目标得到了支持。 培训计划的重点是获得细胞和细胞领域的相关多学科专门知识 分子生物学、生理学、组织工程学、基因疗法和材料科学。为此，一个 “导师团队”包括首席导师James Chodosh博士(MEE/SERI)和共同导师Dr。 Miguel Gonzalez(SERI/UOC)和Shigeto Shimura教授(Keidai)已组装完毕。此外，还有三个 独立合作者将在(1)与MSCs分化相关的研究设计方面为我们的团队提供支持 和特征(加尔松博士)，(2)对参与分化的分子途径的理解(加尔松博士) Sabater)，以及(3)基于纳米颗粒的MRI可检测治疗策略设计(Patra博士)。这群人 导师和合作者将在K99阶段指导我实现我的长期职业目标，成为 一位学术科学家，在我的领域中处于领先地位，具有很强的独立研究背景。我预见到这一点 大奖将为我提供一个很好的平台来过渡到独立研究教员(这是 包括过渡到R00阶段)，从长远来看，成功竞争独立的NIH 资助(R01赠款)。在指导期间，我的培训还将得到专业化课程的补充 发展、改进陈述和写作技能的阶段(赠款和手稿)、手稿 回顾、指导和发展领导能力和实验室管理技能。

Project Summary In this project, we propose to investigate the ability of mesenchymal stem cell (MSCs) to differentiate into functionally active corneal endothelial cells (CECs) to be transplanted in the diseased eye. As sources of MSCs, bone marrow MSCs (BM-MSCs), limbus MSCs (L-MSCs), and adipose derived MSCs (A-MSCs) will be evaluated. We will (1) determine the differentiation potential of different MSCs by varying numerous parameters and evaluate different molecular pathways involved in their differentiation, and (2) determine the ability of the differentiated cells to regenerate functionally active endothelium in diseased conditions. The differentiation, characterization, biocompatibility, and functionalization studies proposed here through in vitro, ex vivo, and in vivo studies will offer qualitative and quantitative information of the degree of biointegration and regenerative potential of the differentiated cells in contact with the host´s corneal cells and extracellular matrix. We expect that CECs differentiated from MSCs can proliferate in vivo after transplantation, maintaining the optimal hydration of the corneal stroma. If autologous or allogeneic MSCs can be differentiated to CECs and transplanted to recover normal endothelial function in patients suffering from endothelial diseases, without causing any immune or inflammatory response and without using whole donor corneas (DCs), this could simplify the treatment of corneal endothelial diseases and increase availability of DCs for other types of keratoplasty. Thus, I believe, this proposed research plan has the potential to revolutionize the treatment of corneal diseases, not only those affecting the corneal endothelium but also those affecting other corneal layers. The research aims are supported by the training plan focused on the acquisition of relevant multidisciplinary expertise in the field of cell and molecular biology, physiopathology, tissue-engineering, gene therapy, and material science. To this end, a “Mentoring Team” which includes the lead mentor, Dr. James Chodosh (MEE/ SERI), along with co-mentors, Dr. Miguel Gonzalez (SERI/UOC) and Professor Shigeto Shimmura (Keidai), has been assembled. Moreover, three independent collaborators will support our team in (1) the design of the studies related to MSCs differentiation and characterization (Dr. Garzon), (2) understanding of molecular pathways involved in the differentiation (Dr. Sabater), and (3) the nanoparticle based MRI detectable therapeutic strategies design (Dr. Patra). This group of mentors and collaborators will guide me during the K99 phase to achieve my long term career goal of becoming an academic scientist, and leader in my field with a strong independent research background. I envision that this award will provide me with an excellent platform to transition to an independent research faculty member (this includes the transition into the R00 phase) and in the long term, to successfully compete for independent NIH funding (R01 grant). My training will additionally be complemented by specialization courses during the mentored phase in the development, improvement of presentation and writing skills (grants and manuscripts), manuscript review, mentoring, and development of leadership abilities and lab management skills.