Differentiation of Human ES and iPS Cells into Lens Cells

描述（由申请人提供）：该项目的长期目标是开发新的模型，通过胚胎干细胞（ES）产生的晶状体细胞来研究人类年龄性白内障。这些模型采用分化为晶状体细胞的患者源性诱导多能干细胞（iPS）来研究晶状体细胞在已知诱导人类白内障发生条件下的稳态。本R21应用程序的直接目标是在确定的细胞培养条件下，进行系统的探索性研究，从人胚胎干细胞（ES）细胞中产生晶状体细胞。与年龄有关的白内障是一种晶状体疾病，全世界近一半的失明是由这种疾病造成的，随着发达国家和不发达国家预期寿命的延长，该病的发病率预计还会增加。老年性白内障发生于40岁以后，是晶状体微结构逐渐破坏的结果。老年性白内障是一种复杂的疾病，涉及遗传和环境因素。晶状体混浊被认为是由于暴露在UV-B、低抗氧化剂摄入和吸烟等环境对晶状体蛋白质和其他结构成分的累积损害造成的。由于缺乏适当的动物模型，以及原代晶状体细胞和器官培养的使用有限，阻碍了系统研究人类白内障的方法。我们现在已经确定了一个实验方案，以产生大量的晶状体细胞祖细胞分化从人类胚胎干细胞系，H1。该提案将(1)建立一个优化的实验方案，以从人类胚胎干细胞中产生高度富集的晶状体祖细胞群体；(2)建立条件，以产生分化的透镜状体，并对其进行表征；(3)将使用6个具有代表性的iPS细胞系对这些实验条件进行测试。这些数据将为利用来自不同白内障患者的iPS细胞生成晶状体细胞奠定基础。患者源性晶状体细胞的数量将足以用于特定抗氧化防御机制的生化研究，以确定在白内障发生中起关键作用的晶状体保护机制。通过确定药物开发的分子靶标，这种新方法将激发对药物的理性探索，以减少衰老对晶状体的影响。总的来说，这些探索性研究将用于R01项目，利用遗传和表型特征的生物材料来模拟人类白内障。

DESCRIPTION (provided by applicant): The long-term goal of this program is to develop new models to study human age-onset cataract via lens cells generated from embryonic stem (ES) cells. These models employ patient-derived induced pluripotent stem (iPS) cells differentiated into lens cells to study lens-cell homeostasis under conditions known to induce human cataractogenesis. The immediate goal of this R21 application is to perform systematic exploratory studies to generate lens cells form human embryonic stem (ES) cells using defined cell culture conditions. Age-related cataract is a disease of the lens that is responsible for nearly half of blindness worldwide, and is expected to increase as a result of increased life expectancy in both developed and underdeveloped countries. Age-onset cataracts develop after the age of 40 as a result of the progressive breakdown of the lens microarchitecture. Age-onset cataract is a complex disease involving both genetic and environmental factors. Lens opacities are thought to originate from the cumulative damage of environmental insults on lens proteins and other structural components of the lens from UV-B exposure, low antioxidant intake, and cigarette smoking. A systemic approach to study human cataract is hampered both by the lack of appropriate animal models, and limited use of primary lens cell and organ cultures. We have now identified an experimental protocol to generate large quantities of lens cells progenitor cells differentiated from human ES cell line, H1. This proposal will (1) establish an optimized experimental protocol to generate a highly enriched population of lens progenitor cells from human ES cells, (2) will establish conditions to generate differentiated lentoid bodies followed by their characterization, and (3) will test these experimental conditions using a representative pool of six iPS cell lines. These data will lay foundation for generation of lens cells using iPS cells from distinct cataract patients. The patient-derived lens cells will be available at quantities sufficient for biochemical studies of specific antioxidant defensive mechanisms to identify those lens protective mechanisms that play key roles in cataractogenesis. By identifying molecular targets for drugs to exploit, this novel approach will inspire a rational search for drugs to reduce the effects of aging on the lens. Collectively, these exploratory studies will be used for R01 projects to model human cataract using genetically and phenotypically characterized biological materials. PUBLIC HEALTH RELEVANCE: This application is relevant to human health as lens cataract is a major cause of worldwide blindness. Age- related cataract develops at some time after 40 years of age as a result of progressive breakdown of the ocular lens structure. Current treatment for senile cataract generally consists of surgery that replaces the opaque lens with an artificial intraocular lens. Although the surgery is performed routinely in the US at a rate of 1.8-2 million patients per annum, it represents a major Medicare reimbursement category. It has been estimated by the National Eye Institute, NIH (Bethesda, MD) that a 10-year delay in the onset of cataracts, could decrease the number of surgeries needed by almost one half, thus significantly decreasing vision care costs.