Generation of retinal pigmented epithelium from iPSCs derived from the conjunctiva of donors with and without age related macular degeneration

Fidelity in pluripotent stem cell differentiation protocols is necessary for the therapeutic and commercial use of cells derived from embryonic and induced pluripotent stem cells. Recent advances in stem cell technology, especially the widespread availability of a range of chemically defined media, substrates and differentiation components, now allow the design and implementation of fully defined derivation and differentiation protocols intended for replication across multiple research and manufacturing locations. In this report we present an application of these criteria to the generation of retinal pigmented epithelium from iPSCs derived from the conjunctiva of donors with and without age related macular degeneration. Primary conjunctival cells from human donors aged 70–85 years were reprogrammed to derive multiple iPSC lines that were differentiated into functional RPE using a rapid and defined differentiation protocol. The combination of defined iPSC derivation and culture with a defined RPE differentiation protocol, reproducibly generated functional RPE from each donor without requiring protocol adjustments for each individual. This successful validation of a standardized, iPSC derivation and RPE differentiation process demonstrates a practical approach for applications requiring the cost-effective generation of RPE from multiple individuals such as drug testing, population studies or for therapies requiring patient-specific RPE derivations. In addition, conjunctival cells are identified as a practical source of somatic cells for deriving iPSCs from elderly individuals.

多能干细胞分化方案的保真度，是胚胎多能干细胞与诱导多能干细胞来源细胞实现治疗性与商业化应用的必要前提。近年来干细胞技术取得长足进展，尤其是一系列化学成分明确的培养基、细胞培养基质与分化组分的广泛可得，使得我们能够设计并实施完全标准化的诱导与分化方案，以支持多研究机构与生产场地间的稳定重现。本研究报告中，我们将上述标准应用于从伴或不伴年龄相关性黄斑变性的供体结膜组织来源的诱导多能干细胞（induced pluripotent stem cells，iPSCs）中诱导生成视网膜色素上皮（retinal pigmented epithelium，RPE）。我们对70至85岁人类供体的原代结膜细胞进行重编程，获得多株iPSC细胞系，并通过一套快速且标准化的分化方案将其诱导为具有功能的RPE。将标准化的iPSC诱导与培养流程，与标准化的RPE分化方案相结合，可从每位供体中稳定获得具有功能的RPE，无需针对个体调整实验方案。本次对标准化iPSC诱导与RPE分化流程的成功验证，为需要从多名供体中低成本获取RPE的应用场景提供了切实可行的方案，例如药物测试、群体研究，或是需要制备患者特异性RPE的治疗应用。此外，本研究证实结膜细胞可作为从老年个体中获取iPSCs的实用体细胞来源。