Enhancing human iPSC-derived retinal organoids by incorporating microglia-like cells

Microglia are the primary resident immune cells in the retina. They regulate neuronal survival and synaptic pruning, which makes them essential for normal development. Following injury, they mediate adaptive responses and under pathological conditions can trigger neurodegeneration exacerbating the effect of the disease. Retinal organoids derived from human induced pluripotent stem cells (hiPSCs) are increasingly used for a range of applications, including disease modelling, development of new therapies and in the study of retinogenesis. Despite their close resemblance to the in vivo retina, they lack some key physiological features including immune cells. To enhance the retinal organoid model, we engineered an hiPSC co-culture system containing the hiPSC-derived retinal organoids and hiPSC-derived microglia-like (iMG) cells and tested their retinal invasion capacity and function. We incorporated iMG into retinal organoids at 13 weeks and tested their effect on function and development at 15 and 21.5 weeks of differentiation. Our key findings showed that iMG cells were able to respond to endotoxin challenge in monocultures and when co-cultured with the organoids. Single cell RNA-Seq transcriptomic analyses, protein expression, electron microscopy imaging and electrophysiological recordings showed that retinal organoids developed normally, expressed key markers, and retained their ability to generate spiking activity in response to light. Thus, this new co-culture immunocompetent in vitro retinal model provides a platform with greater relevance to the in vivo human retina. We engineered an hiPSC co-culture system containing the hiPSC-derived retinal organoids and hiPSC-derived microglia-like (iMG) cells and tested their retinal invasion capacity and function.

小胶质细胞（microglia）是视网膜内主要的驻留免疫细胞，可调控神经元存活与突触修剪，对视网膜正常发育至关重要。机体遭受损伤时，小胶质细胞会介导适应性免疫应答；而在病理状态下，其可诱发神经退行性变，加重疾病进程。人诱导多能干细胞（human induced pluripotent stem cells, hiPSCs）诱导生成的视网膜类器官（retinal organoids）正日益广泛应用于疾病建模、新型疗法开发以及视网膜发生机制研究等诸多领域。尽管这类视网膜类器官与体内视网膜结构高度相似，但仍缺乏包括免疫细胞在内的关键生理特征。为优化视网膜类器官模型，本研究构建了一套人诱导多能干细胞共培养体系，该体系包含人诱导多能干细胞来源的视网膜类器官与小胶质细胞样（microglia-like, iMG）细胞，并对二者的视网膜侵袭能力与功能进行了检测。研究团队于分化第13周将小胶质细胞样细胞引入视网膜类器官，并分别在分化第15周与第21.5周检测其对类器官功能与发育的影响。核心研究结果显示：小胶质细胞样细胞在单独培养与类器官共培养两种情境下，均可对内毒素（endotoxin）刺激产生应答。单细胞RNA测序（single cell RNA-Seq）转录组分析、蛋白质表达检测、电子显微镜成像以及电生理记录结果表明，视网膜类器官发育正常，可表达关键标志物，并保留了对光刺激产生电位发放的能力。综上，这套新型具备免疫活性的体外视网膜共培养模型，为研究人类体内视网膜相关机制提供了更具生理相关性的实验平台。本研究构建了一套包含人诱导多能干细胞来源视网膜类器官与小胶质细胞样细胞的共培养体系，并对其视网膜侵袭能力与功能进行了检测。